Ovulation Induction Surveen Ghumman
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Obstetric Vasculopathies
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Anovulation and Tests for Ovulationchapter 1

Surveen Ghumman,
Ritika Kaur
Disorders of ovulation account for approximately 30–40% of the problems identified in infertile women. They may present with oligomenorrhea or amenorrhea. Classification of ovulatory problems was done by WHO into 3 groups:
• Group I
Hypothalamic pituitary failure (Hypogonadotropic hypogonadism).
• Group II
Hypothalamic pituitary dysfunction (Normogonadotropic, e.g. PCOD).
• Group III
Ovarian failure (Hypergonadotropic hypogonadism).
Usually treatment is simple and effective. However, not all cases of anovulation are amenable to treatment by ovulation induction.1 It is the cause of anovulation that will determine whether ovulation induction is possible (Tables 1.1 and 1.2).
Before taking up a patient for ovulation induction a complete investigation for confirmation and cause of 2anovulation should be carried out (Table 1.3).
Table 1.1   Causes of anovulation suitable for ovulation induction treatment1
Hypothalamic
Low concentration of gonadotropin-releasing hormone (hypogonadism)
Weight or exercise related amenorrhea
Kallmann syndrome
Stress
Idiopathic
Pituitary
Hyperprolactinemia
Pituitary failure (hypogonadotropic hypogonadism)
Sheehan's syndrome
Craniopharyngioma or hypophysectomy
Cerebral radiotherapy
Ovarian
Polycystic ovaries
Other endocrine
Hypothyroidism
Congenital adrenal hyperplasia
Table 1.2   Causes of anovulation not suitable for ovulation induction treatment1
Ovarian failure
Idiopathic
Radiotherapy or chemotherapy
Surgical removal
Genetic
Autoimmune
Chromosomal
Turner's syndrome (45,X)
3
This is necessary to determine which ovulation inducing drugs are to be used, in how much dose and whether any adjuvants are needed.
Table 1.3   Investigations for anovulation1
Investigation
When done
Interpretation
Progesterone
Midluteal phase of cycle (day 21 of 28 day)
>3 ng/mL—confirms ovulation
> 10 ng/mL—shows adequate luteal phase
Follicle-stimulating hormone
Early follicular phase
>10 IU/L indicates reduced ovarian reserve
> 40 IU/L indicates ovarian failure
< 5 IU/L may indicate pituitary or hypothalamic problem
Luteinizing hormone
Early follicular phase
> 10 IU/L indicates polycystic ovaries
< 5 IU/L may indicate pituitary or hypothalamic problem
Testosterone
Any time in cycle
>2.4 nmol/L indicates polycystic ovaries
> 5 nmol/L suggests congenital adrenal
hyperplasia; check DHEAS and 17-OHP
Prolactin
Any time in cycle (but not after exercise or stress)
>100 ng/mL indicates pituitary adenoma
< 100 ng/mL—other causes of hyperprolactinemia
Thyroid-stimulating hormone
Any time in cycle if woman has symptoms or signs of hypothyroidism or has hyperprolactinemia
High thyroid stimulating hormone indicates hypothyroidism4
Transvaginal ultrasound scan
Day 2—baseline scan
Day 9 onward—follicular monitoring
Identifies polycystic ovaries
Ovulation documentation
MRI/CT of pituitary
If two prolactin levels >100 ng/mL
Identifies macroadenomas
Karyotype
Primary amenorrhea and premature menopause
Identifies karyotypic abnormalities—for example, Turner's syndrome (45, X) translocations, and androgen insensitivity syndrome (46, XY)
Body mass index
Oligomenorrhea or amenorrhea
BMI > 30 suggests polycystic ovary syndrome BMI < 20 suggests hypogonadotropic hypogonadism
 
Tests for Ovulation
Clinically ovulation is indicated by regular menstrual cycles, midcycle pain and changes in cervical mucus. There are many methods of confirming ovulation. They are all based on the effects of hormonal events taking place in the body during ovulation. These tests are also used to assess the effectiveness of any ovulation induction treatment.
 
Basal Body Temperature (BBT)
It is body temperature under basal conditions at rest. For practical purposes BBT is measured each morning before arising from bed with an oral glass thermometer, having an expanded scale, typically ranging from 96°F to 100°F and marked in tenths of a degree. Irregular sleep patterns and smoking can interfere with tests results.5
BBT recordings are based on the thermogenic properties of progesterone. As levels rise after ovulation, BBT also increases. BBT varies between 97°F and 98°F during the follicular phase of the cycle. The thermogenic shift in BBT occurs when progesterone concentrations rise above approximately 5 ng/mL, 1 to 5 days after LH surge and up to 4 days after ovulation.2 The temperature rise is usually abrupt but may be gradual and difficult to define. BBT generally falls to its lowest level on the day before ovulation, but the nadir in BBT cannot be reliably identified until after the temperature rises and remains elevated.3 It then increases by 0.4 – 0.8° over the average preovulatory temperature during the luteal phase and falls again to baseline levels just before or after the onset of menses. A biphasic pattern usually is readily evident. A normal luteal phase documents temperature elevation for 11 days at least. Menses begin 12 days or more after the rise in temperature. In pregnancy BBT remains elevated because of the sustained production of progesterone by the corpus luteum stimulated by human chorionic gonadotropin.
 
Advantages
  1. Relatively low cost.
  2. BBT recordings can also reveal an abnormally long follicular phase or short luteal phase.
 
Disadvantages
  1. Increases stress.
  2. Women may menstruate regularly and predictably but do not exhibit a clearly biphasic BBT pattern.
  3. The most fertile period passes once the rise in temperature is seen. BBT tracings are useful when recordings are viewed in retrospect to show ovulatory pathology.
6
 
Luteal Serum Progesterone Levels
Progesterone levels generally remain below 1 ng/mL during the follicular phase, rise slightly on the day of the LH surge (1–2 ng/mL) and steadily thereafter, peak 7 to 8 days after ovulation, and then decline over the days preceding menses. Any level greater than 3 ng/mL provides reliable objective evidence that ovulation has occurred.4 It is usually performed in the midluteal phase around day 21 of menstrual cycle.
Serum progesterone levels have also been used to measure the adequacy of luteal function. Accurate judgment requires daily serum progesterone determination because the corpus luteum progesterone secretion is pulsatile in nature, closely correlating with distinct pulses in pituitary LH release. Levels ranging from as low as 2 ng/mL to as high as 40 ng/mL can be observed, within brief intervals of time.5 However, daily estimations are both costly and impractical. Sampling during the morning hours when progesterone concentrations are generally high and less erratic may be helpful.6 A sum of 3 measurements obtained between the 5th and 9th day after ovulation totaling 30 ng/mL or more have also been recommended.7 A single measurement greater than 10 ng/mL at day 21 of menstrual cycle is often used. However, random serum progesterone concentrations defy confident interpretation of adequacy of luteal phase and have little value beyond documenting ovulation.
 
Advantages
  1. Simple
  2. Reliable
  3. Minimally invasive
  4. Widely available
  5. Assesses adequacy of luteal phase also.
7
 
Disadvantages
  1. False-negative or-positive due to fluctuating levels.
  2. Multiple samples required for accuracy.
 
Midcycle LH Surge
It is a relatively brief event, typically lasting for 48 and 50 hours. LH has a short half-life and is rapidly cleared via the urine. Ovulation predictor kits turn positive when the urinary LH concentration exceeds a threshold level normally seen only during the LH surge. The threshold level for the ELISA kit is 40 mIU/mL.
 
Methods
LH surge can be detected by the following methods:
  1. ELISA: It is the commonly used method.
  2. RIA: It is very accurate.
  3. Slide test.
 
Principle of ELISA Test
It contains 2 antibodies one directed against α subunit which is attached to enzyme alkaline phosphates and the other against the β subunit which is attached to test pad. When LH is present in urine, a sandwich is formed and the enzyme is available to convert a noncolored substance to chromogen (blue). The color intensity produced is proportional to the concentration of LH in the urine sample.
Time of test: The first morning void would be an ideal specimen to test because it is typically the most concentrated. However, results correlate best with the serum LH peak when testing is performed in the late afternoon or early evening hours (4.00 to 10.00 PM), probably because LH surges often 8begin in the early morning hours and are not detected in urine until several hours later.2 Twice daily testing decreases the frequency of false-negative results.
Precautions:
  1. Testing must be done on a daily basis as test is positive on only a single day, occasionally on two consecutive days.
  2. Patients should be advised to avoid drinking large volumes of fluids a short time before they plan to test as results are sensitive to the volume of fluid intake.
Interpretation: Ovulation generally follows within 14 to 26 hours after detection of the urine LH surge and almost always within 48 hours.8 The period of greatest fertility includes the day of LH surge detection and the following 2 days. The day after the first positive test generally is the one best for timed intercourse and artificial insemination.8,9
Accuracy of test: The accuracy of many ovulation predictor kits available varies. The kits predict ovulation with greater than 90% probability.9,10 The positive and negative predictive values of these tests are 90% and 96%, respectively. If the urine is checked twice a day sensitivity increases to 97–99%. About 5–10% of women do not produce positive results either because of failed recognition by the antibody used or because their peak urinary LH concentration does not rise above the threshold set by the kit manufacturers. True false-positive tests are rare but equivocal results are not and can be both confusing and frustrating. About 28.7% of patients undergoing ultrasound-monitored IUI (intrauterine insemination) cycle had a spontaneous LH surge before ovulation triggering was scheduled. This could affect pregnancy rates following IUI. Hence, LH surge may have a more important role than ultrasound monitoring in timing IUI or coitus.119
False-positive: False-positive results may occur with intake of these drugs.
  1. Oral contraceptive pill
  2. Danazol
  3. Exogenous hCG
  4. hMG
  5. Clomiphene citrate.
 
Advantages
  1. Noninvasive
  2. Widely available
  3. Not time consuming
  4. Predict when ovulation will occur unlike other methods which are analyzed in retrospect
  5. Helps to define the length of the follicular and luteal phase and to identify other cycle abnormalities.
 
Disadvantages
  1. Tedious
  2. False-negative results because of short duration of LH surge.
  3. Accuracy is affected by fluid intake.
 
Endometrial Biopsy
Endometrial biopsy is a test of ovulation based on the characteristic histological changes in the endometrium resulting from the action of progesterone. During the follicular phase of the menstrual cycle, the endometrium exhibits a proliferative pattern, reflecting the growth stimulated by rising levels of estrogen derived from the dominant ovarian follicle. During the luteal phase, progesterone secreted by the corpus luteum causes the secretory transformation of the endometrium. 10Anovulatory women are always in the follicular phase and have a proliferative endometrium which can become hyperplastic with extended exposure to a constant estrogen growth stimulus. The histologic features of the secretory endometrium change, with the duration of progesterone exposure. The experienced pathologists can “date” the endometrium, providing a retrospective estimate of how many days have passed since ovulation occurred. The observed date can then be compared to the actual date of sampling. In recent years it has been more accurately defined prospectively, by the number of days elapsed since the detection of the LH surge or ultrasound observation of follicular collapse. Histologic and sampling dates that agree, within a 2-day interval, have generally been considered normal. Dates more than 2 days “out of phase” in two consecutive cycles is the standard criterion for the diagnosis of luteal phase deficiency.12 The best time for the biopsy is controversial. Some advocate the premenstrual phase, when the endometrium might best reflect the cumulative effects of corpus luteum function, while others have argued that the midsecretory phase, coinciding with the putative implantation window, is more relevant being able to identify abnormalities of endometrial maturation which may go undetected.13 A careful and systematic study has revealed that normal variations in histologic characteristics among individuals, between cycles in individuals and among different observers are simply too great to reliably define a specific luteal day or even a narrow interval of days.
To conclude the body of the available evidence supports the conclusion that the traditional endometrial histologic dating is not a valid diagnostic tool. Consequently, endometrial dating cannot be used to guide the clinical management of women with reproductive failure and should no longer be regarded as an important element of their evaluation.11
 
Advantages
  1. Simple office procedure.
  2. Few complications.
 
Disadvantages
  1. Invasive.
  2. Costly.
  3. Not very accurate: Numerous studies and analyzes have described significant intraobserver and interobserver variations in histologic interpretation that are great enough to affect diagnosis and management in 20 to 40% of individual women.14
 
Ultrasonography
Although not providing definite positive proof that ovulation actually occurred, serial transvaginal ultrasound examinations offer details about the size and number of preovulatory follicles and provide the most accurate estimate of when ovulation occurs (See Chapter 14).
In its final stages of development, the preovulatory follicle grows at a predictable pace, approximately 2 mm per day (range of 1–3 mm/day). After ovulation, the follicle abruptly decreases in size, its margins become less distinct, the density of internal echoes increases and fluid in cul-de-sac is seen Table 1.4.15 Abnormal patterns of follicle development can also be observed.
Table 1.4   Signs of ovulation on ultrasonography
1. Margins of follicle become indistinct
2. Increase in density of internal echoes in follicle
3. Irregularity of follicle
4. Abrupt decrease in size of follicle
5. Fluid in cul-de-sac
12
The follicle may grow at an abnormal pace, collapse when still relatively small (atresia), or continue to grow or fail to rupture, persisting as a cyst for days after the LH surge (luteinized unruptured follicle).16 3D ultrasound monitoring is a new introduction and is more closer to physiological monitoring.17
Each of the available tests is useful and no one test is necessarily the best. Some are very simple, noninvasive, and inexpensive, and others are more complicated, invasive and costly. Pregnancy is the only sure positive proof of ovulation.
 
References
  1. Fairley DH, Taylor A. Anovulation. Br Med J. 2003;327:546–9.
  1. Luciano AA, Peluso J, Koch El, Maier D, Kuslis S, Davison E. Temporal relationship reliability of the clinical, hormonal, and ultrasonographic indices of ovulation in infertile women. Obstet Gynecol. 1986;75:412–6.
  1. Quagliarello J, Arny M. Inaccuracy of basal body temperature charts in predicting urinary luteinizing hormone surges. Fertil Steril. 1990;45:334–7.
  1. Wathen NC, Perry L, Lilford RJ, Chard T. Interpretation of single progesterone measurement in diagnosis of anovulation and defective luteal phase: Observations on analysis of the normal range. Br Med J. 1984;288:7–9.
  1. Fillcori M, Butler JP, Crowley WF. Neuroendocrine regulation of the corpus luteum in the human: Evidence for pulsatile progesterone secretion. J Clin Invest. 1984;73:1638–47.
  1. Syrop CH, Hammond MG. Diurnal variations in midluteal serum progesterone measurements. Fertil Steril. 1987;47:67–70.
  1. Jordan J, Craig K, Clifton DK, Soules MR. Luteal phase defect: The sensitivity and specificity of diagnostic methods in common clinical use. Fertile Steril. 1994;62:54–62.
  1. Miller PB, Soules MR. The usefulness of a urinary LH kit ovulation prediction during menstrual cycles of normal women. Obstet Gynecol. 1996;87:13–7.
  1. Martinez AR, Bernardus RE, Vermeiden JP, Schoemaker J. Time scheduled of intrauterine insemination after urinary lutenizing hormone surge detection and pregnancy results Gynecol Endocrinol. 1994;8:1–5.
  1. Nielsen MS, Barton SD, Hatasaka HH, Stanford JB. Comparisons of several one step home urinary luteinizing hormone detection test kits to OvuQuick. Fertil Steril. 1998;76:384–7.
  1. Antaki R, Dean N L, Lapensée L, Racicot M H, Ménard S, Kadoch I J. An algorithm combining ultrasound monitoring and urinary luteinizing hormone testing: a novel approach for intrauterine insemination timing. J Obstet Gynaecol Can. 2011;33(12):1248–52.
  1. Duggan MA, Brashert P, Ostor A, Scurry J, Billson V, Kneafsey P, Difrancesco L. The accuracy and interobserver reproducability of endometrial dating. Pathology. 2001;33:292–7.
  1. Casteibaum AJ, Wheeler J, Coutiferis CB, Mastroianni L, Lessey BA Jr. Timing of the endometrial biopsy may be critical for the accurate diagnosis of luteal phase deficiency. Fertil Steril. 1994;61:443–7.
  1. Scott RL, Snyder RR, Strickland DM, Tyburski CC, Bagnall JA, Reed KR, et al. The effect of interobserver variation in dating endometrial histology on the diagnosis of luteal phase defects. Fertil Steril. 1988;50:888–92.
  1. de Crespigny LC, O'Herlihy C, Robinson HP. Ultrasonic observation of the mechanism of human ovulation. Am J Obstet Gynecol. 1981;139:636–9.
  1. Matijevic R, Grigic O. Predictive value of ultrasound monitoring of menstrual cycle. Curr Opin Obstet Gynecol. 2005;17(4):405–10.
  1. Murtinger M, Aburumieh A, Rubner P, Eichel V, Zech M H, Zech M H. Improved monitoring of ovarian stimulation using 3D transvaginal ultrasound plus automated volume count. Reprod Biomed Online. 2009;19(5):695–9.

Oral Ovulogenschapter 2

Surveen Ghumman
 
Clomiphene
Clomiphene citrate (CC) is an orally active nonsteroidal triphenylethylene derivative approved for clinical trials in 1967. It has both estrogen agonist and antagonist effects by acting on α and β estrogen receptors. It is used in clinical practice as an antagonist, as the agonist properties manifest only when endogenous estrogen levels are very low (Fig. 2.1). The commercially available preparation is a racemic mixture of two sterochemicals in the ratio of 38% zuclomiphene or less active cis-isomer, and 62% enclomiphene or active transisomer which is responsible for the ovulation induction property of clomiphene.1
After oral administration, it undergoes enterohepatic circulation and may be found in serum up to 30 days. Enclomiphene is cleared rapidly, while zuclomiphene has a long half-life. The two clomiphene isomers have mixed estrogenic and antiestrogenic effects with zuclomiphene having a greater estrogenic activity than enclomiphene.15
Fig. 2.1: Mechanism of action
Only 51% of the oral dose is excreted after 5 days. Significant levels of plasma concentration of zuclomiphene can be detected even after one month but there is no evidence of important clinical significance as it is less active isomer.
In normally ovulating woman clomiphene increases GnRH pulse frequency, but in anovulatory PCOS women it increases the pulse amplitude as frequency is already very high.
 
Indications for Use
  1. Anovulation or infrequent ovulation as in PCO (WHO category II).16
  2. Infertility for causes other than ovulatory to time IUI or increase number of oocytes.
  3. Luteal phase defects.
  4. Unexplained infertility.
The mechanism of inadequate corpus luteum can be due to insufficient FSH stimulation during follicular phase. Clomiphene acts by removing any dysfolliculogenesis. In patients of unexplained infertility, controlled ovarian stimulation increases the level of ovarian steroids and may overcome the subtle deficiencies that these women may have. It also increases the number of oocytes available in one cycle.
 
Contraindications
  1. Liver disease
  2. Ovarian cyst
  3. Development of visual symptoms on administration of drug
  4. Ovarian failure
  5. Hypothalamic pituitary failure (WHO Group I)—as clomiphene requires an intact hypothalamic-pituitary ovarian axis for its action.
 
Prerequisites before Clomiphene Therapy
  1. History and examination to determine duration and cause of infertility.
  2. Evaluation of male partner.
  3. Prolactin level assessment—as abnormal levels will require additional treatment besides clomiphene.
  4. Thyroid function test—as specific treatment is needed.17
  5. Pituitary function assessment by baseline hormonal evaluation—as clomiphene requires a functional hypothalamic pituitary ovarian axis for results.
  6. Liver function tests.
  7. Adrenal function assessment—if hirsutism and other evidence of androgen excess are present.
  8. Determination of ovarian tissue responsiveness to gonadotropins.
  9. Tubal factor evaluation—Done only if there is suspicion or evidence of tubal factor involvement, otherwise it is evaluated only on failure of clomiphene treatment. It may also be recommended in women above 35 years to avoid wasting time on ineffective treatment as fertility is rapidly declining.
 
Dose
Clomiphene is started from day 2 to 5 of the spontaneous or progestin induced menstrual cycle and in amenorrheic patients; it can be started immediately if pregnancy is ruled out. Giving clomiphene on day 5 results in increased gonadotropins at the time when the dominant follicle is being selected. Starting it earlier stimulates multiple follicular development which is ideal in order to obtain more than one oocyte. Outcome is similar, if it is started on any day between 2 and 5 of the cycle.2 The starting dose of clomiphene is 50 mg for 5 consecutive days (Fig. 2.2). More sensitive patients may be started with a lower dose of 25 mg. Dose can be increased every month by 50 mg up to 250 mg till ovulation occurs. However, it is seen that cumulative conception rate does not increase substantially beyond a dose of 150 mg due to the adverse effect of clomiphene on cervical mucus and endometrium (Table 2.1).3
18
 
Results
There is an ovulation rate of 80% which decreases with BMI, age, free androgen index and history of oligomenorrhea.5 The cycle fecundity is 15% in women who respond to treatment and increases to 22% if no other infertility factor is present.6 Cycle fecundity for those with unexplained infertility ranges from 3.4 to 8%.7 This is increased up to 9.5% if IUI is added. However, a recent Cochrane review (2010) stated that there is no evidence of clinical benefit of clomiphene citrate for unexplained fertility.8
 
Predictors of Response
It is important to identify women who will be poor responders in order to timely recommend nonresponders to alternative treatments. Negative factors would be obesity, hirsutism, oligomenorrhea, high free androgens and increased mean ovarian volume. It was found that leptin and free androgen index were most accurate predictors of response in normogonadotropic oligomenorrheic women.9
However, there is no accurate way to predict what dose will be required for an individual woman. In a recent study, concentrations of En and Zu were analyzed and it was found that they accumulated throughout treatment but no statistically significant relationship between En or Zu concentrations, and the dose required to induce ovulation was established. The Zu and En concentrations were not different in the patients who failed to respond, and are not a predictor of the ovulation response to CC or of the dose requirement.10
Normograms have been used to predict response. Age, body mass index, free androgen index, and cycle history were 19used to assign a likelihood of response for each patient on the basis of a published nomogram in a study and predicted 80% nonresponders but could not predict the dose required for response.11
Fig. 2.2: Ovulation induction with clomiphene
20
Table 2.1   Ovulatory and conception rates with different doses of clomiphene4
Dose of clomiphene (mg)
Successful ovulation (%)
Cumulative conception rate (%)
50
52.1
52.8
100
21.9
20.7
150
12.31
9.8
200
6.9
4.8
250
4.9
2.2
 
Duration of Treatment
Duration of treatment can extend to 6 to 12 months but 75% conceptions occur in the first 3 months.12
Since fecundability declines with age, those women above 35 should not undergo prolonged treatment with clomiphene citrate and treatment strategy must move on to other forms earlier after an expanded diagnostic evaluation to exclude other factors.6
21
 
Monitoring
Monitoring can be done as follows:
  1. Basal body temperature is simple, inexpensive but tedious and time consuming.
  2. Serum estradiol levels (each follicle secretes 150 to 300 pg/mL per follicle).
  3. LH surge by home monitoring urine tests (best performed by second urination of the day between 7 am and 10 am). It usually occurs between 5 and 12 days of completion of treatment.
  4. Ultrasonography—Follicular monitoring to be started on day 9. Optimal follicular parameters around ovulation are a follicular size of 18 to 20 mm with perifollicular blood flow of 50 to 75% and RI of 0.4 to 0.48.
  5. Midluteal serum progesterone greater than 3 ng/mL is an evidence of ovulation. A level of progesterone more than 10 ng/mL is taken as an adequate luteal phase. However, this value is not reliable as progesterone levels are very variable.
A study comparing cycle fecundity clomiphene cycle monitored by BBT, LH surge detection and ultrasonography are found no advantage of one over the other.14
Before starting the next cycle it is useful to do a ‘clomiphene check’ where previous treatment cycle is reviewed, and pelvic examination or ultrasound ensures that no residual cyst is present. In recent years, this practice has been thought to be unnecessary; however, a regular contact is recommended to review response to treatment and to ensure an additional evaluation and alternative treatment is not delayed.6
 
Antiestrogenic Effect of Clomiphene
It is thought that the antiestrogenic effect on cervical mucus and endometrium may be responsible for the discrepancy 22between ovulation and conception rates in clomiphene induced cycle.
 
Cervical Mucus
With clomiphene the quantity of cervical mucus is decreased. Effect is dose dependant and more readily apparent when the interval between the last dose of clomiphene and ovulation is short. However, it is seen that the effect is usually negated by high serum estradiol levels due to multi-follicular development or because of end-organ sensitivity. Randomized controlled trials have proved that cervical mucus is not very significant as the postcoital test has little predictive value.
 
Endometrial Growth
Clomiphene inhibits estradiol induction of progesterone receptors in endometrium. The effect is usually inconsistent but is important if preovulatory endometrial thickness is persistently less than 6 mm. In such situation, clomiphene citrate can be replaced by tamoxifen or letrozole. Tamoxifen has an estrogen agonist rather than antagonist effect on the endometrium. Letrozole has no adverse effect on endometrium as it acts by decreasing estrogen production rather than receptor antagonism. Its action is easily reversible when the drug is stopped. Patients can also be started on gonadotropins instead of clomiphene.
In recent studies it has been seen that clomiphene affects endometrium thickness on late proliferative days but not on mid-secretory days, and does not alter the echogenic pattern of the endometrium. The endometrial echogenic patterns in mid-secretory phase of women taking clomiphene who had conceived, were not significantly different from those of women who had not conceived.1523
 
Side Effects and Risks
Minor side effects are seen in 10 to 20% of cases.
  1. Hot flushes occur in 10% and are due to central misperception that endogenous estrogens are low causing vasomotor symptoms.
  2. Nausea and vomiting (2%).
  3. Breast discomfort and bloating.
  4. Hair loss and dryness.
  5. Headache.
  6. Visual disturbances (1.6%): Blurred vision, diplopia, scotoma and light sensitivity may occur and need cessation of the drug and change of treatment options. Rarely optic neuropathy develops. Cases of central retinal vein occlusion (CRVO) have been reported and clomiphene may predispose to this condition especially in patients with associated risk factors for CRVO. Patients should be well informed of this side effect before commencement of therapy. If visual disturbances occur, therapy should be terminated and the patient referred for specialist ophthalmic care.16,17
  7. Ovarian cysts (6.4%): They resolve without treatment in a few weeks.13
  8. Ovarian hyperstimulation syndrome (less than 1%): It can be avoided by establishing and using minimum effective dose. Severe forms are rarely seen.
  9. Multiple pregnancy (5–8%): Usually twins (95%) and very rarely triplets may be seen.
  10. Ovarian cancer: Although earlier studies showed a 3-fold increase in incidence of ovarian cancer, recent studies showed only a small increase of incidence (OR-2.43) of borderline serous tumors but not of invasive cancer.18 A pooled analysis of 8 studies showed that 24neither any fertility drug use nor more than 12 months of use was associated with ovarian cancer. Hence, no change in prescribing practice is warranted on these grounds.
  11. Congenital malformations are not increased. However, it was seen that an abnormal karyotype was present in nearly 50% of women undergoing preovulatory oocyte retrieval after clomiphene stimulation.19 Clomiphene has been found to increase interval of time required for oocytes to reach metaphase I compared with oocytes of natural cycle. The interval of time required for metaphase I to reach metaphase II is significantly reduced (2.4 hours versus 10 hours for natural cycle).20
Despite all the above clomiphene with its ease of administration, reduced need for monitoring and limited cost maintains an important place in treatment of anovulation.
 
Tamoxifen
Tamoxifen is a triphenylethylene derivative with a strong antiestrogenic activity.
 
Mechanism of Action
It inhibits estrogen negative feedback on hypothalamus and pituitary by binding to its receptors in a manner similar to clomiphene.
 
Dose
It is administered orally on second or third day of menstrual cycle in a dose of 20 mg/day for 5 days. The dose can be increased to 40 mg/day.25
 
Results
Ovulation is induced in 70 to 75% patients with a pregnancy rate of 35%. In a recent Cochrane review, no evidence of a difference in effect was found between clomiphene versus tamoxifen or clomiphene.21
 
Advantages
Tamoxifen causes a raised estrogen levels due to multi-follicular development and a direct action on the ovaries to enhance estrogen production, hence leading to a favorable response on the cervical mucus and endometrium. It is an alternative to clomiphene when there is persistently poor endometrial response. It also gives better results in patients with poor cervical mucus score.22
 
Side Effects
Side effects include hot flushes, nausea, vomiting, headache, dizziness, liver toxicity, abdominopelvic discomfort, endometrial hyperplasia and endometrial polyps. Complications, similar to clomiphene include multiple pregnancy, hyperstimulation and ovarian enlargement.
Antiestrogens have an important role to play as first-line ovulation induction in infertile women. They are safe with minimal side effects. However, dose and length of treatment must be individualized. It is important to identify when they should be stopped and further treatment with other drugs initiated.
 
Letrozole
Letrozole is an aromatase inhibitor used in breast cancer cases to suppress estrogen. Although it can be used to induce 26ovulation, it is not approved by FDA for this purpose. The use of this drug is not recommended for ovulation induction currently.
It has no effect on plasma androstenedione and testosterone and so no accumulation of androgens (Fig. 2.3). Letrozole has no effect on endometrium and cervical mucus because of its short half-life and absence of estrogen receptor depletion. As can be seen in Table 2.2 although the estrogen levels are lower with letrozole compared to clomiphene the endometrial response is much better.23 It was also associated with lower rate of multiple pregnancy.24
Fig. 2.3: Mechanism of action
27
Table 2.2   Comparison of effect of clomiphene and letrozole23
Endometrial thickness
Ovulation rate
E2 levels
Number of mature follicles
Pregnancy
Clomiphene
5 mm
67%
1278 pg/mL
1.9
Letrozole
8 mm
87%
392 pg/mL
2.4
30%
 
Advantages
  1. No antiestrogenic effect on endometrial lining and cervical mucus.
  2. Induces monofolliculogenesis and hence does not cause hyperstimulation.
  3. Easily reversible action as half-life is 2 days.
 
Dose
It is given in a single dose of 2.5 to 5 mg/day from day 3 to 7 for 5 days. Recently single dose of 20 mg on day 3 has given comparable success rate for ovulation stimulation.25
 
Side Effects
Hair thinning, nausea, hot flushes, peripheral edema and fatigue have been reported in 5% patients.
Aromatase inhibitors warrant additional study to establish their role as treatment drug for ovulation induction and as a option for clomiphene resistant cases.
 
References
  1. McDonough PG. The clomid twins: Waiting for a single isomer heaven (Editorial). Fertil Steril. 1997;68:186–7.
  1. Wu CH, Wenkel CA. The effect of therapy initiation day on clomiphene citrate therapy. Fertil Steril. 1989;52:564–8.
  1. Kusata E, White DM, Franks S. Modern use of clomiphene citrate in induction of ovulation. Hum Reprod. Update. 1997;3:359–65.
  1. Gysler M, March CM, Mishell DR Jr, Baily EJ. A decade's experience with an individualized clomiphene treatment regimen including its effect on the postcoital test. Fertil Steril. 1982;37:161–7.
  1. Imani B, Eijkemans MJ, te Velde ER, Habbema JD, Fauser BC. Predictors of patients remaining anovulatory during clomiphene citrate induction of ovulation in normogonadotropic oligoamenorrheic infertility. J Clin Endocrinol Metab. 1998;83(7):2361–5.
  1. Use of clomiphene citrate in women. The Practice Committee of American Society of Assissted Reproduction. Fertil Steril. 2006;86(5):S187–93.
  1. Fisch P, Casper RF, Brown SE, Wrixon W, Collins JA, Reid RL, et al. Unexplained infertility: evaluation of treatment with clomiphene citrate and human chorionic gonadotropin. Fertil Steril. 1989;51(5):828–33.
  1. Hughes E, Brown J, Collins JJ, Vanderkerchove P. Clomiphene citrate for unexplained subfertility in women. Cochrane Database of Systematic Reviews. 2010; Issue 1. Art. No.: CD000057.
  1. Imani B, Eijkemans MJ, de Jong FH, Payne NN, Bouchard P, Giudice LC, et al. Free androgen index and leptin are the most prominent endocrine predictors of ovarian response during clomiphene citrate induction of ovulation in normogonadotropic oligoamenorrheic infertility. J Clin Endocrinol Metab. 2000;85(2):676–82.
  1. Ghobadi C, Amer S, Lashen H, Lennard MS, Ledger WL, Rostami-Hodjegan A. Evaluation of the relationship between plasma concentrations of en- and zuclomiphene and induction of ovulation in anovulatory women being treated with clomiphene citrate. Fertil Steril. 2009;91(4):1135–40.
  1. Ghobadi C, Nguyen TH, Lennard MS, Amer S, Rostami-Hodjegan A, Ledger WL. Evaluation of an existing nomogram for predicting the response to clomiphene citrate. Fertil Steril. 2007;87(3):597–602.
  1. Speroff L, Glass R, Kase N. In vitro fertilization. In: Speroff L, Glass R, Kase N (Eds). Clinical Gynaecological Endocrinology and Infertility. William and Wilkins:  Baltimore, London. 1989;611–9.
  1. Rust LA, et al. An individualized graduated therapeutic regimen for clomiphene citrate. N Engl J Med. 1994;331:771–6.
  1. Smith YR, et al. Comparison of low technology and high technology monitoring of clomiphene citrate ovulation induction. Fertil Steril. 1998;70:165–8.
  1. Dehbashi S, Parsanezhad ME, Alborzi S, Zarei A. Effect of clomiphene citrate on endometrium thickness and echogenic patterns. Int J Gynaecol Obstet. 2003;80(1):49–53.
  1. Lee VY, Liu DT, Li CL, Hoi-Fan, Lam DS Central retinal vein occlusion associated with clomiphene-induced ovulation. Fertil Steril. 2008;90(5):2011.e11-2.
  1. Viola MI, Meyer D, Kruger T. Association between clomiphene citrate and visual disturbances with special emphasis on central retinal vein occlusion: a review. Gynecol Obstet Invest. 2011;71(2):73–6.
  1. Ness RB, Cramer DW, Goodman MT, Kjaer SK, Mallin K, Mosgaard BJ, et al. Infertility, fertility drugs and ovarian cancer: A pooled analysis of case control studies. Am J Epidemiol. 2002;155:217–24.
  1. Wramby H, Fredga K, Liedholm P. Chromosome analysis of human oocyte recovered from preovulatory follicles in stimulated cycles. N Engl J Med. 1987;316:121–4.
  1. Seibel MM, Smith DM. The effect of clomiphene citrate on human preovulatory oocyte maturation in vivo. J In Vitro Fertil. 1989;6:3–6.
  1. Brown J, Farquhar C, Beck J, Boothroyd C, Hughes E. Clomiphene and anti-oestrogens for ovulation induction in PCOS. Cochrane Database of Systematic Reviews. 2009; Issue 4. Art No.: CD002249.
  1. Annapurna V, Dhaliwal LK, Gopalan S. Effects of two anti-estrogens, clomiphene citrate and tamoxifen, on cervical mucus and sperm cervical mucus interaction. Int J Fertil Women's Med. 1997;42(3):215–8.
  1. Mitwally MF, Casper RF. Aromatase inhibition improves ovarian response to FSH in poor responders. Fertil Steril. 2002;77(4):776–80.
  1. Mitwally MF, Biljan MM, Casper RF. Pregnancy outcome after the use of an aromatose inhibitor for ovarian stimulation. Am J Obstet Gynecol. 2005;192:381–6.
  1. Mitwally MF, Casper RF. Single dose administration of an aromatose inhibitor for ovarian stimulation. Fertil Steril. 2005;83:229–31.

Clomiphene Resistance—What Next?chapter 3

Surveen Ghumman
Many patients do not respond to clomiphene. There may be a genetic basis to clomiphene resistance and it is seen that the chance of resistance to clomiphene is almost double in women with PCOS (polycystic ovary syndrome) harboring the 680-polymorphism Ser/Ser genotype.1 It is important to define clomiphene resistance and failure.
 
 
Clomiphene Resistance
Clomiphene resistance is failure to ovulate with 3 months use of clomiphene at 150 mg/day for 5 days. It occurs in 20% cases more so in PCOS patients.
 
Clomiphene Failure
Patients who ovulate but fail to conceive after treatment with 3 cycles of clomiphene in a dose of 150 mg/day are cases of clomiphene failure. It is usually due to excess LH, androgens or insulin which leads to impaired folliculogenesis, increased 32atresia, poor oocyte quality, poor endometrial receptivity and deficient corpus luteum function. Other potential infertility factors must be ruled out. On diagnostic laparoscopy, it was seen that significant pelvic pathology was present in one third of these patients, 29.3% had minimal pathology and only 32.6% had a normal pelvis.2
 
Management of Clomiphene Failure or Resistance
All cases with clomiphene failure need a complete endocrinal work-up and a diagnostic laparoscopy to rule out any underlying endocrine disorder or pelvic pathology. These women may respond to additional or alternative treatment. A choice of which treatment is to be initiated is based on patient's history, laboratory results and observation of previous unsuccessful clomiphene cycles. These are alternatives that merit consideration depending on patient's age, goals, available resources and risk tolerance.
 
Weight Loss
Women with central fat have high levels of LH, androstenedione, estrone, insulin, triglycerides, very low-density lipoproteins and lower levels of high- density lipoprotein. These altered levels cause disturbances in hypothalamic pituitary ovarian axis. A high waist-hip ratio (more than 0.85) is associated with greater reproductive hormone and insulin derangement. Even moderate obesity with a body mass index of more than 27 kg/m2 is associated with a lesser chance of ovulation.3 Adipose tissue is an active site for 33steroid production and metabolism. It converts androgens to estrogens by aromatase activity (Table 3.1). There is defective clearance and production of androgens in central obesity. Increasing BMI is associated with an increased requirement for clomiphene. Larger doses of clomiphene up to 200 mg/day are required to ensure ovulation in obese women. Doses of gonadotropin required are also more. Weight loss improves the clinical and biochemical parameters that are disordered due to obesity. Loss of 5 to 7% of body weight is effective in restoring ovulation and leads to changes in levels of insulin, IGF and SHBG. Exercise and dietary restraint produce a favorable endocrine status of these patients and a better ovulation rate with clomiphene.4 There is a higher conception rate and a lower miscarriage rate. Even surgically induced weight loss will induce these changes.
 
Extended Course of Clomiphene Treatment
Clomiphene can be given for up to 7 to 10 days to induce ovulation in those cases which do not respond to the 5 day regime.5 Up to 50% of women who are resistant to the standard regime respond to it.
 
Tamoxifen
It can be tried alone or in combination with clomiphene especially in cases of poor cervical mucus score and in cases of poor endometrial response.
Table 3.1   Effect of adipose tissue on steroid hormone levels
• Storage of steroid hormones
• Affects insulin secretion from pancreas
• Conversion of estrogen from inactive to active form
• Peripheral conversion of androgens to estrogens
• Changes in level of sex hormone-binding globulin (SHBG)
34
Tamoxifen causes a raised estrogen levels due to multi-follicular development and a direct action on the ovaries to enhance estrogen; hence, leading to a favorable response on the cervical mucus and endometrium. It is given in a dose of 20 to 40 mg/day from day 2 to 5 days.
 
Aromatase Inhibitors
Letrozole: Letrozole given for 5 days showed a ovulation rate of 33.3% in clomiphene resistant cases.6 However, if administered as long protocol (10 days) it can produce more mature follicles and subsequently more pregnancies than the short letrozole therapy (5 days) in clomiphene resistant women with PCOS.7 In clomiphene resistant PCOS patients, letrozole results were nearly comparable to gonadotropin therapy with an ovulation rate of 79.3% and pregnancy rate of 23.4%. It is found to be most effective when baseline estradiol level was more than 60 pg/mL.8 However, letrozole is no longer approved for use as an ovulation induction drug.
Anastrazole: Anastrazole in PCOS clomiphene resistant women had an ovulation rate of 63.4% and a pregnancy rate of 15.1%, which was found to be comparable to letrozole.9
 
Glucocorticoids
Dexamethasone 0.5 mg/day or prednisolone 5 mg/day is added either continuously or in follicular phase from day 5 to 16 to bring down raised levels of DHEAS (>200 μg/DL) in PCOS patients. It results in an ovulation rate of 80% compared to 20% in cases given placebo and a cumulative pregnancy rate which is 10-fold (40% vs 4%). This effect is seen even 35in cases where DHEAS is normal.10 The pregnancy rate in women with unexplained infertility undergoing IUI after ovulation induction with clomiphene showed a pregnancy rate of 21.4% vs 4.5% with and without dexamethasone, respectively.11
A recent Cochrane review (2009) supported these findings.12 Glucocorticoids diminishes the androgen level in the microenvironment of the ovary by blunting the night time peak of ACTH, thus decreasing the adrenal contribution to circulating androgens. However, the mechanism involves more than androgen suppression. It may directly effect oocyte or induce indirect effects on intrafollicular growth factor and cytokines which may act synergistically with FSH. It may be continued for 3 to 6 cycles if successful and should be discontinued if not (ASRM Guidelines 2006).13 There is no evidence that glucocorticoids have any major side effect in this dose when used. It is stopped if pregnancy occurs.
 
Human Chorionic Gonadotropin (hCG)
About 20% of cases, ovulation does not occur in clomiphene induced cycle because there is a failure of rupture of a developed follicle. hCG is given in a dose of 5,000 to 10,000 IU when follicle is 18 to 20 mm on ultrasonography, in cases where there is repeated evidence of unruptured follicle. It is also helpful in timing intrauterine insemination where midcycle LH surge may remain falsely undetected because of its brief duration. Recombinant hCG can be used in a single dose of 250 μg subcutaneously and has similar pharmacokinetics as the urinary formulation. All studies show that other than these two indications there is no difference in results with or without exogenous hCG.5 Midcycle hCG administration has no effect on luteal function.1436
 
Suppressive Therapy
Suppression can be done either with oral contraceptives or GnRH agonists and is used when there are raised LH or androgen levels as in PCOS women.
 
Oral Contraceptive
Suppression with oral contraceptives decreases ovarian androgens, luteinizing hormone, FSH and 17 β-estradiol and may be responsible for the improved response in patients who previously were resistant to clomiphene citrate. It restores normal function in a patient with dysfunctional hypothalamic pituitary ovarian axis manifesting with anovulation. An ovulation rate of 70% and a cumulative pregnancy rate of 50% was achieved with such treatment in clomiphene resistant cases.15
 
GnRH Agonist
GnRH agonists cause a down regulation of pituitary. It is given in a dose of 0.1 mg/day from day 21 of previous cycle. LH and FSH levels are brought down (E2 < 30 pg/mL, LH < 2.5 IU/L, Progesterone < 2 ng/mL). Then stimulation is begun with gonadotropins and GnRH agonist dose is reduced to half. This decreases high basal LH levels, decreases LH stimulation of ovarian androgen production and eliminates any premature LH surge.
When combined with a oral contraceptive there is a dual advantage of a greater and more sustained reduction of LH, improved luteinizing hormone-to-follicle-stimulating hormone ratio and lower serum androgens, particularly dehydroepiandrosterone sulfate. It also prevents estrogen deficiency which develops on using GnRH agonist without add back therapy (Fig. 3.1).16 Results are good in clomiphene resistant cases.17,18 37
Fig. 3.1: Combined suppressive action of oral contraceptive and GnRH agonist
 
Dopamine Agonists
 
Bromocriptine
For patients with raised serum prolactin levels bromocriptine in a dose up to 2.5 mg bd or tds is used orally or vaginally. Long-acting slow release or depot preparations are also available. Ovulatory dysfunction in presence of galactorrhea responds well to bromocriptine even if prolactin level is normal.19 About 80% patients have restoration of ovulation. However, a recent study found no advantage of adding bromocriptine in clomiphene resistant patients with normal prolactin. No significant differences was seen in ovulation, and serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), dehydroepiandrosterone sulfate (DHEAS), progesterone (P) between treatment and placebo group after treatment. Serum prolactin levels were reduced.20
 
Carbergoline
Carbergoline can be used in bromocriptine resistant cases in a weekly dose of 0.5 to 3 mg orally or vaginally. It has less side effects, the most common being headache.38
 
Insulin Sensitizers
Metformin: Patients with hyperinsulinemia (fasting insulin more than 25 IU or a fasting glucose to insulin ratio of less than 4.5) in PCOS require insulin sensitising drugs like metformin which is given in a dose of 1500 mg/day. Some recommend administration of insulin sensitizers at fasting insulin level of more than 15 IU. Now postprandial insulin levels are also taken into account and levels more than 100 IU are significant Altered GTT is considered the most reliable method of establishing insulin resistance. By reducing hyperinsulinemia metformin causes a reduction in intraovarian androgens. This also leads to reduction in E2 levels and induces an orderly follicular growth restoring ovulation. Ovulation rates are higher when combined with clomiphene (76% vs 46% when used alone).21,22 A recent meta-analysis published in 2008 showed that pregnancy rates were also increased. The live birthrate following up to 6 months of treatment with metformin given along with clomiphene was increased but not so significantly (26.8%vs 22.5% with clomiphene alone).23
Therefore, the use of metformin in improving reproductive outcomes in women with PCOS appears to be limited.24 Metformin may be added to clomiphene citrate in women with clomiphene resistance who are older and who have visceral obesity.25
Ultrashort metformin pretreatment: Women with clomiphene resistant PCOS can be started on 1500 mg metformin daily for 12 days, followed by clomiphene 150 mg daily for 5 days along with metformin. Twelve days of metformin 39pretreatment improves ovulation and pregnancy rates in women with clomiphene-resistant PCOS with 42.5% women ovulating, and 15% conceiving vs 12.5% women ovulating but none conceived in the clomiphene only group as seen in a recent study.26 A Cochrane review 2008 concluded that more randomized controlled trials are required before short treatment can be recommended.27
Pioglitazone and Rosiglitazone: Pioglitazone in a dose of 30 to 50 mg/day or Rosiglitazone in a dose of 4 to 8 mg/day can also be used as monotherapy or in combination with metformin.28,29 A recent study showed rosiglitazone when compared to metformin has higher ovulation rates (64.3% vs 36.4%) and pregnancy rates (50% vs 38.5%) in clomiphene-resistant cases when given at the start of an induced cycle. These findings suggest that short-term use of rosiglitazone with clomiphene is more efficacious than short-term use of metformin with clomiphene in these women.
D-chiroinositol: Administration of D-chiroinositol makes up deficiency of D-chiroinositol containing phosphoglycan which mediates action of insulin in these patients. It is given in a dose of 1200 mg/day for 6 to 8 weeks to correct ovulatory dysfunction.
N-acetyl cysteine: In clomiphene-resistant PCOS women, metformin was more effective than N-acetyl cysteine when added to clomiphene giving significantly higher ovulation and pregnancy rates (69.1% vs 20.0%, and 22.7% vs 5.3%, respectively).30
 
Acarbose
Acarbose is an α-glucosidase inhibitor, used in the management of type 2 diabetes. Acarbose reduces the postprandial rise in both serum glucose and insulin levels 40by inhibiting α-glucosidase, an enzyme responsible for the intestinal absorption of carbohydrates. Acarbose was found to be a safe and effective agent that could be used in cases with clomiphene-resistant PCOS. It was as effective as metformin and is given in a dose of 100 mg tid. When compared with metformin both groups experienced a significant increase in ovulation and monthly mid-luteal serum progesterone levels during the 3-month treatment period compared with pretreatment scores and ovulation rates were similar between the acarbose and metformin groups. Acarbose may be an alternative to metformin for women with PCOS and clomiphene citrate resistance.31
 
Bezafibrate
Dyslipidemia is commonly observed in PCOS patients. Bezafibrate is a drug for dyslipidemia acting through peroxisome proliferator-activated receptors. It was found to be beneficial for ovulation induction in patients with PCOS with dyslipidemia who were resistant to clomiphene citrate in a small study. It was given in a dose of 400 mg/day from day 1 of menses and clomiphene citrate 100 mg/day from day 5 of menses simultaneously until one follicle measuring at least 18 mm in diameter was found by transvaginal ultrasound. Five of seven patients successfully ovulated. Bezafibrate may be effective for ovulation induction in CC-resistant PCOS patients with dyslipidemia. However, larger studies are required.32
 
Naltrexone
Endogenous opiates may affect various aspects of reproductive and metabolic function in patients with polycystic ovary syndrome (PCOS). Naltrexone (50 mg po daily) for 6 months caused long-term inhibition of the opioid system. In CC-resistant 41women with PCOS, there was significant reductions in BMI, fasting serum insulin, luteinizing hormone (LH), LH/follicle-stimulating hormone ratio and testosterone improving a broad range of clinical, endocrine and metabolic derangements characteristic in PCOS. 10% ovulated only on natrexone. 33% patients conceived when clomiphene was added showing that it restored clomiphene sensitivity resulting in a significant number of pregnancies.33 However, larger studies need to be conducted to rule out possible teratogenicity of naltrexone.
 
Gonadotropins
Gonadotropins may be given along with clomiphene to improve results and are specially indicated in unexplained infertility and where there is no success with clomiphene. Sequential treatment is only given if some response is seen with clomiphene. In cases where no response is seen with clomiphene, it is better to directly stimulate with gonadotropins. Treatment is individualized in the same way as traditional gonadotropin therapy based on transvaginal sonography and estradiol levels. Clomiphene citrate is given in a dose of 100 mg from day 2 to day 6. FSH is started on day 6 in a dose of 75 to 150 IU/day till adequate follicular development occurs (Fig. 3.2).
Fig. 3.2: Clomiphene and gonadotropin regime
42
Adequate follicular development is taken as estradiol more than 500 pg/mL or one follicle equal to or more than 14 mm mean diameter. The aim is monofollicular development and not superovulation in these anovulatory infertile women.13 Since clomiphene increases LH, FSH is preferred to hMG. Along with FSH, GnRH antagonist can be added to suppress premature LH surge. A Cochrane review however stated that in women with PCOS, no significant difference could be demonstrated between FSH and hMG, in terms of pregnancy rate. However, given similar cost, potential advantages in terms of purity and a possible reduction in OHSS risk, highly purified or recombinant FSH are likely to be widely adopted.34
 
GnRH Antagonists
They act reversibly by competitive inhibition of GnRH receptors resulting in rapid decline in LH and FSH. There is no stimulatory phase unlike the GnRH agonists. It reduces the dose and duration of gonadotropin treatment as it does not nullify FSH or LH secretion but interrupts the premature LH surge.
There are two protocols used in ovulation induction with antagonists. In both gonadotropins are started as usual. When follicle reaches 14 mm (usually Day 7), in the Lubeck Protocol antagonist, Centrorelix, is started at a dose of 0.25 mg/day (Fig. 3.3) whereas in the French protocol a single dose of 3 mg is given. Cochrane review 2002 has concluded that both protocols were equally effective in preventing premature LH surge.35
 
Surgical Ovulation Induction
Laparoscopic ovarian drilling (LOD) is an alternative to ovulation induction with gonadotropins for polycystic ovarian 43syndrome (PCOS) patients unresponsive to clomiphene.36
Fig. 3.3: Clomiphene with gonadotropins and GnRH antagonist
Since it is as effective as FSH treatment in terms of live births, and reduces the need for ovulation induction or ART in a significantly higher proportion of women. It also increases the sensitivity to clomiphene. Laparoscopic ovarian drilling using electrocautery or laser photocoagulation can be done (Fig. 3.4). The advantage is that it is a single time treatment free of intensive monitoring with no risk of OHSS and multiple pregnancy.37 Ovulation occurs in 70–80% of patients and there is a conception rate of 60%. Complications include adhesion formation (80%) and ovarian atrophy.38 Unilateral ovarian diathermy was as effective and long lasting as bilateral ovarian diathermy in the resumption of menstruation and pregnancy rates.39 However, there are ongoing concerns about long-term effects of LOD on ovarian function.40
Patients with clomiphene resistance require further evaluation and form a challenging diagnostic problem to the infertility specialist. A step-by-step approach to rule out and treat other subclinical endocrinopathies is required.44
Fig. 3.4: Management of clomiphene resistance
45
 
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  1. Sinawat S, Buppasiri P, Lumbiganon P, Pattanittum P. Long versus short course treatment with metformin and clomiphene citrate for ovulation induction in women with PCOS. Cochrane Database Syst Rev. 2008;(1):CD006226.
  1. Stout DL, Fugate SE. Thiazolidinediones for treatment of polycystic ovary syndrome. Pharmacotherapy. 2005;25(2):244–52.
  1. Ortega-Gonzalez C, Luna S, Hernandez L, Crespo G, Aguayo P, Arteaga-Troncoso G. Responses of Serum Androgen and Insulin Resistance to Metformin and Pioglitazone in Obese, Insulin-Resistant Women With Polycystic Ovary Syndrome. J Clin Endocrin Metab. 2005;90(3):1360–5.
  1. Abu Hashim H, Anwar K, El-Fatah RA. N-acetyl cysteine plus clomiphene citrate versus metformin and clomiphene citrate in treatment of clomiphene-resistant polycystic ovary syndrome: a randomized controlled trial. J Women's Health (Larchmt). 2010;19(11):2043–8.
  1. Sönmez AS, Yasar L, Savan K, KoçS, Ozcan J, Toklar A, et al. Comparison of the effects of acarbose and metformin use on ovulation rates in clomiphene citrate-resistant polycystic ovary syndrome. Hum Reprod. 2005;20(1):175–9.
  1. Hara S, Takahashi T, Amita M, Igarashi H, Kurachi H. Usefulness of bezafibrate for ovulation induction in clomiphene citrate-resistant polycystic ovary syndrome patients with dyslipidemia: a prospective pilot study of seven cases. Gynecol Obstet Invest. 2010;70(3):166–72.
  1. Ahmed MI, Duleba AJ, El Shahat O, Ibrahim ME, Salem A. Naltrexone treatment in clomiphene resistant women with polycystic ovary syndrome. Hum Reprod. 2008;23(11):2564-9.
  1. Hughes E, Collins J, Vandekerckhove P. Ovulation induction with urinary follicle stimulating hormone versus human menopausal gonadotropin for clomiphene-resistant polycystic ovary syndrome. Cochrane Database Syst Rev. 2000;(2):CD000087.
  1. Al-Inay H, Aboulghar M. GnRH antagonist in assisted reproduction: a Cochrane review. Hum Reprod Update. 2002;17(4):874–85.
  1. Flyckt RL, Goldberg JM. Laparoscopic ovarian drilling for clomiphene-resistant polycystic ovary syndrome. Semin Reprod Med. 2011;29(2):138–46.
  1. Nahuis MJ, Kose N, Bayram N, van Dessel HJ, Braat DD, Hamilton CJ, et al. Long-term outcomes in women with polycystic ovary syndrome initially randomized to receive laparoscopic electrocautery of the ovaries or ovulation induction with gonadotrophins. Hum Reprod. 2011;26(7):1899–904.
  1. Campo S. Ovulatory cycles pregnancy outcome and complications after treatment of polycystic ovarian syndrome. Obstet Gynecol Survey. 1998;53:297.
  1. Al-Mizyen E, Grudzinskas JG. Unilateral laparoscopic ovarian diathermy in infertile women with clomiphene citrate-resistant polycystic ovary syndrome. Fertil Steril. 2007;88(6):1678–80.
  1. Farquhar C, Lilford RJ, Marjoribanks J, Vandekerckhove P. Laparoscopic ‘drilling’ by diathermy or laser for ovulation induction in anovulatory polycystic ovary syndrome. Cochrane Database Syst Rev. 2007;(3):CD001122.

Gonadotropinschapter 4

Surveen Ghumman
The first birth after gonadotropin stimulation was reported by Alan Trounson in 1981. Since then it has become the cornerstone of ART stimulation protocols. FSH with LH separated by polyvalent antibodies was commercially available by 1987, but these still contained urinary proteins. A highly purified FSH was obtained on removing LH by monoclonal antibodies.1 Finally, the recombinant technology was used for a FSH preparation with absolutely no LH activity.
 
 
Gonadotropin Preparations
  1. Human pituitary gonadotropin.
  2. Human menopausal gonadotropins (75 IU of FSH, 75 IU LH).
  3. Highly purified hMG (75 IU of FSH and 75 IU of LH with <5% of urinary protein).
  4. Purified urinary FSH (75 IU of FSH and < 0.7 IU of LH).
  5. Highly purified urinary FSH (75 IU of FSH and <0.1 IU of LH and < 5% urinary protein).
  6. Recombinant FSH (75 IU of FSH and no LH).
  7. Recombinant LH.51
  8. Human chorionic gonadotropin (hCG).
  9. Recombinant hCG.
 
Follicle Stimulating Hormone
Effective daily dose: It is the dose of gonadotropins which can elicit an ovarian response.
There are two phases of stimulation:
  1. The latent phase: An initial period of 3 to 7 days where there is no measurable ovarian response in spite of follicular growth.
  2. Active phase: After latent phase, a 4 to 7 days period where there is an exponential rise of estrogen with follicular growth.
 
Choice and Dosage of Gonadotropin
The dose and duration of gonadotropin required may vary with the patient and also from cycle to cycle in the same patient. There is a relationship with body weight and dose requirement but the response threshold with all individuals is unpredictable. Implantation, pregnancy, and live birthrates were poorer in obese women and were reduced progressively with each unit of BMI.2
The choice of gonadotropin also depends on indication of controlled ovarian stimulation. In women with hypogonadotropic hypogonadism, the drug of choice is menotropin because it contains both FSH and LH. LH is essential for ovulation and luteinization and in these cases LH levels are low. These patients respond to low doses of gonadotropin stimulation. Luteal phase support may be vital where endogenous LH levels are low (less than 3 IU/L). These patients are prone to hyperstimulation and must be monitored carefully if hCG is given as luteal support (Table 4.1).52
Table 4.1   Choice and dose of gonadotropin
Type of gonadotropin
Dose of gonadotropin
Hypogonadotropic hypogonadism (LH low)
FSH and LH (Menotropin)
Adequate dose
Clomiphene resistance/PCOS (LH high)
Recombinant FSH
Low dose
Unexplained infertility
Any preparation
High dose
In patients with PCOS, LH levels are high. Recombinant FSH is given after down regulation (E2 < 30 pg/mL, LH < 4IU/L). There is a very narrow margin between doses which induce successful ovulation and those which cause hyperstimulation (Table 4.1). Patients with unexplained infertility are older subfertile women and the aim is multifollicular ovulation. Hence, higher doses of gonadotropins are used. In these, normally ovulating women where endocrinopathies have been ruled out, any available gonadotropin preparation can be used. Patients who are obese, above 35 years of age, poor responders, those with a baseline FSH of more than 10 IU/L or those downregulated with GnRH agonists, should be started on a higher dose of 225 IU of FSH. A day 8 LH assay of more than 10 IU/L predicts failure or increased risk of miscarriage if pregnancy occurs (Table 4.2). The CONSORT dosing algorithm individualizes recombinant human FSH (rhFSH) doses for assisted reproduction technologies, assigning 37.5 IU increments according to patient characteristics: basal FSH, body mass index, age and antral follicle count. Use of the CONSORT algorithm achieved an adequate oocyte yield and good pregnancy rates in a preliminary study. Adjustment of the algorithm could reduce cancelation rates.3 53
Table 4.2   Factors influencing the dose of gonadotropin
1. Weight
2. Baseline FSH If > 10 IU/L—give higher dose
3. Age beyond 35 years—a higher dose is required
4. Higher effective daily dose in previous cycle
5. Poor responders
6. PCOS patients are usually started on a lower dose to avoid hyperstimulation
7. Prior down regulation with GnRH agonists—a higher dose is required
8. Hypogonadotropic hypogonadism
 
Anti-Müllerian Hormone Tailored Protocols
AMH-guided, controlled ovarian hyperstimulation protocols can significantly improve positive clinical outcomes, reduce the incidence of complications and reduce the financial burden associated with assisted reproduction. In a recent study, when stimulation protocols were tailored according to the AMH level embryo transfer rates, pregnancy rate per cycle started and live birth- rate increased significantly compared with conventionally treated women. Moreover, the incidence of the ovarian hyperstimulation syndrome (OHSS) fell significantly (6.9 to 2.3%,) and failed fertilization fell from 7.8% to 4.5%. The cost of fertility drug treatment fell by 29% per patient and the overall cost of clinical management of OHSS fell by 43% in the AMH group. Within the AMH-tailored group, the live birthrate was not significantly different between agonist and antagonist-treated groups.4
 
Human Menopausal Gonadotropin vs Highly Purified Human Menopausal Gonadotropin
Patients undergoing controlled ovarian hyperstimulation for IVF that includes HP-hMG preparations produce 54significantly higher implantation (20% vs 8.1%) and pregnancy rates (47.2% vs 19.4%), as compared to the traditional hMG.5
 
Alpha Follitropin vs Beta Follitropin
In a study, it was seen that although both Follitropin beta and alpha achieved a comparable number of retrieved oocytes, the use of follitropin-beta was associated with a tendency toward a lower clinical pregnancy rate (PR), and with significantly higher E2 levels despite the use of significantly lower total gonadotropin dose.6
 
Protocols
The treatment is started within the first 2 days of the menstrual cycle.
 
Step-Up Conventional Protocol
This conventional protocol is started with 150 IU of FSH per day. Serum estradiol is measured on day 8 and transvaginal ultrasonography is done. The dose of gonadotropin is maintained or increased accordingly as indicated. Once the serum estradiol begins to rise, the size and number of the developing follicles is determined every 1 to 2 days along with serum estradiol. If estradiol levels are not increasing then the dose of gonadotropins needs to be increased. A maximum of 300 to 375 IU of FSH can be used. There are studies where doses have gone up to 600 IU. Injection hCG is given once follicle is more than 16 to 17 mm. Patients will ovulate 36 hours after the hCG. This regime is useful for poor responders but has a high rate of multiple pregnancy and ovarian hyperstimulation syndrome. The effective daily dose of gonadotropins must be noted for each cycle. In subsequent stimulation cycles while determining the effective dose of gonadotropins, the response threshold and pattern of 55follicular development observed in previous cycles should be considered (Fig. 4.1).
Fig. 4.1: Step-up conventional protocol
56
 
Step-Up Low Dose Protocol
It is started with an initial dose of 37.5 to 75 IU/day. If no response is seen in terms of estradiol level or follicle, it is increased in increments of 37.5 IU every week. This protocol though safe has an extended duration. Lesser ampules of gonadotropin are used and preovulatory estradiol levels are lower. Age, obesity and raised serum LH levels can adversely affect the outcome of treatment.7 It is useful in patients of PCOS who are prone to hyperstimulation as they have a large number of antral follicles ready to respond to FSH stimulation. Administration of FSH converts already present androgens to estrogens producing very high levels of estrogens and overstimulation of ovary. This is avoided by this protocol as small doses of FSH provide the right amount of stimulation needed to make the process occur in a controlled manner. Homberg et al found in their study comparing conventional vs low dose regimen that patients treated with low dose regimen had a greater pregnancy rate and no hyperstimulation or multiple pregnancy. Patients on conventional treatment had an 11% incidence of OHSS and 33% incidence of multiple pregnancy (Fig. 4.2).8
 
Step-Down Protocol
Since many anovulatory women are very sensitive to low doses of exogenous gonadotropin stimulus the initiating dose of this protocol is determined in one or more pervious stimulation cycles before starting the regime. Total amount of gonadotropins are reduced in this regimen. The treatment is usually started with 225 IU hMG/FSH (or 300 IU in some cases) until follicles of 10 mm is seen. The dose is then reduced to 112.5 IU and 3 days later decreased to 75 IU and this is continued till administration of hCG. This is an effort to promote continued development of the more sensitive dominant follicle while withdrawing support from 57less sensitive smaller follicles in the cohort.
Fig. 4.2: Step-up low dose protocol
It is indicated in oligo-amenorrheic women with PCOS or in high responders in IVF. FSH promotes follicular growth because of 2 events, the ‘FSH threshold’ and the ‘FSH window’. FSH threshold is the level of FSH below which no follicular growth can be 58initiated.9 Usually this level in normal women is 7.8 IU/l. The FSH window is the number of days that serum FSH levels are above the threshold and determines the number of follicles which are activated. Since sensitivity of the follicle increases with development, the required FSH for a follicle will decrease. The balance between the decreasing FSH levels and increasing FSH sensitivity is responsible for the growth of the follicle. Using this concept FSH levels are raised by exogenous FSH to reach the threshold and prolong the window in order to obtain specific number of follicles to be growing. The decreased dose would switch off the recruiting phase and limit the number of dominant follicles. A multiple pregnancy rate of 8% and OHSS rate of 2% is seen. This regimen mimics the physiological normal menstrual cycle.10 It is a second-line protocol in PCOS patients where other regimens do not achieve success (Fig. 4.3).
 
Sequential Step-Up, Step-Down Regimen
It is started similar to the step-up protocol but the dose is reduced by half when leading follicle is 14 mm. This approach reduces the number of lead follicles.11
 
Mild Stimulation Protocol
In the recent years, mild protocols have been introduced aiming at a low stimulation which gives acceptable results with minimal risks and lower cost (See chapter on mild stimulation protocols). Here, the endogenous FSH is also taken advantage of while stimulating. It could be a stimulation regimen in which gonadotropins are administered at a lower than usual dose and/or for a shorter duration throughout a cycle in which GnRH antagonist may or may not be given as co-treatment, or a stimulation in which oral compounds (e.g. antiestrogens) are used either alone or in combination with gonadotropins and GnRH antagonists.12 59
Fig. 4.3: Step-down protocol
Low dose of gonadotropin: A starting dose of 37.5 IU/day FSH may be used in selected cases to prevent ovarian hyperstimulation, without loss of efficacy. In a recent study, follitropin-alpha at 37.5 IU/day was sufficient to achieve ovarian stimulation in 72.8% cycles. A single follicle ≥16 mm in diameter developed in 61.1%. Pregnancy rate was 24.7% with 94.9% singleton and 5.1% twin pregnancies. 4.4% cycles were canceled, mainly due to poor response.13
Role of GnRH antagonist in mild stimulation: The IVF cycle can start with an undisturbed early follicular phase recruitment of follicles by an endogenous FSH rise which occurs in a natural menstrual cycle. The endogenous inter-cycle FSH rise 60is taken advantage of rather than suppressed. FSH is added on day 5 to continue FSH elevation and extend the FSH window, for many follicles to develop. This limits the duration and dose of FSH administration. A premature LH surge may occur with rising estradiol levels which act through positive feedback loop. GnRH, antagonist has an immediate action blocking the pituitary when estradiol levels start rising and approach threshold levels at which an LH surge can occur.
Clomiphene in mild stimulation protocol: The second mild regime includes using clomiphene 100 mg, delayed low dose gonadotropin and a flexible GnRH antagonist administration for ovarian stimulation protocol. Pregnancy rates comparable to the standard stimulation regimens were obtained, with a significant reduction in the total dose of gonadotropin needed and of the economical costs.14
Clomiphene with single dose FSH: Addition of single dose of uFSH on day 3 along with clomiphene recruits the co-dominant follicles earlier at follicular phase and therefore enhances the chances of pregnancy.15 It is not only cost-effective but also prevents multiple pregnancy. In order to determine the ideal dose, a study showed the clinical pregnancy rate was 14.8% when 150 IU of rFSH and 20.4% when only 100 IU of rFSH was given in IUI patients along with clomiphene. The incidence of multiple pregnancy was 41.7% in the first group compared with 12.5% in the second group.16
 
Monitoring
Careful monitoring with serum estradiol levels and ultrasonography is needed to achieve the goal of ovulation without hyperstimulation and multiple pregnancy.61
 
Serum Estradiol Levels
Follicles with a diameter of less than 10 mm produce relatively little estradiol but its level starts rising exponentially doubling every 2 to 3 days before ovulation. A change in the rate of rise of estradiol suggests a need for increasing or decreasing the dose of gonadotropin. For each mature follicle the level of estradiol is 200 to 300 pg/mL. When the gonadotropin injection is given between 5 and 8 pm, the estradiol estimation should be done early morning.
 
Ultrasonography
A baseline ultrasonography is a must. In case there are residual ovarian cysts (more than 10 mm) treatment should be deferred as stimulation in the presence of a cyst is often unsuccessful. In a gonadotropin stimulated cycle follicle exhibits a linear growth but reaches maturity at a much smaller mean diameter. 40% patients ovulate when follicle is of 15 to 16 mm diameter. The rate of growth is 1 to 3 mm/day. Endometrial thickness measurements are also important. Cycle fecundity increases with endometrial thickness. Results are poor if endometrial thickness is less than 7 mm.
 
Problems
  1. Multiple pregnancy (10–40%): Twin births have increased by 50% and higher order births have quadrupled. Risk is reduced if ovulation is not triggered when the estradiol level or number of maturing follicles is excessive.62
  2. Hyperstimulation syndrome: It is seen more in young age, low body weight, PCOS and usage of high doses of gonadotropins. Rapidly rising serum estradiol level, concentration over 2,500 pg/mL and observations of a large number of small and intermediate size follicles indicate a high risk for OHSS. Mild forms of OHSS are seen in 8 to 23%, moderate in 6 to 7% and severe in 1 to 2% of cases stimulated with gonadotropins.
  3. Breast and ovarian cancer: There have been no consistent reports of any causal relationship between gonadotropins and breast or ovarian cancer.
  4. Miscarriage (25%): Miscarriage rates are low in hypogonadotropic hypogonadism and much higher in clomiphene resistant anovulatory women.
 
Recombinant FSH
Recombinant FSH was prepared by transfecting Chinese hamster ovary cell lines with both FSH subunit genes. Starting dose is 50 IU.18 The rFSH is quantified as protein content (mass in μg) rather than biological activity. However, its biological activity is confirmed too. The conversion factor is 75 IU corresponds to between 5 and 5.5 μg fill by mass product.19 The delivery system is a pen-shaped device that is either prefilled or can be adjusted to fill variable doses from the vial. They are either lyophilized powder or liquid formulations found in cartridges or pens.
 
Recombinant vs Urinary FSH
It was seen that although dose and duration of treatment with FSH was less and significantly more number of oocytes were retrieved with rFSH compared with uFSH, there was no difference in pregnancy rate in the two groups.20 However, the pregnancy rates improved significantly when using rFSH 63instead of uFSH in poor responders (33% vs 7%).21 There was no difference in oocytes recovered, dose and duration of treatment and pregnancy rates between alpha folliculotropin (Gonal F) and beta folliculotropin (Puregon).22 Clinical choice of gonadotrophin should depend on availability, convenience and costs as no substantive differences in effectiveness or safety was seen on comparing recombinant FSH with other gonadotropins.23
 
Advantages of Recombinant FSH
  1. It has identical amino acids to natural FSH.
  2. Consistent.
  3. No LH activity.
  4. No contamination with urinary proteins.
  5. Highly specific and pure.
  6. No restriction in supply.
  7. Can be administered subcutaneously.
 
Disadvantages
  1. Cost
  2. Increased incidence of OHSS has been reported in some studies.
 
Recombinant FSH-CTP in IVF
The β subunit of hCG is different from gonadotropic hormones as it has a C-terminal peptide extension which is responsible for reduced clearance resulting in major enhancement of in vivo bioavailabilty. Daily injections of FSH have to be given as it has a short half-life. Genes containing the sequence coding the C-terminal peptide (CTP) of hCG is fused with β subunit of FSH creating an FSH which is long acting with a half-life of 95 hours eliminating need for daily injections. 64Early follicular phase administration of FSH-CTP avoids the need for daily injections as a single injection enables follicular growth over a period of 7 days. Maximum serum levels are obtained after 36 to 48 hours. A second injection 7 days later may cause hyperstimulation. Hence, daily doses of recombinant FSH are given thereafter. It is given as a single subcutaneous injection of 180 μg recombinant FSH-CTP on day 3 followed by daily injections of recombinant FSH 150 IU from day 10 onward combined with GnRH antagonist 0.25 mg subcutaneously to prevent premature surge of LH.24 The pharmacokinetics of corifollitropin alfa and rFSH are quite different but their induced pharmacodynamic effects at the dosages used are similar.25 The safety and efficacy of such regimens is currently being evaluated in large comparative phase III clinical trials. It is recommended that patients should be treated with the appropriate dose of corifollitropin alfa according to their body weight as a lower dose does not result in milder stimulation and a higher dose does not result in an improved ovarian response. Two strengths of corifollitropin are available (for patients ≤ 60 kg and > 60 kg). Compared with a daily dose of 200 IU of rFSH, 150 μg of corifollitropin is equivalent in safety and pregnancy outcomes in women using an antagonist protocol.
In normal responder patients undergoing ovarian stimulation with GnRH antagonist co-treatment for IVF ongoing pregnancy rates of 38.9% for the corifollitropin alfa group and 38.1% for rFSH were achieved showing similar results for number of embryos transferred. Median duration of stimulation was equal (9 days) and incidence of (moderate/severe) ovarian hyperstimulation syndrome was the same (4.1% and 2.7%, respectively).26 Fertilization rates were high, ranging from 66% to 68%. Corifollitropin alfa was generally well tolerated, with a tolerability profile similar to 65that of rFSH. There were no clinically relevant differences in pregnancy complications and the incidence of infant adverse events between the two drugs.27
Hence, compared with seven once-daily injections of rFSH, a single injection of corifollitropin alfa achieves equivalent efficacy, and provides a well tolerated and more convenient treatment option to induce multiple follicular growth prior to assisted reproduction.
 
Recombinant LH
It is known that the follicular selection and final stages of follicular maturation are equally if not more dependent on low circulating levels of LH.28 In addition to stimulating production of thecal androgens as substrate for estrogen synthesis, LH stimulates granulosa cells via LH receptors induced by FSH and estrogen in larger but not smaller follicles. LH then becomes the principal stimulus for final stages of follicular maturation while at the same time declining concentration of FSH starve the smaller more FSH-dependent follicles into atresia. Low dose hCG or recombinant LH can promote larger follicles to grow while hastening the regression of smaller follicles.
Exogenous LH supplementation was consistently associated with higher peak estradiol concentrations. The use of hMG in long GnRH agonist cycles was associated with a 3–4% increase in live birthrate. There was insufficient evidence to make definitive conclusions on the need for exogenous LH activity in GnRH antagonist cycles or the benefit of recombinant LH and hCG protocols. Poor responders and patients 35 years of age and older benefit from exogenous LH.29
Recombinant LH can be used for inducing rupture in a single dose of 15,000 or 30,000 IU which is equivalent to reference treatment of 5000 IU hCG. It is superior with 66regard to incidence of OHSS and has a shorter half-life than hCG. Recombinant LH is also needed in hypogonadotropic hypogonadism in a dose of 75 IU daily for ovulation induction with rFSH for better results.30
 
Human Chorionic Gonadotropin
Human chorionic gonadotropin (hCG) promotes the final stages of follicular maturation helping the oocyte reach metaphase II. Approximately 36 hours are required for completion of meiotic process and oocyte retrieval should be done within this time. It can be derived from human urine or can be recombinant. Recombinant hCG is available in syringes of 250 μg which is equivalent to 5000–6000 IU of hCG. There is no evidence of difference between rhCG or rhLH and uhCG in achieving final follicular maturation in IVF, with equivalent pregnancy rates and OHSS incidence.31 BMI affects hCG levels. The highest levels of hCG were measured in women with the lowest BMI. Patients’ body size, rather than route of hCG delivery, appears to determine circulating levels of hCG.32
Exogenous gonadotropins have been used since last 4 decades. They are highly effective in ovulation induction but are accompanied with the disadvantage of high cost, extensive monitoring and risks of ovarian hyperstimulation and multiple pregnancy.
 
References
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  1. Bellver J, Ayllón Y, Ferrando M, Melo M, Goyri E, Pellicer A, et al. Female obesity impairs in vitro fertilization outcome without affecting embryo quality. Fertil Steril. 2010;93(2):447–54.
  1. Olivennes F, Howies CM, Borini A, Germond M, Trew G, Wikland M, et al. Individualizing FSH dose for assisted reproduction using a novel algorithm: the CONSORT study. Reprod Biomed Online. 2011;22 Suppl 1:S73–82.
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  1. Schoot DC, Harlin J, Shaham Z, Mannerts BM, Lahlou N, Bouchard P, et al. Recombinant human follicle stimulating hormone and ovarian response in gonadotropin deficient women. Hum Reprod. 1994;9(7):1237–42.
  1. Youssef MA, Al-Inany HG, Aboulghar M, Mansour R, Abou-Setta AM. Recombinant versus urinary human chorionic gonadotropin for final oocyte maturation triggering in IVF and ICSI cycles. Cochrane Database Syst Rev. 2011 Apr 13;(4):CD003719.
  1. Elkind-Hirsch KE, Bello S, Esparcia L, Phillips K, Sheiko A, McNichol M. Serum human chorionic gonadotropin levels are correlated with body mass index rather than route of administration in women undergoing in vitro fertilization-embryo transfer using human menopausal gonadotropin and intracytoplasmic sperm injection. Fertil Steril. 2001;75(4):700–4.

Role of GnRH Agonists and Antagonists in Assisted Reproductive Technologychapter 5

Surveen Ghumman
The GnRH agonists and antagonists have occupied an increasingly important position in the ovulation induction protocols. GnRH analogs are able to suppress gonadotropin release and subsequently, the gonadal function. This is the basis for their clinical applications as it controls the premature endogenous luteinizing hormone (LH) surge and therefore, decreases the cycle cancelation rate.
 
GnRH Agonist (GnRHα)
Substitutions in the GnRH molecule cause enhanced affinity for the GnRH receptors and protects against enzyme degradation increasing half-life from 8 minutes to 5 hours.
 
 
Decapeptide Agonists
  • Triptorelin
  • Naferelin
  • Goserelin.
72
 
Nonapeptide Agonists
  • Buserelin
  • Leuprolide
  • Histerelin.
 
Mechanism of Action
The mechanism of action is a “flare effect”, followed by downregulation. Within 12 hours of administration it induces liberation of high amount of FSH and LH and also increases the number of receptors (5-fold increase in FSH, 10-fold increase in LH and 4-fold increase in estradiol receptors). This is the so-called ‘upregulation.’ A continuous administration of GnRH agonist produces the opposite effect. There is a decrease in level of FSH and LH by internalization of the receptor-agonist complex and a reduction in the number of receptors. This is called ‘downregulation’ or desensitization of the pituitary. This eliminates any premature LH surge and decreases LH stimulation of ovarian androgen production. The advantage is reduced cycle cancelation, convenient timing of treatment and higher live birthrates. Stimulation is then begun with gonadotropins. Also it can be used as an ovulation trigger to prevent ovarian hyperstimulation syndrome.
Commonly used GnRH agonists are:
Leuprolide SC 500–1000 μg or IM depot 3.75, 7.5 mg/month
Buserelin SC 200–500 μg/day or 300–400 μg intranasally 3–4 times/day
Goserelin SC implant 3.6 mg/month
Triptorelin SC 100–500 μg/day or IM depot 3.75 mg/month.
 
Route of Administration
  1. Subcutaneous injections.
  2. Sustained release implants.73
  3. Intramuscular depot injections.
  4. Nasal spray.
Subcutaneous: This route is most commonly used because of high bioavailability and low interindividual variation. It results in prolonged and delayed absorption compared to intravenous route.
Intranasal route: Disadvantages of this route include a marked interindividual variation in absorption and considerable losses of peptides by proteolysis and swallowing. Initially preparation like buserelin needed frequent administration (5 times a day); however, nasal preparations like naferelin only require twice a day administration.1 There is an advantage of persistence of drug in the nasal mucosa for up to 24 hours consistent with depot like effects. The drug absorption varies with rhinitis and allergy. It is a more convenient route for the patient as compared to daily injections.
Depot formulation: Since there is more profound suppression of the pituitary gonadal axis with continuous administration of the drug, sustained or continuous release formulations have been developed. Currently available preparations include a suspension of 3.75 mg triptorelin or leuprolide in microcapsules injected intramuscularly once a month or 3.6 mg of goserelin dispersed in a biodegradable polymeric matrix of polylactide-co-glycolide as a cylindrical rod implant injected subcutaneously every month. The pharmacokinetics of the two differ. In both, the drug is released over 30 to 55 days. The number of follicles developed, E2 levels, oocyte quality, fertilization and pregnancy rates were same as with daily administration.2 Because of prolonged desensitization the dose of gonadotropins needed is more and stimulation is longer.274
 
Use of GnRH Agonist in COH Protocols
The use of GnRH agonists decreased cancelation rates in IVF cycle from 20 to 2% and improved fertilization and implantation rates.3 Three protocols have been described:
  1. Long protocol.
  2. Short protocol.
  3. Ultrashort protocol.
 
Protocols for Administration
 
Long Protocol
75
 
Advantages of Long Protocol
  1. It prevents unwanted LH surge and hence, cancellation of cycles.
  2. It helps plan the time of ovum pick-up.
  3. Improves overall pregnancy rates especially in patients with raised LH levels.
  4. Allows synchronization ingrowth of follicles.
  5. Reduces intensive monitoring of cycles to detect premature LH surge.
  6. Most studies show a better result with the long protocol than with short or ultrashort protocol.
 
Disadvantages of Long Protocol
  1. Extends or prolongs treatment cycle.
  2. Higher doses of gonadotropins are needed.
  3. More expensive.
  4. It is associated with symptoms of depression in hypogonadal phase.4
 
How Long Can we Continue with GnRH Agonist if Suppression is not Achieved?
In a recent study suppression was obtained after 14 GnRHa days in 75.70% and 24.30% required a mean ± SD (range) of 10 ± 4 (7–28) additional days to achieve complete suppression. In a standardized long GnRHa protocol, prolonging desensitization to achieve complete ovarian suppression does not affect the outcome in terms of pregnancy rate, oocytes retrieved, cycle cancelation, implantation rate, quality of embryos and live birthrates.576
 
Oral Contraceptive Pretreatment before Suppression by GnRH Agonist
Administration of a gonadotropin-releasing hormone analog (GnRHa) causes a flare effect leading to formation of ovarian cysts that can be functional and can impair downregulation and stimulation. Hence, they must be treated before commencement of stimulation. Oral contraceptives (OC) prevent the formation of ovarian cysts during GnRHa administration through a dual effect of pituitary suppression and ovarian protection. OC may be given for 14 days prior to downregulation. Pretreatment with an OC abolishes ovarian cyst formation, shortens the time required to achieve pituitary suppression, and decreases gonadotropin requirements without having a negative effect on pregnancy rates.6
 
Short Protocol
77
 
Advantages of Short Protocol
  1. Better for older patients and poor responders as it causes greater follicular recruitment.
  2. The flare effect offers an advantage in hypogonadotropin hypogonadism.
  3. Shorter protocol
  4. Less expensive.
  5. Less chances of ovarian hyperstimulation because of lower E2 levels.
 
Disadvantages of Short Protocol
In PCOS it causes irregular growth of follicle and is not helpful where LH levels are raised.
 
Ultrashort Protocol
This differs from the short protocol in discontinuing the agonist once it has stimulated the flare. This reduced the doses of both drugs. However, this protocol had poor pregnancy rates as compared to long protocol because of premature LH surge.
 
Which Protocol to Use?
The pregnancy rate was found to be higher when GnRHa was used in a long protocol as compared to a short or ultrashort 78protocol but there was no difference in live birthrate. There was no evidence of a difference in the outcomes amongst various long protocols, nor that stopping or reducing GnRHa at the start of stimulation was associated with a reduced pregnancy rate.7
 
Disadvantages of GnRH Agonists
  1. Increased time for stimulation in the long protocol.
  2. Short protocol may add to increase in premature LH surge.
  3. Luteal phase support is needed.
  4. Increased cost due to increased requirement of gonadotropin.
  5. It may cause hyperstimulation due to flare response in luteal phase in the long protocol leading to high estradiol levels and ovarian cyst.8
 
Advantages of GnRH Agonists
  1. Decreases the need for close monitoring to detect spontaneous LH surge.
  2. Less cycle cancelation.
  3. Better response.
  4. More flexible schedule.
  5. Higher oocyte recovery and pregnancy rate.
 
Side Effects and Risks of GNRH Agonists
  1. Ovarian cyst: It is seen in 14 to 29%, more with the short protocol.
  2. Ovarian hyperstimulation syndrome: The increased incidence of OHSS is due to increased pregnancy rate and higher doses of gonadotropins being used.
  3. Luteal phase defect.
  4. Transient neurological disturbances like parasthesia or headaches occur in 5%.79
 
GNRH Agonist for Ovulation Trigger
In patients at risk to develop OHSS, the only option available earlier was to withhold the ovulatory dose of human chorionic gonadotropin leading to canceled cycles. GnRH agonist, when administered in a single dose, bring about the LH surge and triggers ovulation like hCG, because of its flare effect.
Dose: Leuprolide acetate: 1 mg given either subcutaneously as a single, or two doses 12 hours apart can act as an ovulation trigger.
 
Advantages
  1. Its short duration of action is more physiological unlike extended surge with the use of hCG.
  2. Decrease in multiple pregnancy.
  3. Prevents OHSS as it has shorter duration of action compared to hCG. Luteotropic action is prolonged in hCG administration leading to development of multiple corpora lutea and supraphysiological levels of E2.
  4. Can be used along with antagonist.
 
Disadvantages
  1. Cannot be used in cases of hypogonadotropic hypogonadism.
  2. Not used for cases downregulated with GnRH agonist as the flare effect of LH release by a single dose of the agonist will not occur because of pituitary downregulation, making ovulation trigger ineffective.
  3. It causes pituitary desensitization lead lowered pregnancy and live birth- rates. Hence, luteal support is necessary in these women. Triggering final oocyte maturation with GnRH agonist instead of hCG in IVF cycles dramatically decreases luteal levels of inhibins, reflecting significant 80inhibition of the corpus luteum function. This effect may explain, at least in part, the mechanism of ovarian hyperstimulation syndrome prevention by the use of GnRH agonist.9 A recent Cochrane review 2011 has not recommended that GnRH agonists be routinely used as a final oocyte maturation trigger in fresh autologous cycles because of lowered live birthrates and ongoing pregnancy rates. An exception could be made for women with high risk of OHSS, after appropriate counseling.10 A defective corpus luteum function resulting from the relatively short endogenous luteinizing hormone surge may be detrimental to endometrial receptivity. Adequate estradiol and progesterone supplementation in the luteal phase and the first trimester is recommended. An alternative approach is the use of adjuvant low-dose human chorionic gonadotropin, although caution should be exercised in view of the associated risk of OHSS development.11 After modified luteal support there is now a non-significant difference of 6% in delivery rate in favor of hCG triggering.12
 
GnRH Antagonists
Antagonists act by competitive inhibition of GnRH receptors preventing the native GnRH from exerting its stimulatory effect on the pituitary cells, resulting in rapid decline in LH and FSH lasting for 10 to 100 hours. There is no stimulatory phase unlike the GnRH agonists. Due to competitive nature of action this effect is dose-dependant and depends on the equilibrium between endogenous GnRH and the antagonists. Their action is easily reversible. Antagonists neither deplete the LH and FSH stores nor inhibit their synthesis.81
 
Contraindications
  1. Hepatic dysfunction.
  2. Renal dysfunction.
  3. Hypersensitivity to GnRH analogs.
 
Protocols
Antagonist can be given in two ways:
  1. Lubeck protocol (Multidose protocol): Gonadotropins are started as usual. When follicle reaches 14 mm or on a fixed day of protocol, antagonist is added at a dose of 0.25 mg/day until the day before ovulation (Fig. 5.1). This protocol can be either fixed or flexible.
    1. Fixed Protocol – Daily injections of small doses initiated on a fixed day of stimulation till hCG administration
    2. Flexible Protocol – Daily injections of small doses initiated depending on the size of the dominant follicle (14 mm) or on estradiol levels till hCG administration.
  2. French protocol (Single dose protocol): Gonadotropins are started as usual. Antagonist is given in a single dose of 3 mg when E2 is about 150 to 200 pg/mL and follicular size is 14 mm (Fig. 5.2).
Fig. 5.1: Lubeck protocol
82
Fig. 5.2: French protocol
 
Single Versus Multiple Dose GnRH Antagonist Protocol
Advantage of single dose protocol was lesser injections but if hCG needs to be delayed additional daily doses are given. About 10% women require additional doses. Cochrane review 2002 has concluded that both protocols were equally effective in preventing premature LH surge.13 It was seen that the single dose protocol may lead to extreme suppression of LH but pregnancy rates were similar with both protocols.14
 
Fixed Versus Flexible Antagonist Administration
In an analysis of three studies, a flexible GnRH antagonist protocol was compared to the fixed protocol. In stimulated cycles it was seen that intense ovarian response with more number of follicles led to an early rise in estradiol. Thus, threshold levels of estradiol, which initiates a LH surge, are reached earlier before follicles reach an optimum size. In these cases the flexible protocol which is dependent on the size of the follicle on ultrasound to start antagonist in order to suppress surge, may no longer be accurate to determine time of initiation of GnRH antagonist.15 This observation might explain the observed lower efficacy of the flexible protocol 83compared with a fixed protocol in a meta-analysis of four studies.16
Hence, for patients with a profound ovarian response, early initiation of the GnRH antagonist may be needed.
 
Which GnRH Antagonist is to be used?
Cetrorelix and ganirelix both effectively prevented LH surge. However, cetrorelix required significantly fewer injections, increasing patient convenience.14
 
Should FSH dose be Increased in Antagonist Cycle ?
Although with antagonist cycles, gonadotropin dose needed is lower as there is no pituitary suppression; however, a reduced oocyte recovery was notice. An initial higher dose of FSH gave a higher oocyte recovery but there was no difference in pregnancy rates.17 It was seen that increasing dose of FSH or hMG after starting antagonist did not increase pregnancy rates.18,19
 
Role of Oral Contraceptive Pill Pretreatment in Ovarian Stimulation with GnRH Antagonists for Cycle Scheduling
Pretreatment with an oral contraceptive (OC) in antagonist cycle has been suggested to allow greater control over patient response rate and to avoid follicular asynchrony. Scheduling of cycle is no longer based on menstruation but on discontinuation of OCP. A meta-analysis proved that no increase in pregnancy rate was seen with OC.20 However, it has been associated with increased duration of treatment and higher doses of gonadotropin.21 In a recent study OCP (0.030 ethinyl E(2)/0.15 desogestrel) for 12–16 days, and controlled ovarian hyperstimulation with GnRH antagonist was started on day 5 after OCP treatment. On comparison with long 84protocol no differences were observed in the fertilization rates (68.1% vs 64.8%), total number of embryos obtained (5.9 vs 6.2), mean number of embryos transferred (1.8 vs 1.8), implantation rate (36% vs 39%), miscarriage rate (8.9% vs 17%), ongoing pregnancy rate (47.8% vs 53.9%), or live birthrate (44.3% vs 47%).22
 
LH Supplementation with GnRH Antagonist
An abrupt suppression of endogenous LH by GnRH antagonist occurs in the mid-follicular phase, at a critical stage for follicular development. However, studies have shown no increase in pregnancy rate with LH supplementation or increase of hMG dose on initiation of antagonist.19,23 The decision to add LH must be individualized. It has been seen that the direction and rate of change in LH concentrations are the important factors governing the follicular unit development, not the LH concentration itself.24 It was found in a study that in 12–14% of downregulated patients the initial response to FSH is suboptimal (in terms of follicular growth and estradiol rise) and their day 8 LH concentration decreased from 1.2 to 0.7. It was suggested that these patients are the candidates for LH supplementation. Normal responders increased their mean LH concentrations from 1.5 to 4.3 after 8 days of stimulation. It was suggested that the follicular unit is sensitive not necessarily to the current concentration of LH, but rather to the dynamics of the change in these concentrations and hence, LH supplementation must be individualized.25 In women on antagonist-based cycles for the first time, it is preferable to add recombinant LH or partly switch to hMG on the day of antagonist administration.26 85
 
Luteal Support
It is seen that pituitary suppression may continue into luteal phase with poor development of endometrium. Hence, luteal support is a must and has shown to improve pregnancy rates.
 
Advantages of GnRH Antagonists over GnRH-agonist in ART
  1. Short, simple and convenient method of stimulation, which is well tolerated by the patient.
  2. There is immediate suppression with no stimulatory phase. So no ovarian cyst formation takes place as in agonist cycle.
  3. There are no symptoms of estrogen deprivation.
  4. Minimal local reactions.
  5. Decreased risk of OHSS: A recent Cochrane (2011) showed a decreased risk of OHSS but similar live birthrates compared to GnRH agonists.27
  6. Clinical results of IVF cycles are comparable in both (Cochrane 2011)27
  7. Decrease in the overall cost of treatment.
  8. Immediate reversibility.
  9. Reduced dose of gonadotropins.
  10. Effect on endometrium: Simón et al (2005) observed that the endometrial development after GnRH antagonist mimics the natural endometrium more closely than after GnRH agonist.28
  11. LH levels and embryo quality: GnRH antagonists administration during the late follicular phase resulted in lower serum LH levels and better embryo quality in comparison to GnRH agonists.29
86
Studies have concluded that antagonists should be the first choice in IVF treatment, with less of the complications and risks of controlled ovarian hyperstimulation and an acceptable success rate.29
 
Disadvantages of Antagonists
  1. Easy patient scheduling is lacking, because of the reliance on spontaneous menstrual cycles.
  2. Lack of stimulatory effects on folliculogenesis typical of GnRH agonist regimens.
  3. Need to replace LH if recombinant FSH is used.
 
Gonadotropin-Releasing Hormone
It is mainly used in WHO group I anovulatory women but can be used in PCOS patients. The advantages are that significant follicular monitoring is not needed and it has less risk for OHSS and multiple pregnancy. It can be given either subcutaneously in a dose of 20 μg or intravenously 5 μg at 90 min interval through a portable programmable mini pump worn by the patient throughout. If there is no response in weekly estradiol level the dose is increased by increments of 5 μg. Luteal phase 87support can be given by continuation of the pump. Problems encountered are malfunction of the pump and local effects like thrombophlebitis, cellulitis, urticaria or anaphylaxis. Ovulation rates of 90%, conception rates of 20 to 30% per cycle and cumulative pregnancy rate of 80 to 90% after 12 months are seen in WHO group 1 women.31 In PCOS cumulative pregnancy rate is 30 to 40%. Abortion occurs in 20% cases and multiple pregnancies in 5%.
Thus, to conclude the GnRH antagonist reduces the dose and duration of gonadotropin treatment as it does not nullify FSH or LH secretion but interrupts the premature LH surge. It has the disadvantage of loss of easy patient rescheduling because of reliance on natural recruitment of follicles, lack of stimulatory effect on folliculogenesis unlike GnRH agonists and need for replacement of LH if recombinant FSH is used.
 
References
  1. Anik ST, McRae G, Narenberg C, Worden A, Foreman J, Hwang JY, et al. Nasal absorption of naferelin acetate, the decapeptide (D-Nal{2}6) LHRH, in rhesus monkeys. J Pharm Sci. 1984;73:684–5.
  1. Albuquerque LE, Saconato H, Maciel MC. Depot versus daily administration of gonadotrophin releasing hormone agonist protocols for pituitary desensitization in assisted reproduction cycles. Cochrane Database Syst Rev. 2005 Jan 25;(1):CD002808.
  1. Akagbosu FT. The use of GnRH agonists in infertility. In: Brinsden R (Ed). A Textbook of In Vitro Fertilization and Assisted Reproduction (2nd ed): London: Parthenon Publishing;  1999:83-9.
  1. Bloch M, Azem F, Aharonov I, Ben Avi I, Yagil Y, Schreiber S, et al. GnRH-agonist induced depressive and anxiety symptoms during in vitro fertilization-embryo transfer cycles. Fertil Steril. 2011;95(1):307–9.
  1. Dessolle L, Ferrier D, Colombel A, Fréour T, Jean M, Barrière P. Prolonging GnRH-agonist to achieve ovarian suppression does not compromise the results of a long protocol. Eur J Obstet Gynecol Reprod Biol. 2011;159(1):111–4.
  1. Biljan MM, Mahutte NG, Dean N, Hemmings R, Bissonnette F, Tan SL. Effects of pretreatment with an oral contraceptive on the time required to achieve pituitary suppression with gonadotropin-releasing hormone analogues and on subsequent implantation and pregnancy rates. Fertil Steril. 1998;70(6):1063–9.
  1. Maheshwari A, Gibreel A, Siristatidis CS, Bhattacharya S. Gonadotrophin-releasing hormone agonist protocols for pituitary suppression in assisted reproduction. Cochrane Database Syst Rev. 2011 Aug 10;(8):CD006919.
  1. Depenbusch M, Diedrich K, Griesinger G. Ovarian hyperresponse to luteal phase GnRH-agonist administration. Arch Gynecol Obstet. 2010;281(6):1071–2.
  1. Nevo O, Eldar-Geva T, Kol S, Itskovitz-Eldor J. Lower levels of inhibin A and pro-alpha C during the luteal phase after triggering oocyte maturation with a gonadotropin-releasing hormone agonist versus human chorionic gonadotropin. Fertil Steril. 2003;79(5):1123-8.
  1. Youssef MA, Van der Veen F, Al-Inany HG, Griesinger G, Mochtar MH, Aboulfoutouh I, et al. Gonadotropin-releasing hormone agonist versus HCG for oocyte triggering in antagonist assisted reproductive technology cycles. Cochrane Database Syst Rev. 2011 Jan 19;(1):CD008046.
  1. Engmann L, Benadiva C. Ovarian hyperstimulation syndrome prevention strategies: Luteal support strategies to optimize pregnancy success in cycles with gonadotropin-releasing hormone agonist ovulatory trigger. Semin Reprod Med. 2010;28(6):506–12.
  1. Humaidan P, Kol S, Papanikolaou EG. Copenhagen GnRH Agonist Triggering Workshop Group. GnRH agonist for triggering of final oocyte maturation: time for a change of practice? Hum Reprod Update. 2011;17(4):510–24.
  1. Al-Inay H, Aboulghar M. GnRH antagonist in assisted reproduction: a Cochrane review. Hum Reprod Update. 2002;17(4):874–85.
  1. Wilcox J, Potter D, Moore M, Ferrande L, Kelly E. CAP IV Investigator Group. Prospective, randomized trial comparing cetrorelix acetate and ganirelix acetate in a programmed, flexible protocol for premature luteinizing hormone surge prevention in assisted reproductive technologies. Fertil Steril. 2005;84(1):108–17.
  1. Al-Inany HG, Aboulghar M, Mansour R, Serour GI. Optimizing GnRH antagonist administration: meta-analysis of fixed vs flexible protocol. Reprod Biomed Online. 2005;10:567–70.
  1. Tarlatzis BC, Fauser BC, Kolibianakis EM, Diedrich K, Rombauts L, Devroey P. GnRH antagonists in ovarian stimulation for IVF. Hum Reprod Update. 2006;12:333–40.
  1. Out HJ, Rutherford A, Fleming R, Tay CCK, Trew G, Ledger W, et al. A randomized, double-blind, multicentre clinical trial comparing starting doses of 150 and 200 IU of recombinant FSH in women treated with the GnRH antagonist ganirelix for assisted reproduction. Hum Reprod. 2004;19:90–5.
  1. Propst AM, Bates GW, Robinson RD, Arthur NJ, Martin JE, Neal GS. A randomized controlled trial of increasing recombinant follicle-stimulating hormone after initiating a gonadotropin-releasing hormone antagonist for in vitro fertilization-embryo transfer. Fertil Steril. 2006;86(1):58–63.
  1. Aboulghar MA, Mansour RT, Serour GI, Al-Inany HG, Amin YM, Aboulghar MM. Increasing the dose of human menopausal gonadotropins on day of GnRH antagonist administration: randomized controlled trial. Reprod Biomed Online. 2004;8:524–7.
  1. Griesinger G, Venetis CA, Marx T, Diedrich K, Tarlatzis BC, Kolibianakis EM. Oral contraceptive pill pretreatment in ovarian stimulation with GnRH antagonists for IVF: a systematic review and meta-analysis. Fertil Steril. 2008;90(4):1055–63.
  1. Bendikson K, Milki A, Speck-Zulak A, Westphal L. Comparison of GnRH antagonist cycles with and without oral contraceptive pill pretreatment in poor responders. Fertil Steril. 2003;80(Suppl. 3):s188.
  1. Garcia-Velasco JA, Bermejo A, Ruiz F, Martinez-Salazar J, Requena A, Pellicer A. Cycle scheduling with oral contraceptive pills in the GnRH antagonist protocol vs the long protocol: a randomized, controlled trial. Fertil Steril. 2011;96(3):590–3.
  1. Griesinger G, Schultze-Mosgau A, Dafopoulos K, Schroeder A, Schroer A, von Otte S, Hornung D, et al. Recombinant luteinizing hormone supplementation to recombinant follicle stimulating hormone induced ovarian hyperstimulation in the GnRH antagonist multiple-dose protocol. Hum Reprod. 2005a;20:1200-06.
  1. Huirne JA, van Loenen ACD, Schats R, McDonnell J, Hompes PGA, Schoemaker Joop, et al. Dose-finding study of daily GnRH antagonist for the prevention of premature LH surges in IVF/ICSI patients: optimal changes in LH and progesterone for clinical pregnancy. Hum Reprod. 2005;20:359–67.
  1. De Placido G, Alviggi C, Perino A, et al. Recombinant human LH supplementation versus recombinant human FSH (rFSH) step-up protocol during controlled ovarian stimulation in normogonadotrophic women with initial inadequate ovarian response to rFSH. A multicentre, prospective, randomized controlled trial. Hum Reprod. 2005;20:390–6.
  1. Kol S. To add or not to add LH: consideration of LH concentration changes in individual patients. Reprod BioMed Online. 2005;11:664–6.
  1. Al-Inany HG, Youssef MAFM, Aboulghar M, Broekmans FJ, Sterrenburg MD, Smit JG, et al. Gonadotrophin-releasing hormone antagonists for assisted reproductive technology. Cochrane Database of Systematic Reviews. 2011, Issue 5. Art. No.: CD001750. doi: 10.1002/14651858.CD001750.pub3
  1. Simon C, Oberye J, Bellver J, Vidal C, Bosch E, Horcajadas JA, et al. Similar endometrial development in oocyte donors treated with either high- or standard-dose GnRH antagonist compared to treatment with a GnRH agonist or in natural cycles. Hum Reprod. 2005;20(12):3318–27.
  1. Xavier P, Gamboa C, Calejo L, Silva J, Stevenson D, Nunes A, et al. A randomised study of GnRH antagonist (cetrorelix) versus agonist (buserelin) for controlled ovarian stimulation: effect on safety and efficacy. Eur J Obstet Gynecol Reprod Biol. 2005;120:185–9.
  1. Tarlatzis BC, Kolibianakis EM, Griesinger G, et al. GnRH antagonists in ovarian stimulation for IVF. Hum Reprod Update. 2006;12:333–40.
  1. Ghosh C, Buck G, Priore R, Wende JW, Severino M. Follicular response and pregnancy among infertile women undergoing ovulation induction and intrauterine insemination. Fertil Steril. 2003;80:328–35.

Mild Ovarian Stimulationchapter 6

Surveen Ghumman
The basis of successful assisted reproduction is appropriate ovarian stimulation. Since IVF procedures have not produced a 100% result, ovarian stimulation has become the compensatory mechanism to improve results by increasing the number of oocytes retrieved; thus, providing the choice of the best quality embryos for transfer.
In the conventional protocol downregulation is started in the luteal phase and continued into next cycle, involving high doses and prolonged administration of FSH. It has high chances of complications like ovarian hyperstimulation syndrome (OHSS). The prolonged treatment with high cost results in larger dropout rates.
In the recent years, mild protocols have been introduced that aim at a low stimulation that gives acceptable results with minimal risks. International Society of Mild Approaches in Assisted Reproduction (ISMAAR) defines a “mild” IVF cycle either as (a) a stimulation regimen in which gonadotropins are administered at a lower-than-usual dose and/or for a shorter duration throughout a cycle in which GnRH antagonist 93is given as co-treatment, or (b) a stimulation in which oral compounds (e.g. antiestrogens) are used either alone or in combination with gonadotropins and GnRH - antagonists.1
 
Role of GnRH Antagonist in Mild Stimulation
With use of antagonists downregulation is not needed. The IVF cycle can start with an undisturbed early follicular phase recruitment of follicles by an endogenous FSH rise that occurs in a natural menstrual cycle. The endogenous inter-cycle FSH rise is taken advantage of rather than suppressed. FSH is added on day 5 to continue FSH elevation and extend the FSH window, for many follicles to develop. This limits the duration and dose of FSH administration. A premature LH surge may occur with rising estradiol levels that act through positive feedback loop. GnRH, antagonist has an immediate action blocking the pituitary when estradiol levels start rising and approach threshold levels at which an LH surge can occur. The action of GnRH antagonists is immediate suppression of the pituitary release of gonadotropins and a rapid reversibility of normal gonadotropin secretion when the drug is withdrawn.
 
Protocols
Antagonist can be given in three ways:
  1. Single large dose on, usually sixth day of stimulation with gonadotropins
  2. Fixed Protocol: Daily injections of small doses initiated on a fixed day of stimulation till hCG administration.
  3. Flexible Protocol: Daily injections of small doses initiated depending on the size of the dominant follicle or on estradiol levels till hCG administration.
In an analysis of three studies, a flexible GnRH antagonist protocol was compared to the fixed protocol. In stimulated 94cycles it was seen that intense ovarian response with more number of follicles led to an early rise in estradiol. Thus, threshold levels of estradiol that initiates a LH surge are reached earlier before follicles reach an optimum size. In these cases the flexible protocol, which is dependent on the size of the follicle on ultrasound to start antagonist to suppress surge, may no longer be accurate to determine time of initiation of GnRH antagonist.2 This observation might explain the observed lower efficacy of the flexible protocol compared with a fixed protocol in a meta-analysis of four studies.3 Hence, for patients with a profound ovarian response, early initiation of the GnRH antagonist may be needed.
 
Clomiphene, Gonadotropins and GnRH Antagonist
The second mild regime includes using clomiphene 100 mg, delayed low dose gonadotropin and a flexible GnRH antagonist administration for ovarian stimulation protocol. Pregnancy rates comparable to the standard stimulation regimens were obtained, with a significant reduction in the total dose of gonadotropin and hence, the cost. The number of recovered oocytes, obtained embryos, transferred embryos, peak of estradiol on the day hCG administration and OHSS were significantly higher in conventional group but there were no significant difference in clinical pregnancy rate and ongoing pregnancy rate between two groups.4,5
 
Mild vs Conventional Protocols
 
Dose of FSH
The mild form used lower dose FSH for a shorter period. Mean dose of FSH used was 1183 IU in the mild group versus 1836 IU for the conventional group (Table 6.1).695
Table 6.1   Comparison of mild and conventional protocols
Mild
Conventional
Low oocyte recovery
Pregnancy rate per embryo transfer – Good
Pregnancy rate per embryo transfer – Poor
Optimal outcomes
5 oocyte recovery
10 oocyte recovery
Poor outcome
> 8 oocytes recovered
> 18 and < 4 oocytes recovered
Proportion of abnormal and mosaic embryos
73%
55%
Dose of FSH used
1836
1183
 
Correlation of Number of Oocytes Recovered with Pregnancy Rate
A higher pregnancy rate is achieved when there is a moderate ovarian response. The highest ongoing pregnancy rate per embryo transfer of 30.7% in mild stimulation is observed where five oocytes were obtained but a 28.5% ongoing pregnancy rate with a median of 10 oocytes is seen in the conventional protocol. A sharp decline in implantation rates was seen with retrieval of more than 8 oocytes in the mild stimulation protocol. These differences between the two stimulation regimens were statistically significant (P = 0.045).6
Excessive follicles do not lead to a higher pregnancy rate. In both protocols, the number of pregnancies following the retrieval of 18 oocytes or more was very low (Table 6.1). Hence, when a few oocytes are recruited they are better quality oocytes from a homogenous group that lead to a higher pregnancy rate. Mild stimulation protocol simulates 96a natural cycle where the natural selection of good quality oocytes remains. It decreases detrimental effect of ovarian stimulation on the growing follicle. By delaying the initiation of ovarian stimulation to the mid-follicular phase, exogenous FSH may only stimulate the naturally selected most mature follicles giving rise to the best quality oocytes.
A second study reinforced this, showing that those patients where stimulation was started on day 5, fewer cycles were characterized by a total fertilization failure or by abnormal embryo development. After stronger ovarian stimulation, only 7% of the patients who retrieved less than 5 oocytes conceived, whereas after “mild” stimulation 67% of these patients conceived.7 The retrieval of a less number of oocytes gives higher chance of ongoing pregnancy per embryo transferred in mild stimulation but poor results in conventional protocols. The implication of a poor ovarian response in the conventional protocol usually means a low ovarian reserve resulting in poor IVF outcomes whereas a poor response in mild ovarian stimulation is probably a normal response with natural selection of follicle with best receptor endowment.
Overall, no differences were found among the two stimulation protocols as far as the pregnancy rate per started cycle was concerned because of the higher number of oocytes retrieved in the conventional group, more number of embryos were available to select the best. No significant difference in the quality of the best transferred embryo was observed among the two groups.7 The higher pregnancy rate per oocyte retrieved seen with mild protocol may be due to better development of embryo that has been naturally selected with higher chance of being chromosomally normal. Secondly, detrimental effects seen with very high levels of hormones on endometrium are absent, as ostradiol levels are lower than the conventional protocol.97
 
Endometrial Factor
The conventional regime has supra-physiological circulating estradiol levels that have a well-documented negative impact on the developmental and implantation potential of human embryos.8 A higher endometrial receptivity is seen in the natural cycle and since the “mild” stimulation regimen, like the natural cycle has a lower peak estradiol levels, implantation is thought to be better.9,10
 
Chromosomal Abnormality with Mild Ovarian Stimulation
In a recent study, embryos were biopsied on day 3 when at least six blastomeres were present after stimulation with both regimes. One or two cells were removed for a fluorescence in situ hybridization procedure. Both regimens finally generated the same number (1.8/cycle) of chromosomally normal embryos. As the conventional protocol had higher number of oocytes per cycle, overall abnormality rates (abnormal and mosaic embryos) were 55% following mild and 73% following conventional ovarian stimulation (Table 6.1). However, the proportion of mosaic embryos per patient was more significantly increased following conventional ovarian stimulation (65% vs 37%; P = 0.004). This observation indicates that the increase in abnormal embryos is mainly due to an increase in mitotic segregation errors in early embryonic cleavage divisions.11 Ovarian stimulation might disrupt mechanisms involved in maintaining accurate chromosome segregation leading to differences in rates of mosaic embryos.12 Hence, a lower embryo aneuploidy rate was present following mild stimulation. Several animal studies have supported this. Increased incidences of morphological and chromosomal abnormalities have been observed in mouse oocytes after exposure to high doses of gonadotropins during in vitro maturation of oocytes.1398
Ovarian stimulation strategies should avoid maximizing oocyte yield, but aim at generating a sufficient number of chromosomally normal better quality embryos by reduced interference with ovarian physiology.
 
Cumulative Pregnancy Rate
In another study of the “mild” treatment group the pregnancy rate per cycle was significantly lower in the “mild” stimulation group vs the conventional group (17.6% vs 28.6%, p < 0.0001). However less dose of FSH was used since it was cheaper, patient discomfort was less resulting in higher acceptance of patient for a second IVF. The one year cumulative live birthrate was 43.4% with the mild protocol, 44.7% with the standard regimen.14 Hence, there was not much difference in the cumulative pregnancy rate.
 
Multiple Pregnancy
The second advantage was that twin pregnancy was 0.5% in comparison to conventional protocol which had a rate of 13.1%.14 This also contributed to the cost effectivity of the regime as multiple pregnancy requires a more intensive monitoring and subsequent neonatal care.
 
Cryopreservation
The pooled data from a meta-analysis shows that the ongoing pregnancy rate per started cycle sorts out to be 15% in the “mild” group and 29% in the classical group clearly showing conventional protocol to be more effective. Also, the conventional protocol will give excess embryos for a subsequent frozen embryo transfer that will get down both the cost and patient discomfort and increase the overall IVF pregnancy chance per oocyte pick-up by approximately 10–15%.15 99
 
Cancelation
The other factor that decreases the effectiveness of “mild” strategy is the relatively high rate of cycle cancelation due to mono- or bifollicular response (around 15–20%). It is recommended that next cycle stimulation should be started earlier on day 2. Ovarian aging and high BMI have been identified as relevant variables to predict the risk of insufficient response to “mild” stimulation, and a predictive model has been developed in order to minimize the need of canceling the cycle.16
 
Poor Ovarian Reserve and Mild Ovarian Stimulation
The standard approach to women who are poor responders is based on starting with high doses of stimulation of 300 FSH IU and going up to 600 FSH IU. However, IVF outcomes in terms of pregnancy rates with 225 FSH UI/day and those receiving 450 UI/day was shown to be similar, despite the latter obtained more oocytes.17,18
High gonadotropin doses lower the cycle cancelation rate, but the likelihood of clinical pregnancy and live birthrate have been observed to reduce and to increase the risk of spontaneous miscarriage because of its adverse effects on endometrium and oocyte and embryo quality generated from follicles that were rescued from atresia if a natural cycle had taken place.19
In patients with poor ovarian reserve, the choice of a mild stimulation protocol instead of a classical, high dose regimen could be particularly indicated. Although these patients have a very low risk of OHSS, the quality of both their oocytes and their endometrium may be better when a smoother stimulation approach is used.
A combination of clomiphene citrate (CC) plus gonadotropins and GnRH antagonists has been proposed 100as a “mild” stimulation alternative for poor responders. The treatment protocol consisted of a daily dose of clomiphene citrate 100 mg for 5 days and gonadotropin injections daily from cycle day 4 onward. Cetrorelix, 0.25 mg/day, was started when the leading follicle reached 14 mm. Induction of ovulation was triggered with human chorionic gonadotropin. The combination of clomiphene with gonadotropins may counterbalance its undesired antiestrogenic effect on the endometrium and clomiphene; at the same time their effects may reduce the amount of gonadotropins required, because of the combined synergistic effect on the ovary. A significantly higher blastocyst development rate and a very good (41.2%) ongoing pregnancy rate was found with this regimen.20
In CC/Gn/GnRH antagonist cycles, it was seen that in some cases LH was profoundly suppressed by GnRH antagonist and the circulating level of LH were less than one third at the time of hCG than it was at the beginning of stimulation. In these cases, both the pregnancy (18% vs 39%) and implantation rates are significantly reduced. This suggests that in these patients medications containing LH or hCG rather than FSH alone should be associated with CC in this kind of protocol.21
 
Risk of Ovarian Hyperstimulation Syndrome (OHSS)
Severe OHSS has an incidence of 1–3% in IVF programs involving standard ovarian stimulation regimens. The incidence of severe OHSS is significantly lower when GnRH antagonists are used instead of agonists probably due to the smaller cohort of recruited follicles and to the lower circulating estradiol levels during ovarian stimulation.22
The risk of developing severe OHSS is further significantly reduced using “mild” stimulation regimens. In one study, the incidence of OHSS was 1.4% with the mild protocol and 3.7% with the long protocol.11 While in another study, 101there was no case of OHSS in the group treated with “mild” stimulation versus 6% in the group treated with conventional stimulation.4 Further, the risk of severe OHSS is reduced if ovulation trigger is elicited using a single dose of GnRH agonist instead on hCG; this is possible if a stimulation with GnRH antagonists has been applied.23
 
Emotional Stress
Emotional stress represents a well-known negative side effects associated with IVF treatment, and probably one of the most important reasons for dropping out of the program.
In the mild stimulation, a lower incidence of side effect faced with a conventional protocol; maybe the reason for a lower dropout rate and patients going in for a repeat IVF sooner as the psychological burden is lower. This leads to a higher cumulative success rate.24
However, a lower pregnancy chance with mild stimulation may itself cause psychological problems associated with failure. Also, the increased number of repeat oocyte retrievals with this protocol is a stressing event.
 
Economical Costs
A milder ovarian stimulation is associated with a lower medication consumption per cycle thus lowering the cost. The balance between lower costs and lower “per cycle” results must be kept in mind when taking a decision. Also, the frequent need to repeat treatment actually increases cost as amount of FSH required to achieve one pregnancy goes up. However, in a recent study it was seen that although there is a significantly increased average number of IVF cycles (2.3 versus 1.7), lower average total costs over a 12-month period (8333 euro versus 10,745 euro) were observed using the mild strategy. Despite an increased mean number of IVF 102cycles within 1 year, from an economic perspective, the mild treatment strategy is more advantageous per term live birth.25
 
Lower Effectiveness
The lower effectiveness of IVF procedure can also become a problem for IVF clinics choosing the mild strategy, who will compete on the market with clinics following classical stimulation concepts. If the clinic loses patients for the lower “per cycle” effectiveness of its IVF program, it could be forced to go back to classical stimulations, or alternatively, to increase prices, finally weighting on patients’ budget.
In conclusion, lower number of oocytes retrieved during mild stimulation is associated with favorable pregnancy outcomes as there may be natural selection. As the mild stimulation did not show better pregnancy rates compared with a conventional stimulation protocol with GnRH agonist co-treatment, the benefits of low cost should be balanced with the decrease in pregnancy rate per cycle. With current recommendation of maximum two embryo transfer, there seems to be no need of aggressive stimulation to obtain large number of oocytes at the cost of good quality oocyte selection and adequately primed endometrium.
 
References
  1. Nargund J, Fauser BCJM, Macklon NS, Ombelet W, Nygren K, Frydman R. The ISMAAR proposal on terminology for ovarian stimulation for IVF. Hum Reprod. 2007;11(14):2801–04.
  1. Al-Inany HG, Aboulghar M, Mansour R, Serour GI. Optimizing GnRH antagonist administration: meta-analysis of fixed vs flexible protocol. Reprod Biomed Online. 2005;10:567–70.
  1. Tarlatzis BC, Fauser BC, Kolibianakis EM, Diedrich K, Rombauts L, Devroey P. GnRH antagonists in ovarian stimulation for IVF. Hum Reprod Update. 2006;12:333–40.
  1. Karimzadeh MA, Ahmadi S, Oskouian H, Rahmani E. Comparison of mild stimulation and conventional stimulation in ART outcome. Arch Gynecol Obstet. 2010;281(4):741–6.
  1. Williams SC, Gibbons WE, Muasher SJ, Oehninger S. Minimal ovarian hyperstimulation for in vitro fertilization using sequential clomiphene citrate and gonadotropin with or without the addition of a gonadotropin-releasing hormone antagonist. Fertil Steril. 2002;78(5):1068–72.
  1. Verberg MFG, Eijkemans MJC, Macklon NS, Heijnen EMEW, Baart EB, Hohmann FP, et al. The clinical significance of the retrieval of a low number of oocytes following mild ovarian stimulation for IVF: a meta-analysis. Hum Reprod Update. 2009;15:5–12.
  1. Hohmann FP, Macklon NS, Fauser BC. A randomized comparison of two ovarian stimulation protocols with gonadotropin-releasing hormone (GnRH) antagonist co-treatment for in vitro fertilization commencing recombinant follicle-stimulating hormone on cycle day 2 or 5 with the standard long GnRH agonist protocol. J Clin Endocrinol Metab. 2003;88(1):166–73.
  1. Valbuena D, Jasper M, Remohi J, Pellicer A, Simon C. Ovarian stimulation and endometrial receptivity. Hum Reprod. 1999;14(2):107–11.
  1. Check JH, Choe JK, Nazari A, Summers-Chase D. Ovarian hyperstimulation can reduce uterine receptivity. A case report. Clin Exp Obstet Gynecol. 2000;27(2):89–91.
  1. Check JH, Check ML. A case report demonstrating that follicle maturing drugs may create an adverse uterine environment even when not used for controlled ovarian hyperstimulation. Clin Exp Obstet Gynecol. 2001;28(4):217–8.
  1. Baart EB, Martini E, Eijkemans MJ, Van Ostal D, Beckers NG, Verhoeff A, et al. Milder Ovarian stimulation for in vitro fertilization reduces aneuploidy in the human preimplantation embryo: a randomised controlled trial. Hum Reprod. 2007;22(4):980–8.
  1. Hodges CA, Ilagan A, Jennings D, Keri R, Nilson J, Hunt PA. Experimental evidence that changes in oocyte growth influence meiotic chromosome segregation. Hum Reprod. 2002;17:1171–80.
  1. Roberts R, Iatropoulou A, Ciantar D, Stark J, Becker DL, Franks S, Hardy K. Follicle-stimulating hormone affects metaphase I chromosome alignment and increases aneuploidy in mouse oocytes matured in vitro. Biol Reprod. 2005;72:107–18.
  1. Hejinen EMEW, Eijkemans MJC, De Klerk C, Polinder S, Beckers NGM, Klinkert ER, et al. A mild treatment strategy for in vitro fertilization: a randomised non inferiority trial. Lancet. 2007;369(9563):743–9.
  1. Revelli A, Casano S, Salvagno F, Piane LD. Milder is better? advantages and disadvantages of “mild” ovarian stimulation for human in vitro fertilization. Reprod Biol Endocrinol. 2011;9:25–30.
  1. Verberg MF, Eijkemans MJ, Macklon NS, Heijnen EM, Fauser BC, Broekmans F. Predictors of low response to mild ovarian stimulation initiated on cycle day 5 for IVF. Hum Reprod. 2007;22(7):1919–24.
  1. Land JA, Yarmolinskaya MI, Dumoulin JC, Evers JL. High-dose human menopausal gonadotropin stimulation in poor responders does not improve in vitro fertilization outcome. Fertil Steril. 1996;65(5):961–5.
  1. Lekamge DN, Lane M, Gilchrist RB, Tremellen KP. Increased gonadotrophin stimulation does not improve IVF outcomes in patients with predicted poor ovarian reserve. J Assist Reprod Genet. 2008;25(11-12):515–21.
  1. Pal L, Jindal S, Witt BR, Santoro N. Less is more: increased gonadotropin use for ovarian stimulation adversely influences clinical pregnancy and live birth after in vitro fertilization. Fertil Steril. 2008;89(6):1694–701.
  1. Takahashi K, Mukaida T, Tomiyama T, Goto T, Oka C. GnRH antagonist improved blastocyst quality and pregnancy outcome after multiple failures of IVF/ICSI-ET with a GnRH agonist protocol. J Assist Reprod Genet. 2004;21(9):317–22.
  1. Yanaihara A, Yorimitsu T, Motoyama H, Ohara M, Kawamura T. The decrease of serum luteinizing hormone level by a gonadotropin-releasing hormone antagonist following the mild IVF stimulation protocol for IVF and its clinical outcome. J Assist Reprod Genet. 2008;25(4):115–8.
  1. Kolibianakis EM, Collins I, Tarlatzis BC, Devroey P, Griesinger G. Among patients treated for IVF with gonadotrophins and GnRH analogues is the probability of live birth dependent on the type of analogue used? A systematic review and meta-analysis. Hum Reprod Update. 2006;12a(6):651–71.
  1. Humaidan P, Papanikolaou EG, Tarlatzis BC. GnRHa to trigger final oocyte maturation: a time to reconsider. Hum Reprod. 2009;24(10):2389–94.
  1. de Klerk C, Heijnen EM, Macklon NS, Duivenvoorden HJ, Fauser BC, Passchier J, et al. The psychological impact of mild ovarian stimulation combined with single embryo transfer compared with conventional IVF. Hum Reprod. 2006;21(3):721–7.
  1. Polinder S, Heijnen EM, Macklon NS, Habbema JD, Fauser BJ, Eijkemans MJ. Cost-effectiveness of a mild compared with a standard strategy for IVF: a randomized comparison using cumulative term live birth as the primary end point. Hum Reprod. 2008;23(2):316–23.

Premature Luteinizationchapter 7

Surveen Ghumman,
Monika Gupta
Premature luteinization (PL) has been defined as the rise of progesterone on the day hCG is given. It is an important entity in women who are undergoing ovulation induction. There is no strict criterion of diagnosis and no definite etiology has been identified.
 
Diagnosis
 
Serum Progesterone on Day of hCG
Various studies have quoted a cut-off of 0.8 to 2 ng/mL.1 As more number of follicles produce more progesterone, it may be important to link ovarian response to estrogen and progesterone levels rather than absolute progesterone values. Hence, P/E2 ratio may be better in detecting PL. A level of more than 1 is thought to differentiate between progesterone secretion from a dysmature follicle as occurs in PCOS from that of a mature healthy follicle.2 A study showed that PL seems unrelated to preovulatory luteinizing hormone (LH) elevation and LH/hCG content of gonadotropins and could be associated with poor ovarian response and the presence of dysmature follicles.3107
 
Progesterone/Estradiol Ratio on Day of hCG
Cycles with elevated P/E2 ratios are associated with lower clinical pregnancy and live birth rates, which decrease further as the P/E2 ratio rises. P/E2 ratio improves the prediction of IVF outcome when compared to serum P levels alone.4
 
Ultrasound Appearance
Collaborative ultrasound appearance of the follicle shows a thickened follicular wall and appearance of irregular echogenic structures within the follicle.
 
Incidence
It varies in various studies from 13 to 71%, using P only to define PL. It was found that 41% had a P/E2 ratio.2 The incidence varies with different stimulation protocols. It is maximum with flare protocol being 85%.5 It is 54.7% in women undergoing COH with CC hMG and a single 2.5 mg dose of the GnRH antagonist, cetrorelix.6 With GnRH agonist protocol incidence varied from 5 to 35% and with antagonist from 20 to 35%.7,8
 
Causes
 
Increased Levels of hCG Accumulated with hMG Administration
It has been seen that there are higher levels of hCG in women with PL suggesting that the LH activity in hMG is responsible for PL. Hence, in these cases recombinant or highly purified FSH may be given.9108
 
Increased LH Levels
Although GnRH agonists suppress LH, it has been seen that in some cases suppression may be incomplete. LH levels may be enough to stimulate granulose cells to produce progesterone but not enough to cause rupture of follicle.
 
Increased LH Sensitivity of Granulose Cells to FSH
It is postulated that there is a higher sensitivity of LH receptors of granulose cells to FSH, which may be due to increased estradiol levels.
 
Increased LH Sensitivity in Poor Responders
PL is seen more often in poor responders and raised progesterone may be because of adversely developing cumulus oocyte complex and not because of a raised LH.
 
Effect on Reproductive Outcomes
 
Adverse Effects on Oocyte Maturation, Fertilization or Early Cleavage
A study showed that the mean number of retrieved oocytes, recovered mature oocytes, embryos and top quality embryos were significantly higher in the non-prematurely luteinized group than in the prematurely luteinized group. Although fertilization rates and implantation rates were similar between the two groups, the clinical pregnancy rate was higher in the non-prematurely luteinized group than in the prematurely luteinized group.3 However, many authors have not found a negative impact of PL.10109
 
Effect on Endometrium
Since many studies did not find an adverse oocyte quality, it was suggested that PL has an adverse effect on endometrium. There is an abnormally accelerated endometrial maturation leading to impaired endometrial receptivity.11
 
Pregnancy Rate
A systemic review and meta-analysis stated that no statistically significant association between progesterone elevation and the probability of clinical pregnancy was detected in women undergoing ovarian stimulation with GnRH analogs and gonadotropins for IVF.12
With COH cycles using GnRH antagonists and where serum P is measured by ELISA there does not seem to be any disadvantage of higher serum P levels up to 2 ng/mL at the time of hCG in IVF-ET cycles.13
 
Prevention
 
Flexible Antagonist Protocol
In flexible antagonist protocol, antagonist is initiated when follicle size is more than 14 mm or estradiol is more than 600 pg whereas in the fixed protocol it is administered on day 6. In stimulated cycles, it was seen that intense ovarian response with more number of follicles led to an early rise in estradiol. Thus, threshold levels of estradiol that initiates a LH surge are reached earlier before follicles reach an optimum size. In these cases the flexible protocol, which is dependent on the size of the follicle on ultrasound to start antagonist to suppress surge, may no longer be accurate to determine time of initiation of GnRH antagonist.2 This observation might explain the observed lower efficacy of the flexible protocol compared 110with a fixed protocol in a meta-analysis of four studies.3 Hence, for patients with a profound ovarian response, early initiation of the GnRH antagonist may be needed.14
Treatment with ganirelix effectively prevents premature LH rises; luteinization in subjects undergoing stimulated IUI. Low-dose rFSH regimen combined with a GnRH antagonist may be an alternative treatment option for subjects with previous proven luteinization.15
 
Low-dose hCG Alone in the Late Follicular Stage
Women who were undergoing COS with recombinant FSH/hMG followed by low-dose hCG (200 IU/day) alone [66 ]. This regimen did not cause PL.16
 
Mifepristone
Mifepristone was started in a daily dose of 40 mg along with stimulation. 50 mg progesterone was given along with hCG to counteract the antiprogesterone effect of mifepristone. No PL was seen in any case. However, endometrial receptivity status requires additional evaluation after decreasing RU-486 doses.17
 
hCG Administration Preponed to day of Progesterone Rise
Progesterone was monitored from day 7 and hCG was given when a rise was detected >1.0 ng/ml. It was seen that the quality of embryos and implantation rate was better than when hCG was delayed in cases of PL.18
 
Aspiration of Single Lead Follicle
Better pregnancy rates were seen if a single lead follicle was aspirated and other follicles continued to grow and were aspirated later. No premature LH surge was seen.19111
Premature luteinization is a diagnostic and therapeutic challenge for the infertility specialist. Its impact on ART results is controversial and more randomized studies are required before one can be definite about it.
 
References
  1. Hofmann GE, Bentzien F, Bergh PA, Garrisi GJ, Williams MC, Guzman I, et al. Premature luteinization in controlled ovarian hyperstimulation has no adverse effect on oocyte and embryo quality. Fertil Steril. 1993;60:675–9.
  1. Younis JS, Simon A, Laufer N. Endometrial preparation: lessons from oocyte donation. Fertil Steril. 1996;66:873–84.
  1. Ou YC, Lan KC, Chang SY, Kung FT, Huang FJ. Increased progesterone/estradiol ratio on the day of hcg administration adversely affects success of in vitro fertilization–embryo transfer in patients stimulated with gonadotropin-releasing hormone agonist and recombinant follicle-stimulating hormone Taiwan. J Obstet Gynecol. 2008;47:168–74.
  1. Keltz MD, Stein DE, Berin I, Skorupski J. Elevated progesterone-to-estradiol ratio versus serum progesterone alone for predicting poor cycle outcome with in vitro fertilization. J Reprod Med. 2012;57(1-2):9–12.
  1. Sims A, Seltman HJ, Muasher SJ. Early follicular rise of serum progesterone concentration in response to a flare-up effect of gonadotrophin-releasing hormone agonist impairs follicular recruitment for in-vitro fertilization. Hum Reprod. 1994;9:235–40.
  1. Seow KM, Lin YH, Huang LW, Hsieh BC, Huang SC, Chen CY, et al. Subtle progesterone rise in the single-dose gonadotropin-releasing hormone antagonist (cetrorelix) stimulation protocol in patients undergoing in vitro fertilization or intracytoplasmic sperm injection cycles. Gynecol Endocrinol. 2007;23:338–42.
  1. Edelstein MC, Seltman HJ, Cox BJ, Robinson SM, Shaw RA, Muasher SJ. Progesterone levels on the day of human chorionic gonadotropin administration in cycles with gonadotropin-releasing hormone agonist suppression are not predictive of pregnancy outcome. Fertil Steril. 1990;54:853–7.
  1. Bosch E, Valencia I, Escudero E, Crespo J, Simon C, Remohi J, et al. Premature luteinization during gonadotropin-releasing hormone antagonist cycles and its relationship with in vitro fertilization outcome. Fertil Steril. 2003;80:1444–9.
  1. Copperman AB, Horowitz GM, Kaplan P, Scott RT, Navot D, Hofmann GE. Relationship between circulating human chorionic gonadotropin levels and premature luteinization in cycles of controlled ovarian hyperstimulation. Fertil Steril. 1995;63:1267–71.
  1. Martinez F, Coroleu B, Clua E, Tur R, Buxaderas R, Parera, et al. Serum progesterone concentrations on the day of hCG administration cannot predict pregnancy in assisted reproduction cycles. Reprod Biomed Online. 2004;8:183–90.
  1. Yovel I, Yaron Y, Amit A, Peyser MR, David MP, Kogosowski A, et al. High progesterone levels adversely affect embryo quality and pregnancy rates in vitro fertilization and oocyte donation programs. Fertil Steril. 1995;64:128–31.
  1. Bosch E. Comment on: is progesterone elevation on the day of human chorionic gonadotrophin administration associated with the probability of pregnancy in vitro fertilization? A systematic review and meta-analysis. By Venetis et al (2007). Hum Reprod Update. 2008;14:194–5.
  1. Katsoff B, Check JH, Wilson C, Choe JK. Effect of serum progesterone level on the day of human chorionic gonadotropin injection on outcome following in vitro fertilization-embryo transfer in women using gonadotropin releasing hormone antagonists. Clin Exp Obstet Gynecol. 2011;38(4):322–3.
  1. Al-Inany HG, Aboulghar M, Mansour R, Serour GI. Optimizing GnRH antagonist administration: meta-analysis of fixed vs flexible protocol. Reprod Biomed Online. 2005;10:567–70.
  1. Lambalk CB, Leader A, Olivennes F, Fluker MR, Andersen AN, Ingerslev J, et al. Treatment with the GnRH antagonist ganirelix prevents premature LH rises and luteinization in stimulated intrauterine insemination: results of a double-blind, placebo-controlled, multicentre trial. Hum Reprod. 2006;21(3):632–9.
  1. Filicori M, Cognigni GE, Gamberini E, Parmegiani L, Troilo E, Roset B. Efficacy of low-dose human chorionic gonadotropin alone to complete controlled ovarian stimulation. Fertil Steril. 2005;84:394–401.
  1. Escudero EL, Boerrigter PJ, Bennink HJ, Epifanio R, Horcajadas JA, Olivennes F, et al. Mifepristone is an effective oral alternative for the prevention of premature luteinizing hormone surges and/or premature luteinization in women undergoing controlled ovarian hyperstimulation for in vitro fertilization. J Clin Endocrinol Metab. 2005;90:2081–8.
  1. Harada T, Katagiri C, Takao N, Toda T, Mio Y, Terakawa N. Altering the timing of human chorionic gonadotropin injection according to serum progesterone concentrations improves embryo quality in cycles with subtle P rise. Fertil Steril. 1996;65:594–7.
  1. Barash A, Shoham Z, Lunenfeld B, Segal I, Insler V, Borenstein R. Can premature luteinization in superovulation protocols be prevented by aspiration of an ill-timed leading follicle? Fertil Steril. 1990;53:865–9.

Polycystic Ovarian Syndrome and Insulin Sensitizerschapter 8

Surveen Ghumman
Polycystic ovarian syndrome (PCOS) is a heterogeneous collection of signs and symptoms that form a spectrum of mild to severe disturbance of reproductive, endocrine and metabolic functions. It was first described by Stein and Leventhal in 1935.1 The disorder is multifactorial in origin. It is thought to have a genetic etiology but the severity and course is determined by lifestyle, especially body mass index. 80 to 90% of women suffering from anovulation have PCOS. Prevalence of PCOS has been studied in several populations and it appears that it affects as many as 5–10% of women of reproductive age.
 
Diagnosis
Presence of two out of the following three criteria are essential for diagnosis.1
  1. Oligo and/or anovulation.
  2. Hyperandrogenism (clinical and/or biochemical).
  3. Polycystic ovaries with exclusion of other etiologies.
115
 
Histopathological Criteria
  1. Atretic follicles and/or degenerating granulosa cells.
  2. Hypertrophy and luteinization of the inner theca cell layer.
  3. Thickened ovarian tunica.
 
Transvaginal Sonography
Diagnosis is on the basis of these criteria:2
  1. Presence of 12 or more cysts of 2 to 9 mm
  2. Ovarian volume equal to or more than 12 cm3
  3. Bright echogenic stroma.
Degree of insulin resistance is correlated well with the ovarian volume and stromal echogenicity whereas serum LH and testosterone is related well with ovarian volume, stromal echogenicity and follicle number. Ovarian volume was found to be the best predictor for hyperandrogenism.3
 
Insulin Resistance
Insulin resistance is defined as reduced glucose response to a given amount of insulin. It occurs in 80% of obese women and 30 to 40% of women with normal weight with PCOS. Hyperinsulinemia is more common in patients with more than 10 follicles in the ovary, enlarged ovarian volume or elevated day 10 LH.
Peripheral target tissue insulin resistance can be due to decreased number of peripheral insulin receptors, decreased insulin binding or a post receptor failure. In PCOS it is caused by the post-receptor defect because of excessive serine phosphorylation of beta chain of insulin receptor and of adrenal and ovarian cytochrome P450c17 enzyme. This enzyme catalyzes 17 hydroxylase and the 17, 20 lyase activities 116that is a rate limiting step in androgen biosynthesis thus leading to hyperandrogenemia. There is evidence of adrenal hyperandrogenemia in 15% of PCOS women (Fig. 8.1).
Peripheral target tissue resistance leads to hyperinsulinemia as a compensatory mechanism. When the beta cells of pancreas fail to meet this challenge there are declining insulin levels and an impaired GTT finally leading to type 2 NIDDM. Hyperinsulinemia leads to adverse lipid effects (Increased triglycerides and VLDL cholesterol and a decreased HDL) (Fig. 8.2).
Insulin when in excess binds to IGF-I receptors and also decreases insulin-like growth factor binding protein I (IGFBP-I) production in liver thus, increasing levels of IGF-I. IGF-I augments theca androgen response to LH. Another theory states that insulin binds to its own receptors causing steroidogenesis. Increased IGF-I activity in endometrium may also be responsible for the endometrial growth and increased risk of endometrial cancer in these patients. Plasminogen activator inhibitor-I is increased causing impaired fibrinolysis.
In PCOS the ovary does not secrete increased amount of estrogen. Levels of estrone are increased because of peripheral conversion of increased amount of androstenedione to estrone.
Fig. 8.1: Mechanism of hyperinsulinemia and hyperandrogenemia4
117
Fig. 8.2: Effect of hyperinsulinemia in polycystic ovarian syndrome
Serum prolactin levels may be high in 30 to 40% of PCOS women because of increased estrogen levels.
Sex hormone binding globulin are decreased because raised insulin levels inhibit hepatic synthesis of SHBG (Fig. 8.2). Also the increased testosterone level suppresses SHBG. This further increases levels of estradiol and testosterone. High levels of estradiol cause increased LH secretion and suppress FSH secretion 40% of cases will have an increased level of LH. As FSH secretion is not totally suppressed follicular growth is continuously stimulated but not to the point of full maturation and ovulation. Small follicles 2 to 10 mm in diameter are present that may last for months. These are surrounded by hyperplastic theca cells that under the influence of LH get luteinized. The tissue derived from follicular atresia contributes to stromal compartment that secretes androstenedione and testosterone. High androgens 118prevent normal follicular development and premature atresia of follicles.
All anovulatory women who are androgenic should be assessed for glucose tolerance and insulin resistance with measurement of 2 hour glucose and insulin value after 75 g load.4
 
Clinical Presentation
  1. Hyperandrogenism (acne, hirsutism, alopecia—not virilization).
  2. Menstrual disturbance.
  3. Infertility.
  4. Obesity.
  5. Clinical evidence of insulin resistance – Acanthosis nigricans.
 
Late Sequelae
  1. Diabetes mellitus.
  2. Dyslipidemia.
  3. Hypertension and cardiovascular disease.
  4. Endometrial carcinoma.
  5. Breast cancer.
 
Serum Endocrinology
  1. Fasting insulin levels (Normal < 25 IU/L) in glucose tolerance test preferred (Tables 8.1 and 8.2).
  2. Postprandial insulin >100 ug/mL
  3. Fasting blood sugar:insulin ratio (> 4.5)
  4. Glucose tolerance test (Table 8.1)
  5. Total and free testosterone (Normal total testosterone 20–80 ng/DL)
  6. DHEAS (Normal–Less than 350 μg/DL)119
    Table 8.1   Blood glucose value after 75 g load4
    Normal
    Impaired
    Diabetes mellitus
    Fasting (mg/DL)
    < 100
    100–126
    > 126
    2 hour value (mg/DL)
    < 140
    140–199
    > 200
    Table 8.2   Hour insulin value after 75 g load4
    Likely insulin resistance
    Insulin resistance
    Severe insulin resistance
    Value (μg/mL)
    100–150
    150–300
    > 300
  7. LH raised (2–10 IU/L) Measured on day 2–3 or on any day if amenorrheic
  8. FSH normal (2-8 IU/L)
  9. Decreased SHBG (normal 16–119 nmol/L)
  10. Free androgen index (FAI) T × 100/SHBG (Normal <5)
  11. Estradiol, estrone are increased (not measured routinely)
  12. Serum prolactin increased (normal < 20 ng/mL). Measure if oligo-/amenorrheic.
  13. Thyroid stimulating hormone—increased sometimes
  14. Lipid profile—Low-density lipoprotein and high-density lipoprotein cholesterol levels.
 
Treatment
 
Goals
  1. Reducing insulin.
  2. Treating anovulation.
  3. Regularizing cycles.
  4. Antagonizing androgens.
  5. Maintaining a normal endometrium.120
 
Weight Loss
Weight loss must be initiated as part of treatment plan alone or along with drug therapy for all patients of PCOS irrespective of whether they desire pregnancy or not. It improves ovarian function and hormonal abnormality by decreasing insulin and androgens and increasing SHBG. Central obesity and BMI are major determinants of insulin resistance, hyperinsulinemia and hyperandrogenemia. 5–10% of weight loss is enough to decrease the visceral fat by 30% and restore reproductive function. It should be encouraged prior to ovulation as patients respond better and require less dose of ovulation inducing drugs.
 
Diet
Food with low glycemic index such as vegetables, fruits, and fiber should be consumed. Regular aerobic exercise is beneficial. Two hours of exercise a week is sufficient.
 
Drugs
Orlistat: Orlistat reduces up to 30% lipid adsorption and produced a significant reduction in weight and total testosterone. The reduction in total testosterone was similar to that seen after treatment with metformin.5 Orlistat improves the hormonal and metabolic profile in women with PCOS after 6 months of treatment, independent of BMI changes.6
Sibutramine: A new drug, sibutramine increases satiety and energy expenditure caused by thermogenesis in brown adipose tissue. Sibutramine in combination with lifestyle intervention results in significant weight reduction in obese patients with PCOS. Sibutramine 15 mg once daily together with brief lifestyle modification has lead to a weight loss of 7.8 kg in 6 months compared with a weight loss of 2.8 kg in 121those who followed only lifestyle changes.7 In addition to the weight loss, sibutramine seems to have beneficial effects on metabolic and cardiovascular risk factors.
 
Surgery
Bariatric surgery can be considered in morbidly obese women who do not respond to any treatment.
 
Insulin Sensitizers
The insulin sensitizers used in ovulation induction are shown in Table 8.3.
 
Metformin
Metformin is a water-soluble oral biguanide that lowers insulin, LH, free testosterone levels, PAI-I and endothelin I levels in overweight women with polycystic ovaries.
Table 8.3   Insulin sensitizers
Insulin sensitizers
Dose
1. Biguanides metformin
1500–2000 mg/day
2. Thiazolidinediones
   Rosiglitazone
4–8 mg/day
   Pioglitazone
30–50 mg/day
3. D-chiroinositol
1200 mg/day
122
Action
  1. Blunting of hepatic gluconeogenesis.
  2. Decreased intestinal absorption of glucose.
  3. Increased peripheral glucose uptake and utilization.
However, it is seen that the ability of metformin to alter insulin sensitivity in morbidly obese (BMI 40 kg/m2) is limited.8 Metformin by reducing hyperinsulinemia causes a decrease in intraovarian androgens. This in turn leads to a reduction in E2 levels and favors orderly follicular growth in response to exogenous gonadotropins. There is a decrease in testosterone, free testosterone, DHEAS, androstenedione and LH, normalization of LH: FSH ratio and an increase in SHBG.
Indication for Metformin Therapy
  1. PCOS patients with increased androgens.
  2. Documented ovulation induction failure after clomiphene.
  3. Fasting insulin level more than 25 IU/L (some take a value of 15 IU/L).
  4. Altered glucose tolerance test.
Justification of administrating insulin lowering agents in patients with normal insulin values needs to still be evaluated although some therapeutic benefit is seen in terms of reproductive function.
Precautions: Metformin can cause lactic acidosis in 1:33,000 cases. It is a serious condition with a mortality of 50%, mainly occurring in women with renal impairment. Symptoms are often nonspecific like fatigue, myalgia, abdominal distension, vomiting and respiratory depression. Immediate cessation of the drug is indicated on observing any of the symptoms. Serum electrolytes, blood glucose, ketones, pH, serum lactate level and serum metformin levels, if possible, should be done.123
To take precautions against this condition metformin should be discontinued 48 hours before any planned surgery or any radiographic study utilizing intravenous contrast dye. Ethanol potentiates the effect of metformin and patients should be warned against high alcohol intake. Hemodialysis may be needed to resolve the situation.9
Minor side effects like nausea, vomiting, diarrhea, bloating, flatulence and metallic taste occurs in 20% patients. It resolves if drug is taken with food. Since this effect is dose dependent, the dose of metformin should be increased in an incremental fashion. If discomfort is significant, the drug should be discontinued. There may be weight loss associated with the nausea and vomiting accompanying the drug. Megaloblastic anemia may occur in some patients because of subnormal B12 levels.
Hypoglycemia does not occur with metformin in euglycemic patients. It may be seen in special cases where there is:
  1. Deficient caloric intake.
  2. Concomitant use with sulfonylureas.
  3. Strenuous exercise is not compensated with adequate intake.
  4. Excessive alcohol consumption.
Contraindications
  1. Renal disease: If serum creatinine is more than 1.5 mg/DL or creatinine clearance is less than 60% of normal for that age, metformin is not given.
  2. Metabolic acidosis.
  3. Myocardial infarction.
Drug interaction: Drug interaction occurs with diuretics, oral contraceptives and phenytoin.
Dose: The drug is usually started in the follicular phase in a dose of 500 mg/day for 5 to 7 days. This is increased in 124weekly increments of 500 mg up to 1500 to 2000 mg/day. If patient is amenorrheic pregnancy needs to be ruled out.
Results: Metformin induces regular cycles in 68 to 95% patients treated for 4 to 6 months. It improved ovulation, hirsutism, hyperandrogenemia and insulin resistance. Lowering of fasting insulin levels are seen in 2 to 3 months. A repeat test is required only after this period. If amenorrhea persists clomiphene or rosiglitazone is added. Ovulation rates are higher when combined with clomiphene (76% versus 46% when used alone).10 Patients with elevated pretreatment levels of testosterone show the best results in resumption of ovulation with significant reduction in testosterone. Those with raised fasting insulin responded less and those with normal testosterone showed no effect.
A Cochrane review 2010 concluded that metformin is of benefit in improving clinical pregnancy and ovulation rates. However, there is no evidence that metformin improves live birthrates whether it is used alone or in combination with clomiphene, or when compared with clomiphene. Hence, the use of metformin in improving reproductive outcomes in women with PCOS appears to be limited.11
Metformin in ART Cycles
A recent Cochrane review found that the risk of OHSS in women with PCOS and undergoing IVF or ICSI cycles was reduced with metformin. There was no evidence that metformin treatment before or during ART cycles improves live birth or pregnancy rates.12
Role of Metformin After Conception
It has been suggested that metformin be continued after conception as it decreases rate of miscarriage and gestational diabetes without any teratogenicity as it is a category B drug.13 The risk of miscarriage is thought to be due to increased 125levels of PAI-I that accompany hyperinsulinemia suggesting the possibility that placental thrombosis induces miscarriage. However, there are no randomized controlled trials.
 
Thiazolidinediones
They markedly improve insulin sensitivity mainly by improved peripheral glucose utilization in skeletal muscle. Troglitazone decreases hyperinsulinemia, androgens, PAI-I, LH and increases SHBG. Cycles become ovulatory. However, it was withdrawn from the market because of liver toxicity. Pioglitazone and rosiglitazone have been reported free of liver toxicity but liver function should be monitored every 2 months. Minor side effects like weight gain and fluid retention may be present. Pioglitazone in a dose of 30 to 50 mg/day and rosiglitazone in a dose of 4 to 8 mg/day can also be used as monotherapy or in combination with metformin.14,15 They produce an ovulatory response in cases that were resistant to metformin.16 They are both Category C drugs and need to be discontinued when patient becomes pregnant.
 
Acarbose
Acarbose is used orally in the management of type 2 diabetes. Acarbose reduces the postprandial rise in both serum glucose and insulin levels by inhibiting α-glucosidase, an enzyme responsible for the intestinal absorption of carbohydrates. It was as effective as metformin when given in a dose of 100 mg tid in clomiphene resistant PCOS women. It significantly reduced LH, LH:FSH ratio, and testosterone and fasting insulin concentrations, and increased FSH concentrations versus pretreatment values. There was a significant increase in ovulation. Acarbose was found to be a safe and effective agent that could be used in cases with clomiphene-resistant PCOS.17 Acarbose improved hirsutism, acne, and menstrual 126irregularities through reduction in androgen concentrations and increased androgen binding. Markers of cardiovascular risk were also significantly improved following 6 months of acarbose therapy in obese women with PCOS.18 The rate of flatulence and diarrhea was significantly lower for acarbose compared to metformin (38% vs 80%).19
 
D-chiroinositol
Administration of D-chiroinositol makes up deficiency of D-chiroinositol containing phosphoglycan that mediates action of insulin in these patients. It is given in a dose of 1200 mg/day for 6–8 weeks to correct ovulatory dysfunction.20 86% of women ovulated compared to 26% in the placebo group.
 
Endometrial Biopsy
It is indicated in all women who have a clinical history of long-term unopposed estrogen exposure even when the endometrial thickness is normal 5 to 12 mm and in those where endometrial thickness is greater than 12 mm even though clinical suspicion of the disease is low.
 
Ovulation Induction
Aim of ovulation should be to correct the underlying disturbance and achieve unifollicular ovulation.
 
Problems of Ovulation Induction in PCOS
  1. Disturbed folliculogenesis leading to poor response to induction.
  2. Large number of antral follicles sensitive to FSH leading to multiple follicular development, OHSS and multiple pregnancy.127
  3. Tonically elevated serum LH levels leading to premature luteinization, low pregnancy rates and high miscarriage rates.
Several modes of inducing ovulation are used (Fig. 8.3). Maybe a combination of all will work (Table 8.4).
  1. Weight loss: It is important to take into consideration the patients weight while prescribing any drug. Weight loss helps in restoring ovulation and improves response to ovulation inducing drugs.
  2. Clomiphene citrate: In PCOS patients the usual dose of 50 mg may need to be lowered to 25 mg as patients are prone to hyperstimulation. 20% of women do not respond to clomiphene.
    Fig. 8.3: Modes of ovulation induction in PCOS
    128
    Table 8.4   Options for ovulation induction in PCOS
    1. Weight loss
    2. Clomiphene citrate
    • Clomiphene alone
    • Clomiphene along with adjuvants
      1. hCG
      2. Metformin
      3. Glucocorticoids
      4. Bromocriptine
    3. Gonadotropin therapy
    • Gonadotropin alone
    • Gonadotropins with addition of
    • GnRH agonist
    • GnRH antagonist
    4. Insulin sensitizers like metformin
    5. Surgical ovulation induction—laparoscopic ovarian drilling
    Dose can be increased to 250 mg (see Chapters 2 and 3). In cases which do not respond clomiphene may be given for a period of 8 days.21 Prior administration of progesterone intramuscularly in a dose of 50 mg for 5 days can cause a suppression of LH secretion. Injection hCG is given in a dose of 5000 to 10,000 IU as ovulation trigger in cases where there is a delayed or absent LH surge despite presence of well-developed follicle.
  3. Insulin sensitizing drugs: Metformin in a dose of 1500 to 2000 mg may be started along with clomiphene in cases with hyperinsulinemia as has been discussed in detail earlier in this chapter.
  4. Naltrexone: It is an opiate receptor antagonist that lowers insulin, LH and androgen levels and induces ovulation in 86% clomiphene resistant cases with a pregnancy rate of 55%.22129
  5. Glucocorticoids: 50% of patients of PCOS show involvement of an adrenal component with raised DHEAS.23 The desired effect should be to normalize without suppressing the adrenal component, with dexamethasone (0.25–0.5 mg/day) or prednisolone (5–10 mg/day). Dexamethasone has the advantage of having a longer half-life compared to prednisolone although this may cause over suppression at times. Dose of 0.25 mg/day is seen to suppress 50% of patients. Two regimes can be followed. Dexamethasone may be started with clomiphene and stopped when ovulation is documented. The other regime is continuous administration of glucocorticoid till pregnancy is achieved. The logic behind this regime is that the follicle takes 100 days for development and the effect of the drug is present through that period. It is usually never given beyond 6 months. Dexamethasone gives best results when administered at night as adrenals are most active early morning. DHEAS and testosterone levels are monitored after one month. Glucocorticoids with gonadotropins have been tried and show conflicting results with some quoting an improved pregnancy rate and others showing no change.24,25 The side effect most commonly seen is weight gain that can be upto 4 to 5 kg in 4 months. It is a common cause for discontinuation of the therapy. There may be adrenal suppression and inability to cope with stresses of major surgery if dose is more than 0.5 mg or duration of therapy is more than one month. There may be osteoporosis, worsening of glucose tolerance, skin changes, gastritis, lipid abnormalities and hypertension on prolonged use.
  6. Bromocriptine: In cases of a raised serum prolactin or galactorrhea bromocriptine in a dose of 2.5 to 7.5 mg/day may be added along with clomiphene to improve results.130
  7. Gonadotropins: These patients are more prone to hyperstimulation. This is not due to the difference in FSH threshold level but to the fact that they contain twice the number of FSH sensitive antral follicles. Due to this the chronic low dose regime is employed where FSH is started on day 3 at a low dose of 37.5 to 75 IU for 14 days in first cycle and 7 days in subsequent cycles. Thereafter increments of 25 to 37.5 IU are given at weekly intervals until follicular development is initiated. The dose that initiates follicular development is continued till criteria for giving hCG is attained. Monitoring is done as usual. Compared to the conventional step-up protocol pregnancy rates improved (40% vs 24%), uniovulation was induced in more (74% vs 27%) and there was no OHSS or multiple pregnancy that was prevalent (11% OHSS and 33% multiple pregnancy) in the conventional therapy.26 The chances of multiple pregnancy were reduced if a strict criteria is followed for administration of hCG. There should be no more than three follicles greater than 14 mm and estradiol should be less than 1500 pg/mL. In poor responders conventional step-up protocol may be tried where the gonadotropin is started at 150 IU and increased by 75 IU according to response. Recently, some studies showed better results regarding unifollicular ovulation with step-down regimes compared with low dose step-up protocol. These involve starting higher dose of FSH and bring it down after follicular recruitment (see Chapter 4). It is thought that pure FSH may be better in cases where LH levels are high. This theory has been refuted by a few studies where no difference in ovulation, pregnancy, or miscarriage rate are seen following administration of hMG versus purified FSH.27131
  8. GnRH agonists: They can be used to downregulate patients with high LH levels prior to giving ovulation induction drugs. There was a miscarriage rate of 17.6% vs 39% and a cumulative live birthrate of 64% vs 26% for regimen with GnRH agonist vs those without.28 It has the disadvantage of requiring higher dose of gonadotropin for longer periods and increased incidence of multiple follicular development leading to OHSS and multiple pregnancy. This treatment is reserved for women with high serum concentration of LH with failure to conceive on gonadotropins, repeated premature luteinization, and early miscarriages. They are usually started as a long protocol on day 21 of previous cycle. Gonadotropins are added after confirming down- regulation on day 2 (Details in Chapter 5).
  9. GnRH antagonist: They have the advantage of acting by competitive binding that allows a modulation of the degree of hormonal suppression by their dose. They act within hours, have no flare effect and gonadal function assumes without a lag effect. Compared with an agonist treated cycle this would have the advantage of a shorter cycle treatment, promise more conceptions, and fewer miscarriages, reduce the amount of gonadotropins needed and increase the incidence of monofollicular ovulation with a consequent reduction in prevalence of OHSS and multiple pregnancy. Centrorelix can be given as a single dose of 3 mg once follicle reaches 14 mm (Detail in Chapter 5). However, they cannot bring down raised basal LH.
  10. Surgical ovulation induction: Laparoscopic ovarian drilling is done by laser or electrocautery where multiple punctures are made in the ovary. This induces ovulation 132in 70 to 92% patients with a pregnancy rate of 40 to 80%. There is no need for intensive monitoring as there is no hyperstimulation or multiple pregnancy (Details in Chapter 9).
  11. IVF: If all other treatment is ineffective IVF is the next option. Although smaller percentage of oocytes are fertilized, the large number of oocytes recovered in these cases balances out the pregnancy rate.
133
 
Treatment in those who do not Desire Pregnancy
Treatment is given to these patients to improve symptoms and prevent complications.
 
Prevention of Endometrial Hyperplasia
  1. Monthly medroxyprogesterone: Medroxyprogesterone in a dose of 5 to 10 mg/day or norethindrone in a dose of 5 to 15 mg for 10 to 14 days each month should be given. This avoids abnormal endometrial proliferation but does not suppress the ovarian androgen production.
  2. Low dose oral contraceptive pills.
  3. GnRH agonists: They are used where there is no response to hormones or there are severe side effects.
 
Regularization of Cycles
  1. Low dose contraceptive pill: Advantages of contraceptive pills are contraception, prevention of endometrial hyperplasia, regularization of cycles and treatment of hirsutism. Oral contraceptive pills containing estradiol in low dose and progestogen desogestrel that is lipid friendly and has no androgenic effect, are used (Fig. 8.4).
  2. Metformin: Metformin may be given in cases where insulin levels are high. Menstrual cyclicity returns in 65 to 95% cases when treated for 4 to 6 months.
 
Hyperinsulinemia
Hyperinsulinemia should be treated because of the metabolic effects it has (Fig. 8.2). Metformin, rosiglitazone or pioglitazone may be given. Details have already been discussed earlier in the chapter.134
Fig. 8.4: Management of PCOS
135
 
Hyperandrogenism
Hyperandrogenism may be determined clinically by hirsutism, acne and alopecia and biochemically by raised testosterone, free testosterone index, dehydroepiandrosterone sulfate (DHEAS) and androstenedione.
Other tests in patients showing increased androgen levels are:
  1. 17 hydroxyprogesterone: It is a screening test for adult onset congenital adrenal hyperplasia. This should be done once PCOS is ruled out. The sample should be drawn at 8 AM. Basal follicular phase serum 17 OHP levels above 5 ng/mL suggest this disorder.
  2. Overnight dexamethasone suppression test: It should be performed in women with physical features of cortisol excess such as hypertension, central obesity, facial plethora, easy bruisibility, striae and proximal muscle weakness.1 mg of dexamethasone is administered orally at 11 PM and serum cortisol measurement is taken at 8 am, the following morning. Serum cortisol level below 5 μg (140 nmol/L) rules out Cushing's syndrome but may be present in PCOS women.
Treatment consists of:
  1. Oral contraceptive pills: They suppress ovarian androgen production and increase sex hormone binding globulin thereby reducing free testosterone. Oral contraceptive pills containing 2 mg cyproterone acetate and 35 μg ethinyl estradiol or those containing desogesterol with ethynyl estradiol are recommended.
  2. Cyproterone acetate: It antagonizes the androgen receptors in the skin and acts as a weak progestogen that inhibits gonadotropin secretion thereby decreasing androgen production. Dose 50 to 100 mg/day for 5 to 15 along with cyclic estrogen to regularize menstruation. It can also be 136given as oral contraceptive pill. It has been shown that it increases ovulation rate in women on clomiphene or gonadotropins.
  3. Spironolactone: It is an oral aldosterone antagonist with antiandrogenic properties. It increases metabolic clearance of testosterone and reduces cutaneous 5 alpha reductase activity. It reduces the hirsuitism score by 40% and is effective alone in 50% women. Dose 50 to 200 mg/day. The most common side effect is irregular menses and hence, it should be used along with oral contraceptive pills.
  4. Flutamide: It is a nonsteroidal antiandrogen at the receptor level given in a dose of 250 mg once or twice a day. It reduces the levels of free and total testosterone reducing the hirsuitism and regularizing the menstrual cycle.
  5. Finasteride: It acts as a type II 5 alpha reductase inhibitor given in a dose of 5 mg/day.
  6. GnRH agonist: They can be given in a depot preparation with the goal of reducing serum testosterone to 40 ng/DL for patients resistant to above therapy.
  7. Metformin: It is added if there is hyperinsulinemia.
  8. Ketoconazole: By suppression of steroidogenesis it brings down androgen and LH levels. It is a known teratogen and hence, has limited use in young woman with PCOS trying to conceive.
Management of PCOS is complex and requires multiple metabolic and endocrine factors to be taken into account. Since the spectrum of clinical presentation and metabolic pathology is vast all ovulation induction protocols have to be individualized in PCOS (Fig. 8.4).137
 
References
  1. Fauser B, Tarlatzis B, Chang J, Azziz R, et al. The Rotterdam ESHRE/ASRM sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovarian syndrome (PCOS). Hum Reprod. 2004;19:41–7.
  1. Balen AH, Laven JSE, Tan SL, Dewailly D. Ultrasound assessment of polycystic ovary: international consensus definitions. Human Reprod Update. 2003;9:505–14.
  1. van Santbrink EJP, Hop WC, Fauser BCJM. Classification of normogonadotropic infertility: Polycystic ovaries diagnosed by ultrasound versus endocrine characteristics of polycystic ovary syndrome. Fertil Steril. 1997;67:452–9.
  1. Speroff L, Fritz MA. Anovulation and Polycystic Ovary. In: Speroff L, Fritz MA (Eds): Clinical Gynecological Endocrinology and infertility. Williams and Wilkins  Philadelphia USA. 2005;465–98.
  1. Jayagopal V, Kilpatrick ES, Holding S, Jennings PE, Atkin SL. Orlistat is as beneficial as metformin in the treatment of polycystic ovarian syndrome. J Clin Endocrinol Metab. 2005;90(2):729–33.
  1. Diamanti-Kandarakis E, Katsikis I, Piperi C, Alexandraki K, Panidis D. Effect of long-term orlistat treatment on serum levels of advanced glycation end-products in women with polycystic ovary syndrome. Clin Endocrinol (Oxf). 2007;66(1):103–9.
  1. Lindholm A, Bixo M, Björn I, Wölner-Hanssen P, Eliasson M, Larsson A, et al. Effect of sibutramine on weight reduction in women with polycystic ovary syndrome: a randomized, double-blind, placebo-controlled trial. Fertil Steril. 2008;9(5):1221–8.
  1. Kahn SE, Prigeon RL, Mcculloch DK, Bayco EJ, Bergman RN, Schwartz MW, et al. Quantification of relationship between insulin sensitivity and b cell function in human subjects. Diabetes. 1993;42:1663–72.
  1. Heaney D, Majid A, Junor B. Bicarbonate hemodialysis as a treatment of metformin overdose. Nephrol Dial Transplant. 1997;12:1046–7.
  1. Lord JM, Flight IH, Norman RJ. Insulin-sensitising drugs (metformin, troglitazone, rosiglitazone, pioglitazone, D-chiroinositol) for polycystic ovary syndrome. Cochrane Database Syst Rev. 2003;(3):CD003053. Comment in: ACP J Club. 2004;140(3):75–80.
  1. Tang T, Lord JM, Norman RJ, Yasmin E, Balen AH. Insulin-sensitising drugs (metformin, rosiglitazone, pioglitazone, D-chiroinositol) for women with polycystic ovary syndrome, oligo amenorrhoea and subfertility. Cochrane Database Syst Rev. 2010 Jan 20;(1):CD003053.
  1. Tso LO, Costello MF, Albuquerque LE, Andriolo RB, Freitas V. Metformin treatment before and during IVF or ICSI in women with polycystic ovary syndrome. Cochrane Database Syst Rev. 2009 Apr 15;(2):CD006105.
  1. Glueck CJ, Wang P, Kobayashi S, Philips H, Seive -Smith, Wang P. Continuing metformin throughout pregnancy in women with polycystic ovary syndrome appears to safely prevent first trimester spontaneous abortion: A pilot study. Fertil Steril. 2001;75:46–52.
  1. Stout DL, Fugate SE. Thiazolidinediones for treatment of polycystic ovary syndrome. Pharmacotherapy. 2005;25(2):244–52.
  1. Ortega-Gonzalez C, Luna S, Hernandez L, Crespo G, Aquayo P, Arteaga-Troncoso G, et al. Responses of serum androgen and insulin resistance to metformin and pioglitazone in obese, insulin-resistant women with polycystic ovary syndrome. J Clin Endocrin Metab. 2005;90(3):1360–5.
  1. Glueck CJ, Moreira A, Goldeberg N, Sieve L, Wang P. Pioglitazone and metformin in obese women with polycystic ovary syndrome not optimally responsive to metformin. Hum Reprod. 2003;18:1618–22.
  1. Sönmez AS, Yasar L, Savan K, KoçS, Ozcan J, Toklar A, et al. Comparison of the effects of acarbose and metformin use on ovulation rates in clomiphene citrate-resistant polycystic ovary syndrome. Hum Reprod. 2005;20(1):175–9.
  1. Kircher C, Smith KP. Acarbose for polycystic ovary syndrome. Ann Pharmacother. 2008;42(6):847–51.
  1. Hanjalic-Beck A, Gabriel B, Schaefer W, Zahradnik HP, Schories M, Tempfer C, et al. Metformin versus acarbose therapy in patients with polycystic ovary syndrome (PCOS): a prospective randomised double-blind study. Gynecol Endocrinol. 2010;26(9):690–7.
  1. Nestler JE, Jakubowicz DJ, Reamer P, Gunn RD, Allan G. Ovulatory and metabolic effects of D-chiro-inositol in the polycystic ovary syndrome. New Eng J Med. 1999;340:314.
  1. Lobo RA, Granger LR, Davajan V, Mishcell DR Jr. An extended regime of clomiphene citrate in women unresponsive to standard therapy. Fertil Steril. 1982;37:762–6.
  1. Hadžiomerović-Pekić D, Wildt L, Weiss JM, Moeller K, Mattle V, Seeber BE. Metformin, naltrexone, or the combination of prednisolone and antiandrogenic oral contraceptives as first-line therapy in hyperinsulinemic women with polycystic ovary syndrome. Fertil Steril. 2010;94(6):2385–8.
  1. Gonzalez F. Adrenal involvement in polycystic ovarian disease. Semin in Reprod Endocrinol. 1997;15:137–57.
  1. Evron S, Navot D, Laufer N, Diamant YZ. Induction of ovulation with combined human gonadotropins and dexamethasone in women with polycystic ovarian disease. Fertil Steril. 1983;40:183–6.
  1. Bider D, Blankstein J, Levron J, Tur Kaspa I. Gonadotropins and glucocorticoid therapy for low responders: A controlled study. J Assist Reprod Genet. 1997;14:328–31.
  1. Homberg R, Levy T, Ben Rafael Z. A comparative prospective study of conventional regimen with chronic low dose administration of follicle stimulating hormone for anovulation associated with polycystic ovary syndrome. Fertil Steril. 1995;63:729–33.
  1. Venturoli S, Paradisi R, Fabbri R, Porcu E, Orsini LF, Flamigni C. Induction of ovulation in polycystic ovary: Human menopausal gonadotropins or human urinary follicle stimulating hormone? Int J Fertil. 1987;32:66–70.
  1. Homberg R, Levy T, Berkowitz D. GnRH agonist reduces miscarriage rate for pregnancies conceived in polycystic ovary syndrome. Fertil Steril. 1993;59:527–31.
  1. Hosseini MA, Aleyasin A, Saeedi H, Mahdavi A. Comparison of gonadotropin-releasing hormone agonists and antagonists in assisted reproduction cycles of polycystic ovarian syndrome patients. J Obstet Gynaecol Res. 2010;36(3):605–10.
  1. Vause TD, Cheung AP, Sierra S, Claman P, Graham J, Guillemin JA, et al. Society of Obstetricians and Gynecologists of Canada. Ovulation induction in polycystic ovary syndrome. J Obstet Gynaecol Can. 2010;32(5):495–502.

Surgical Ovulation Inductionchapter 9

Lalita Badhwar,
Surveen Ghumman
Polycystic ovarian disease is a heterogeneous group of disorders that may present with a wide variety of clinical syndromes. It may present with obesity, hyperandrogenism, menstrual cycle abnormalities, infertility and ultrasound finding of increased ovarian volume, multiple small cysts and bright echogenic stroma.
The aim of therapy of patients with polycystic ovarian disease is treatment of infertility, hirsutism, endometrial hyperplasia and irregular cycles. Patients desirous of pregnancy are treated with clomiphene citrate, gonadotropins with or without GnRH agonists or antagonists and insulin sensitizers (Refer to Chapter 8). The medical method of inducing ovulation fails in 25% of patients due to lack of adequate response or complications like OHSS. In these cases surgical option becomes important.
Wedge resection was first reported in 1935. With the introduction of clomiphene citrate, medical management replaced surgical management as the primary mode of treatment. Interest in laparoscopic surgical management was 142renewed in 1984 by Gjonnaess.1 Today, surgical management has definite indications and occupies an important place in management of PCOS patients.
 
Mechanism of Action
Proposed theories of how laparoscopic ovarian drilling improves chances of ovulation in these patients are many.
  1. Puncturing of follicles release androgen rich fluid and decrease androgen producing stroma so as to decrease circulating androgens.
  2. Crowding of cortex is reduced allowing progress of normal follicles to the surface resulting in resumption of normal ovulation.
  3. There is a fall in LH that results in increased secretion of FSH.
 
Indications
  1. Failure of medical therapy for ovulation induction.
  2. Persistent hypersecretion of LH.
  3. Intolerable side effects to drug therapy.
  4. Need for laparoscopic evaluation of pelvis.
  5. Inability to attend any intensive monitoring protocols required with drug therapy.
 
Techniques
 
Wedge Resection
It is no longer the surgery of choice as it requires that 50% of ovarian substance be removed. This results in a loss of ovarian tissue and adhesion formation.143
 
Operative Laparoscopy
It is a minimally invasive technique and has replaced ovarian wedge resection. The advantages of this procedure are:
  1. Multiple ovulatory cycles after a single treatment.
  2. No cyclic monitoring of ovulation induction is required.
  3. No risks of multiple pregnancy or ovarian hyperstimulation.
  4. Spontaneous abortion rate is less than that of medical induction.
  5. Less expensive and less cumbersome compared to gonadotropin treatment.
 
Steps of Surgical Technique
Instruments used are shown in Figures 9.1 and 9.2.
The procedure is done under general anesthesia with the patient in extended lithotomy position with Trendelenburg tilt (Fig. 9.3).
The (right handed) surgeon stands to the left of the patient.
Currently used is the three port laparoscopy technique (Fig. 9.4).
  • A 10 mm scope is used as the visual axis
  • The first port with 5 mm sheath is placed on left side lateral to the inferior epigastric vessels. The instruments inserted through this port are the atraumatic grasper used for grasping and retraction (surgeon's left hand).
  • Suprapubic portal is used for suction, irrigation and monopolar needle cautery (surgeon's right hand).
  • Optional right lateral portals for alternate grasper.
At the outset the uterus, tubes, ovaries and peritoneal folds are examined for any pathology (Fig. 9.5; See accompanying interactive CD-ROM).
Then by grasping the ovarian ligament or simply flipping over the ovary, the ovary is lifted and rotated to a more convenient position for puncture (Fig. 9.6).144
Fig. 9.1: Imaging system and insufflator
Fig. 9.2: Grasper, suction, etc.
145
Fig. 9.3: Extended lithotomy position
Fig. 9.4: Patients abdomen showing 3 points of entry
Fig. 9.5: View of bilateral polycystic ovaries
146
Fig. 9.6: Grasping of ovarian ligament
The needle is held perpendicularly to the ovarian surface and the follicles are punctured with simultaneous application of current for approximately one second or less (Fig. 9.7). This prevents accidental injury that may occur due to tangential slipping of the needle with energy and minimizes ovarian surface damage.
Attempt is made to puncture almost all the prominent follicles that amount to 4 to 10 punctures on each ovary (Fig. 9.8).
Fig. 9.7: Position of needle on ovary before puncture
147
Fig. 9.8: Appearance of ovary after drilling
The ovary is then irrigated thoroughly with normal saline to ensure hemostasis as well as cool it down at once (Fig. 9.9).
Please note that the surface damage to the ovary as well as the thermal damage should be minimized by reducing the number of punctures, the time for that the current is passed as well as by entering the ovary perpendicularly (Figs 9.10 and 9.11).
Fig. 9.9: Ovary irrigated with normal saline
148
Fig. 9.10: Excessive injury to ovarian surface
Fig. 9.11: Correct appearance of ovary after drilling
 
Caution
Sites to be avoided during drilling are:
  1. Area close to the mesovarium.
  2. Area close to the attachments of the infundibulopelvic and ovarian ligaments.
  3. Corpus luteum.
These sites tend to have troublesome bleeding while attempting to control may increase the ovarian damage.149
 
Various Laparoscopic Approaches
 
Simple Needle Puncture
This is thought to have minimal stromal damage, but was seen to be associated with bleeding and periovarian adhesions. Fertility rate is poor compared to other procedures.
 
Electrocauterization
Here monopolar cautery is used at 20 to 30 watts in cutting mode. Pure cutting current may be used on the thick ovarian surface. Cortex is usually penetrated at 4 to 10 sites for a depth of 3 to 5 mm. Attempt is made to puncture almost all the prominent follicles. The fluid mixed with the irrigating solution is completely aspirated out through the suction cannula. The ovaries are lavaged with Ringer's solution containing hydrocortisone and heparin to minimize adhesions. The bleeding is usually self-limiting and is arrested by diathermy if required.
 
Laser
KTP and CO2 lasers are used and the technique is similar to that of electrocautery, namely the ovarian cortex is vaporized over the follicles. As the energy is more precisely focused, there is less peripheral thermal damage. Either an ultrapulse CO2 laser (40–80 watts, 25–200 MJ) or the super pulse CO2 (25–40 watts) is used. All visible subcapsular follicles are vaporized and a 2 to 4 mm crater is made in the ovarian stroma. It is recommended that the number of punctures be more, 25 to 40, so that all visible follicles are drained. With Nd:YAG laser the technique is somewhat different. There is much more thermal diffusion in the non-contact mode with this laser. Since it is divergent once it passes the tip of the delivery system, the coagulation of the tissue is achieved. A 150wedge-shaped area of 4 to 10 mm is thus coagulated without opening the cortex. This laser has also been used in the contact mode to cut out a wedge-shaped portion of the ovary, not unlike the bilateral ovarian resection. The laser is now regarded to produce more excessive ovarian surface trauma than stromal damage leading to adhesion formation.
As a result, attention is now shifting toward the use of monopolar needle electrodes, which are insulated where they contact the ovarian surface. The needle punctures the ovarian surface to the depth of the insulated hub and the deeper stroma can be cauterized with minimal surface damage. The most commonly used instrument is the monopolar needle.
 
Unilateral vs Bilateral Diathermy
It was seen that unilateral diathermy restored bilateral ovarian activity with the contralateral ovary often being the first to ovulate after the treatment. Unilateral ovarian diathermy was as effective and long lasting as bilateral ovarian diathermy in the resumption of menstruation and pregnancy rates.2
 
Complications
 
Postoperative Adhesion
A rate of adhesion formation of 19.3% was seen that decreased to 16.6% when the peritoneal lavage was done.3 The greater the damage to the surface of the ovary more the peritubal adhesion formation. Hence, Armar recommended only 4 diathermy points per ovary for 4 seconds at 40 W.4 A recent study reported a high rate of adhesion formation up to 60% and their extent and severity was not influenced by the number of ovarian punctures; however, the left ovary appeared more prone to develop severe adhesions than the contralateral one.5151
Wedge resection has been seen to cause adhesions in nearly 100% cases. To prevent adhesions 200 mL of Hartmann's solution is instilled in the Pouch of Douglas that by cooling the ovary prevents heat injury to adjacent tissues and reduces adhesion formation. The risk may further be removed by abdominal lavage. Laser treatment may have a lower adhesion formation rate than diathermy.
The risks of periovarian adhesions can be reduced by minimizing the ovarian surface damage. This is achieved by reducing the total number of punctures, and ensuring that the needle enters perpendicular to the surface, not tangentially.
 
Ovarian Failure
Ovarian failure has been reported by some.6 If FSH and LH levels are not too high, avoid too many punctures as it may lead to ovarian failure. The incidence is very uncommon.7
However, a recent study stated most of the changes in the ovarian reserve markers observed after LOD could be interpreted as normalization of ovarian function rather than a reduction of ovarian reserve. LOD, if applied properly, normalizes the exaggerated ovarian morphologic and endocrinologic properties.8
The risk of premature ovarian failure can be reduced by minimizing the thermal damage to the ovary. This is achieved by reducing the number of punctures, using only very short bursts of cutting current and lavaging the ovary to cool it down.
 
Advantages
  1. In subsequent ovulation induction ovaries become more responsive and lower dose of drugs are needed to induce ovulation.
  2. There is decreased pregnancy loss.152
  3. No risk of hyperstimulation or multiple pregnancies.
  4. Effect lasts for 12 to 18 months. Recent studies have shown a beneficial effect up to 9 years.9
 
Results of Laparoscopic Treatment
 
Ovulation
Ovulatory rate that varies between 70 to 92% is influenced by body weight. Ovulation rate being higher in slim and moderately obese but lower in obese patients.1
 
Pregnancy Rate
A pregnancy rate of 40 to 80% is seen.1 In cases with clomiphene resistance pregnancy rate was similar with laparoscopic ovarian drilling and gonadotropins.10 However, a recent study showed a 67% cumulative pregnancy rate with gonadotropins after 6 months but only a 37% pregnancy rate with laparoscopic ovarian drilling.11
 
Abortion
The abortion rate is 6 to 7% with laparoscopic ovarian drilling compared to 26–28% in treatment with gonadotropins.12
Keeping in view the problems of postoperative periovarian adhesions and ovarian atrophy a combined approach for ovulation induction is recommended whereby low-dose diathermy is followed by low dose ovarian stimulation.13
Ovarian drilling by hydrolaparoscopy is an effective treatment for CC-resistant.14 A recent Cochrane review (2007) stated that there was no evidence of a difference in live birth, clinical pregnancy or miscarriage rate between LOD and gonadotropins. Multiple pregnancy rates were lower with ovarian drilling than with gonadotropins (1% versus 16%) making it an attractive option. However, there are 153ongoing concerns about long-term effects of LOD on ovarian function.15
 
Follow-up
If in 12 ovulatory cycles there is no conception, one should proceed for assisted reproductive techniques rather than waiting.
Surgical treatment for PCOS has a definite role in management of the infertile women with the advantage of no subsequent cycle monitoring. Caution during the procedure is needed to avoid ovarian damage that may cause adhesions and ovarian failure.
 
References
  1. Gjonnaess H. Polycystic ovarian syndrome treated by ovarian electrocautery through the laparoscope. Fertil Steril. 1984;41:20–5.
  1. Al-Mizyen E, Grudzinskas JG. Unilateral laparoscopic ovarian diathermy in infertile women with clomiphene citrate-resistant polycystic ovary syndrome. Fertil Steril. 2007;88(6):1678–80.
  1. Naether OGJ, Fischer R, Weise HC, Geiger-Kotzler L, Delfs T, Rudolf K. Laparoscopic electrocoagulation of the ovarian surface in infertile patients with polycystic ovarian disease Fertil Steril. 1993;60:88–94.
  1. Armer NA, Mc Garrigle HH, Honour J, Holownia P, Jacobs HS, Lachelin GC. Laparoscopic ovarian diathermy in management of anovulatory infertility in women with polycystic ovaries: endocrine changes and clinical outcome. Fertil Steril. 1990;53:45–9.
  1. Mercorio F, Mercorio A, Di Spiezio Sardo A, Barba GV, Pellicano M, Nappi C. Evaluation of ovarian adhesion formation after laparoscopic ovarian drilling by second-look minilaparoscopy. Fertil Steril. 2008;89(5):1229–33.
  1. Dabirashrafi H. Complications of laparoscopic ovarian cauterization. Fertil Steril. 1989;52:878–83.
  1. Cohen BM. Laser laparoscopy for polycystic ovaries. Fertil Steril. 1989;52:167–8.
  1. Api M. Is ovarian reserve diminished after laparoscopic ovarian drilling? Gynecol Endocrinol. 2009;25(3):159–65.
  1. Amer SAKS, Banu Z, Li TC, Cooke ID. Long-term follow-up of patients with polycystic ovarian syndrome after laproscopic ovarian diathermy: endocrinal and ultrasonic outcomes. Hum Reprod. 2002;11:2851–7.
  1. Ferquhar CM, Williamson K, Gudex G, Johnson NP. A randomized controlled trial of laparoscopic ovarian diathermy versus gonadotropin therapy for women with clomiphene citrate resistant polycystic ovary syndrome. Fertil Steril. 2002;78:404–11.
  1. Bayram N, van Wely MK, Kajik EM, Bossuyut PMM, van der Veen. Using an electrocautery strategy or recombinant follicle stimulating hormone to induce ovulation in polycystic ovarian syndrome: randomized controlled trial. Br Med J. 2004;328:192–5.
  1. Abdel GA, Mowafi RS, Alnaser HM, Alrashid AH, et al. Ovarian electrocautery versus HMG and pure FSH therapy in treatment of PCOS. Clin Endocrinol. 1990;33:585–92.
  1. Farhi J, Soule S, Jacob H. Effect of laparoscopic ovarian electrocautry on ovarian response and outcome of treatment with gonadotropins in clomiphene resistant patients with PCOS. Fertil Steril. 1995;64:930–5.
  1. Poujade O, Gervaise A, Faivre E, Deffieux X, Fernandez H. Surgical management of infertility due to polycystic ovarian syndrome after failure of medical management. Eur J Obstet Gynecol Reprod Biol. 2011;158(2):242–7.
  1. Farquhar C, Lilford RJ, Marjoribanks J, Vandekerckhove P. Laparoscopic ‘drilling’ by diathermy or laser for ovulation induction in anovulatory polycystic ovary syndrome. Cochrane Database Syst Rev. 2007;(3):CD001122.

Hypogonadotropic Hypogonadism and Ovulation Inductionchapter 10

Surveen Ghumman
Hypogonadotropic hypogonadism is a disorder characterized by low or undetectable LH or FSH levels leading to low estradiol levels and anovulation. These women respond well to ovulation induction.
 
Causes
Causes could be pituitary or hypothalamic. Common functional disturbance like eating disorders, emotional stress or excessive exercise are responsible in most cases. Exercise increases prolactin, GH, testosterone, ACTH, adrenal steroids, and endorphins and decreases gonadotropins (Figs 10.1 and 10.2).
156
Fig. 10.1: Causes of hypogonadotropic hypogonadism
Fig. 10.2: Mechanism of exercise induced hypogonadotropic hypogonadism
157
 
Clinical Presentation
Clinical presentation depends on degree of GnRH suppression (Table 10.1).
  1. Mild GnRH suppression—Inadequate luteal phase.
  2. Moderate GnRH suppression—Anovulation and menstrual irregularity.
  3. Profound GnRH suppression—Amenorrhea. Failure to demonstrate withdrawal bleeding with progesterone.
Other symptoms like vaginal dryness, hot flushes, decreased breast tissue, libido and muscle mass may present. Decreased bone density is also seen due to low estrogen levels.
 
Diagnosis
 
Laboratory Parameters
Tests done are mainly hormonal and imaging so as to establish diagnosis and to identify cause:
  1. Low gonadotropins (LH and FSH).
  2. Low estradiol levels.
    Table 10.1   Grades of severity of hypothalamic amenorrhea
    Grade I :
    Positive response with clomiphene
    Grade Ia: Normal ovulatory
    Grade Ib: Ovulatory with LPD
    Grade Ic: Anovulatory
    Grade II:
    Progesterone withdrawal + but clomiphene response negative
    Grade III:
    Response to 100 mg IV bolus
    Grade IIIa :
    Normal adult type
    Grade IIIb:
    Blunted prepubertal type
    Grade IIIc:
    No response
    158
  3. Normal prolactin and imaging.
  4. Normal testosterone and DHEA
  5. Pituitary dynamic testing:
    1. Gonadotropin-releasing hormone challenge test: IV bolus injection of 100 μg of GnRH is given. LH and FSH measured at regular interval of 0, 30, 60, 90 min, respectively after application of GnRH.
    2. Pituitary capacity test: Differentiates pituitary cause from hypothalamic origin. An IV bolus of GnRH 100 μg is given followed by subsequent stimulation by pulsatile GnRH 15 μg/90 min for 4 days to prime pituitary. One week after discontinuation of stimulation pituitary challenge test is repeated. If second response same or lower than first pituitary gonadotrophs are deficient.
  6. Bone mineral density: Bone mineral density shows lower values as these women are estrogen deficient.
 
Differential Diagnosis of Hypogonadotropic Hypogonadism with Amenorrhea
Conditions like hyperprolactenemia, hypothyroidism, PCOS and ovarian failure can mimic this condition (Table 10.2).
 
Treatment
These women have anovulatory infertility. This can be treated by GnRH therapy or replacement of gonadotropins. The important thing to be kept in mind with these women is: 159
Table 10.2   Differential diagnosis of hypogonadotropic hypogonadism
Tests
Rules out or confirms
LH/FSH/Prolactin/TSH
Excludes ovarian failure, hyperthyroidism,
hypothyroidism, prolactin secreting adenoma
Androgen –Testosterone, androstenedione, insulin
PCOS
Corticotropin stimulation test, or perform dexamethasone suppression test
Cushing's syndrome/Addison's disease
Pituitary stimulation test
Panhypopituitarism
All normal except elevated 24 hour urinary cortisol
Functional hypothalamic chronic anovulation
  1. Low values of LH and their impact on ovulation, fertilization and pregnancy rates.
  2. The other aspect is the effect of low estradiol level on the pregnancy rate. Estradiol does not seem to be important for the fertilization but it is highly essential for pregnancy and its maintenance.1,2 It is important for a normal endometrial receptivity.
 
Gonadotropin Therapy
 
Role of LH Supplementation
LH supplementation is essential in follicular phase as low endogenous levels of LH acts on corpus luteum to produce progesterone which prepares the endometrium. One key aspect of the effect of LH activity on the developing follicle is the follicle-stimulating hormone (FSH) mediated acquisition of LH receptors in granulosa cells that occurs in the mid-follicular 160phase of the normal menstrual cycle, once ovarian follicles reach a diameter of 10 mm. At this stage LH can stimulate granulosa cell function and folliculogenesis independently of FSH activity.
Stimulation without LH induces normal follicular growth with accompanying low estradiol level leading to absence of normal oocyte maturation and fertilization competence.3 The poor oocyte quality is because, proper maturation of oocyte requires both gonadotropins.4 Optimal follicular development is obtained if exposure to endogenous and/or exogenous LH is sufficient. This is the ‘threshold’ concept. However, it is seen that excessive levels of LH inhibit follicular growth. This is the “ceiling” concept, i.e. level beyond that normal follicular growth and oocyte quality will be compromised by LH. Follicular development was achieved in 65.4% of patients receiving lutropin alfa and 15.4% of patients receiving placebo.5
 
Which Gonadotropin Should be Used?
Gonadotropins that can supplement LH are used:
  1. hMG: hMG, with a fixed LH:FSH ratio, represents a non-physiological stimulation where the ratio of LH and FSH is fixed. However, in the natural cycle early part of the follicular phase is FSH dominated and latter part has increasing levels of LH that are important (Fig. 10.3).
  2. Stimulation with recombinant FSH and LH
  3. hCG supplementation: Activity of LH can also be obtained by the addition of hCG (50 IU/day) as has been shown in a woman with secondary amenorrhea.6
 
What Dose of Gonadotropins is Needed?
Higher doses of gonadotropin and longer duration of stimulation are required as there is no endogenous production. 161In IVF higher doses given for a longer time yielded lower peak E2 levels and lesser number of oocytes compared to women with unexplained infertility (Fig. 10.3).7
 
At What Level should LH Supplementation be Started?
A study showed that rhLH is indicated in LH deficient women (defined by an endogenous LH level < 1.2 IU/L).8 A transition from LH dependence to independence was observed between basal LH values of ≥1.2IU/L and ≤1.6IU/L.9
Fig. 10.3: hMG protocol for hypogonadotropic hypogonadism
162
 
What Dose of LH should be Given?
  • A study carried out in women with hypogonadotropic hypogonadism showed that when there was no LH supplementation E2 levels on last day of FSH were only 65 pmol/L. This level increased marginally to 195 pmol/L on supplementing with 25 IU of LH but increased to 1392 pmol/L when a supplementation of 75 IU was given. Difference in serum E2 levels resulted in different endometrial growth. The follicular growth increased from 27% to 79% with supplementation between 75 IU and 225 IU and endometrium became excellent. They also responded better to hCG permitting successful luteinization of follicles.
  • Daily dose of 75 IU rLH is sufficient for promoting optimal follicular development in the majority of HH patients. High doses of exogenous LH greater than 225 IU lead to atresia of secondary follicles LH ceiling concept. Androstenedione levels were higher among patients treated with the higher doses of LH.10
 
How Do We Monitor LH Dose?
Such low doses do not lead to a measurable change in serum LH trough levels and hence, are difficult to monitor.10
 
Adverse Effects
Most adverse events are mild to moderate in severity.11 Adverse events were seen in same incidence as women taking only FSH and consisted of pelvic and abdominal pain, headache, breast pain, nausea, ovarian enlargement and somnolence.10
 
Results of Gonadotropin Therapy
Cumulative PR after 6 treatment cycles was 89%. Severe OHSS occurred in 1% and multiple pregnancy rate in 30%.12163
 
Poor Response with Gonadotropins and Role of Growth Hormone
Growth hormone (GH) replacement may be beneficial in hypopituitarism when response to conventional treatment is absent. A study showed that GH (1 IU/day) alone for 3 months followed by GH and hMG gave good results. Other have tried higher doses up to 12 IU/day.13,14
 
GnRH Therapy
GnRH therapy needs an intact pituitary gland. It can be administered by an autosyringe or a peristaltic pump.
 
Dose and Frequency of Administration
Depends on:
  1. Characteristics of amenorrhea.
  2. Pulse frequency and dose.
  3. Route of administration.
 
Characteristic of Amenorrhea
The classification shown in Table 10.1 determines the dose needed. Grade II to IIIb requires 2.5–5 μg per pulse but for grade IIIc 15–20 μg per pulse is required.
 
Pulse, Frequency and Dose
With 60 μg/day dose of GnRH a 60 minutes interval gave a 94% ovulation and a 120 minute interval gave a 70% ovulation. A low frequency of pulsatile GnRH in women decreases mean LH levels, blunts the midcycle gonadotropin surge, does not increase follicle-stimulating hormone concentrations, and is associated with a reduced rate of ovulation (Fig. 10.4).15 164
Fig. 10.4: GnRH pulsatile therapy: Suitable for women with intact pituitary gland
Higher dose leads to increased ovulation rates and multiple pregnancy.
 
Route of Administration—IV or SC Administration
Earlier and higher LH peaks with IV than SC administration are seen. Daily resorption from subcutaneous sites may lead to a continuous pattern resulting in desensitization. Irreversible loss of 30% may be seen in SC infused GnRH because of local catabolism of the hormone by GnRH proteases. Site of SC administration—upper arm vs abdomen—has twice the concentration. GnRH antibodies may form with SC therapy because of binding of GnRH to large protein molecules around tip of the needle. SC therapy requires higher pulse doses and prolonged duration of treatment and results in lower ovulation and pregnancy rates.165
 
Failure of Response to GnRH Therapy
Combined use of Pulsatile GnRH and gonadotropins: If there is no response to GnRH therapy a combination of pulsatile GnRH with gonadotropins can be given. An early rise in estradiol keeps the FSH levels suppressed below threshold level for ongoing follicular growth. Addition of hMG may bring these above threshold resulting in ovulation.
 
Advantages of GnRH Pulsatile Therapy
It induces a menstrual cycle like the natural one with monofollicular development, physiological estradiol levels and subsequent luteal phase characteristics similar to a normal cycle. The cumulative pregnancy rate was higher with this than hMG stimulated cycle.16
  1. High ovulation rate – 90%
  2. High pregnancy rate – 23% per cycle. Cumulative conception rate – 6 cycles – 78%, 12 cycles – 93%
  3. Multiple pregnancy rate – lower
  4. Low abortion rate
  5. No OHSS
  6. No monitoring
  7. No frequent office visits
  8. Avoidance of gonadotropin therapy.
 
Disadvantages of GnRH Pulsatile Therapy
  1. Carrying an infusion pump requires adaptation. Unfortunately, many patients displayed psychological problems and were irritated by the carrying of an electronic pump on their body for several days or weeks.
  2. Slight risk of infection bacteremia or sepsis.
  3. Reaction at skin site.
  4. Hematoma.
  5. Development of GnRH antibodies.166
 
Monitoring of Thyroid Activity
Constant monitoring of thyroid to maintain free thyroxine at upper limit of normal is done.
 
Long-term Therapy
  • Hormone replacement therapy for bone health is needed. Bone loss is greater in the initial period, hence, early treatment is required.
  • Exercise, adequate diet, calcium supplementation and counseling and support are also essential.
 
Pregnancy in Cases with Hypogonadotropic Hypogonadism
 
Pregnancy Monitoring
  • Luteal and early pregnancy progesterone support is a must in these cases. Luteal support with progestogen is continued till 12 weeks
  • Thyroxine intake is raised to 150 μg/day in first and 200 μg/day in second trimester to maintain free thyroxine at a higher limit of normal 1.5 ng/ml.
 
Pregnancy Complications
A study showed that pregnancies in hypopituitary women had the following complications
  • Postpartum hemorrhage – 8.7%,
  • Transverse lie in 16%,
  • Small for gestational age – 42.4%
  • Uterine dysfunction caused by hormone deficiency: In women with severe hypopituitarism oxytocin supplementation was given in third trimester with the 167aim to establish physiologic conditions and to prevent postpartum uterine inertia.17
Women with hypogonadotropic hypogonadism may present with a spectrum of presentations according to the severity. They respond well to ovulation induction therapies. LH supplementation must be done in these women. Pregnancies in these women require special care and must receive luteal support.
 
References
  1. Sonntag B, Loebbecke KC, Nofer JR, Kiesel L, Greb RR. Serum estradiol and progesterone in the mid-luteal phase predict clinical pregnancy outcome in IVF/ICSI cycles. Gynecol Endocrinol. 2013;29(7):700–3.
  1. Murray AA, Swales AKE, Smith RE, Molinek MD, Hillier SG, Spears N. Follicular growth and oocyte competence in the in vitro cultured mouse follicle: Effects of gonadotropins and steroids. Mol Hum Reprod. 2008;14(2):75–83.
  1. Balasch J, Miró F, Burzaco I, et al. The role of luteinizing hormone in human follicle development and oocyte fertility: Evidence from in-vitro fertilization in a woman with long-standing hypogonadotrophic hypogonadism and using recombinant human follicle stimulating hormone. Hum Reprod. 1995;10(7):1678–83.
  1. Huirne JA, van Loenen AC, Schats R, McDonnell J, Hompes PG, Schoemaker J, et al. Dose-finding study of daily GnRH antagonist for the prevention of premature LH surges in IVF/ICSI patients: optimal changes in LH and progesterone for clinical pregnancy. Hum Reprod. 2005;20(2):359–67.
  1. Shoham Z, Smith H, Yeko T, O'Brien F, Hemsey G, O'Dea L. Recombinant LH (lutropin alfa) for the treatment of hypogonadotrophic women with profound LH deficiency: A randomized, double-blind, placebo-controlled, proof-of-efficacy study. Clin Endocrinol. 2008;69(3):471–8.
  1. Filicori M, Cognigni GE, Taraborrelli S, Spettoli D, Ciampaglia W, de Fatis CT. Low-dose human chorionic gonadotropin therapy can improve sensitivity to exogenous follicle-stimulating hormone in patients with secondary amenorrhea. Fertil Steril. 1999;72L:118-1120.
  1. Kumbak B, Kahraman S Women with hypogonadotropic hypogonadism: cycle characteristics and results of assisted reproductive techniques. Acta Obst Gyne Scand. 2006;85:1453–7.
  1. Loumaye E. Ovarian stimulation: is exogenous LH necessary in all patients?. Gynecol Obstet Fertil. 2002;30(11):890–5.
  1. O'Dea L, O'Brien F, Currie K, Hemsey G. Follicular development induced by recombinant luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in anovulatory women with LH and FSH deficiency: evidence of a threshold effect. Cur Med Res Opin. 2008;24:2785–93.
  1. The European Recombinant Human LH Study Group. Recombinant human luteinizing hormone (LH) to support recombinant human follicle-stimulating hormone (FSH)-induced follicular development in LH- and FSH-deficient anovulatory women: a dose-finding study. J Clin Endocrinol Metabol. 1998;83:1507–14.
  1. Dhillon S, Keating GM Lutropin alfa, Drugs. 2008;68(11):1529–40.
  1. Messinis IE. Ovulation induction: a mini review. Hum Reprod. 2005;20:2688–97.
  1. Salle A, Klein M, Pascal-Vigneron V, Dousset B, Leclere J, Weryha G. Successful pregnancy and birth after sequential cotreatment with growth hormone and gonadotropins in a woman with panhypopituitarism: a new treatment protocol. Fertil Steril. 2000;74(6):1248–50.
  1. Park JK, Murphy AA, Bordeaux BL, Dominguez CE, Session DR. Ovulation induction in a poor responder with panhypopituitarism: a case report and review of the literature. Gynecol Endocrinol. 2007;23(2):82–6.
  1. Filicori M, Flamigni C, Campaniello E, Ferrari P, Meriggiola MC, Michelacci L, et al. Evidence for a specific role of GnRH pulse frequency in the control of the human menstrual cycle. Am J Physiol. 1989;257(6 Pt 1):E930–6.
  1. Martin KA, Hall JE, Adams JM, Cowley WF Jr. Comparison of exogenous gonadotropins and pulsatile gonadotropin-releasing hormone for induction of ovulation in hypogonadotropic amenorrhea. J Clin Endocrinol Metab. 1993;77(1):125–9.
  1. Kübler K, Klingmüller D, Gembruch U, Merz WM. High-risk pregnancy management in women with hypopituitarism. 2009;29:89–95.

Hyperprolactinemiachapter 11

Surveen Ghumman,
Ritika Kaur
Hyperprolactinemia is the condition when there is increased prolactin secretion from pituitary lactotrophs that may lead to galactorrhea, amenorrhea or infertility. Prolactin is a single chain polypeptide containing 199 amino acids similar to growth hormone and placental lactogen in structure. It is secreted by lactotroph cells from the anterior pituitary. This secretion is controlled by the hypothalamus via prolactin inhibiting neurotransmitter, dopamine that acts on the D2 receptors. Gamma-aminobutyric acid is also inhibitory. The stimulatory effect on prolactin secretion is through thyrotropin releasing hormone, vasoactive intestinal peptide, and angiotensin II. Prolactin circulates in various forms with structural modification that are the result of glycosylation, phosphorylation, deletion and addition. They differ in their bioactivity (assessed by clinical symptoms like galactorrhea and amenorrhea) and immunoreactivity (recognized by immunoassay). Prolactin is found in serum in different molecular forms differing in molecular size, i.e. monomeric prolactin (molecular mass 23 kDa), “big prolactin” (50–60 kDa, possibly a dimer or a complex with receptor) and “big-big prolactin” or “macroprolactin” (150–170 kDa), usually 171a complex with immunoglobulin G. The little prolactin constitutes 80% of the prolactin and is biologically most active. The big prolactin account for 10 to 25% of the hyperprolactinomas reported.
 
Prolactin and Ovarian Function
High levels of prolactin in proliferative phase may interfere with follicle and oocyte development, cause atresia of dominant follicle and inhibit ovulation. In the secretory phase, high levels of prolactin interfere with corpus luteal function, downregulate LH receptors, and cause premature destruction of corpus luteum leading to corpus luteum deficiency.
 
Incidence
Hyperprolactinemia occurs in 0.4% of normal population and 9 to 17% of patients with reproductive disorders. It is seen in 40% of PCOS patients.
 
Etiology
  1. Physiological
    1. Pregnancy and lactation
    2. Chest wall stimulation, i.e. by sucking
    3. Sleep
    4. Stress.
  2. Pathological
    1. Hypothalamic pituitary stalk damage
      1. Tumors: They disrupt the inhibitory influence of dopamine on prolactin secretion—craniopharyngioma, supraseller pituitary mass extension, meningioma, dysgerminoma, and metastasis.172
      2. Empty sella syndrome
      3. Lymphocytic hypophysitis
      4. Adenoma with stalk compression
      5. Granulomas
      6. Irradiation
      7. Trauma.
    2. Pituitary hypersecretion
      1. Prolactinoma
        1. Micro-adenoma: These tumors are less than 1 cm in diameter, do not invade parasellar area and resolve spontaneously.
        2. Macro-adenoma: These are tumors more than 1 cm in diameter, locally invasive and compress adjacent structures.
      2. Acromegaly
      3. Hyperplasia of lactotrophs.
    3. Systemic disorders
      1. Chronic renal failure
      2. Hypothyroidism
      3. Cirrhosis
      4. Epileptic seizures.
    4. latrogenic causes: Drug-induced hypersecretion:
      1. Dopamine receptor blockers: Phenothiazines, butyrophenones, thioxanthenes, metoclopromide
      2. Dopamine synthesis inhibitors
      3. Catecholamine depletors
      4. Opiates
      5. H2 antagonist: Cimetidine, ranitidine
      6. Imipramines: Amitryptilene, amoxapine
      7. Serotonin re-uptake inhibitors: Fluoxetine
      8. Calcium channel blockers: Verapamil
      9. Hormones: Estrogens, oral contraceptive pills.
173
 
Idiopathic Hyperprolactinemia
Hyperprolactinemia in patients where no cause can be found is termed as idiopathic. Usually prolactin level never exceeds 100 ng/mL. It could be because of fluctuating levels of prolactin or macroprolactinemia where the large prolactin molecule is immunoreactive but not biologically active.
 
Transient Hyperprolactinemia
Transient preovulatory rise is seen in prolactin levels with some patients of unexplained infertility. This lasted 2 to 3 days and coincided with estradiol peak. 40% of patients conceived with bromocriptine as compared to 1% in controls.1
 
Clinical Features of Hyperprolactinemia
  1. Menstrual irregularities like amenorrhea and oligomenorrhea.
  2. Galactorrhea: 33% women with hyperprolactinemia have galactorrhea.
  3. Infertility.
  4. Estrogen deficiency—Vaginal dryness, dysparenia, osteoporosis (if hyperprolactinemia persists for a long duration).
  5. Reduced libido.
  6. Delayed puberty.
  7. Visual defects: They occur in 5% women due to CNS compression.
 
Hyperprolactinemia and Infertility
Hyperprolactinemia inhibits ovulation leading to menstrual irregularities as well as amenorrhea. High prolactin levels directly inhibit the amount of GnRH secretion. This results 174in inhibition of LH and FSH release leading to anovulation and hypogonadism. The high prolactin levels interfere with the positive effect of estrogen on midcycle surge and may directly inhibit ovarian steroidogenesis. It hinders normal corpus luteum formation by directly acting on the ovary leading to luteal phase defect and recurrent pregnancy loss.2 Mild transient hyperprolactinemia is seen in 30% of PCOS patients. As prolactin concentration increases women progressively show increasingly severe reproductive abnormality (Fig. 11.1).
Ovulatory cycles and regular menses are achieved within 6 months of therapy with 80% of patients. Infertile women with galactorrhea and normal prolactin have responded to treatment.3 The choice of treatment for infertility is medical. About 60–80% patients with hyperprolactinemia become ovulatory with bromocriptine. There is a good response to clomiphene induction in previously resistant cases. It is a safe drug in pregnancy. In some studies, bromocriptine has been given only in follicular phase.
Fig. 11.1: Progressive reproductive abnormality seen in hyperprolactinemia
175
 
Diagnostic Evaluation
 
Serum Prolactin Estimation
Normal level is 1 to 20 ng/mL. It has a circadian rhythm with maximum levels between 3 AM to 9 AM; thus, sample should be collected between 9 AM and noon. It is to be done in fasting state with no prior breast or pelvic examination, exercise or sexual activity.
 
Fallacy
  1. Occasionally in presence of a large pituitary tumor falsely low levels of serum prolactin are caused by an effect known as ‘high dose hook effect’ where extremely large doses of prolactin prevent accurate assessment by antibody assay. Serial dilutions may reveal a high level.
  2. Sometimes, high levels of relatively inactive prolactin in absence of tumor can be due to the creation of macromolecules of prolactin by antiprolactin autoantibodies.4
 
What is Macroprolactin?
Hyperprolactinemia can occur in physiological and pathological conditions. Prolactin has 3 different isoforms—little prolactin, big prolactin and big-big prolactin. Macroprolactin is a complex of little prolactin and an immunoglobulin G antibody and the weight of the complex is more than 150 kDa. In physiological condition, macroprolactin comprises up to 1% of all circulating prolactin in the serum blood. In pathological condition, percentage of macroprolactin can increase in the serum blood. The prevalence of macroprolactinemia is found in around 10–26% of all patients with hyperprolactinemia. Women with high serum prolactin concentration should be 176screened for macroprolactinemia. Presence of macroprolactin should always be suspected when a patient's clinical history and/or radiological data are incompatible with his/her prolactin value. Thus, it may be useful to screen all patients with high sera prolactin levels by tests, such as the polyethyleneglycol precipitation method although gel filtration chromatography remains the gold standard. Macroprolactinemia doesn't require any pharmacological treatment or other medical procedures. Confirmation prevents unnecessary procedures such as laboratory controls, MRI of the pituitary, treatment with dopamine agonists or even pituitary surgery.5,6
 
T3, T4 and TSH
It is done to rule out compensated or primary hypothyroidism. Both of them may be found in cases of hyperprolactinemia.
 
Goldmann's Perimetry
Visual field defects may be detected.
 
Radioimaging of the Sella Turcica
 
Cone Down view X-ray of Sella Turcica
With newer accurate imaging techniques this is no longer the investigation of choice. If the view is normal, then it is unlikely there is an extrasellar extension of the tumor. Macroadenoma may show imaging findings of an asymmetrically enlarged pituitary fossa with a double contour to its floor and erosion of the clinoid process.7,8
 
MRI/CT Scan of the Sella Turcica
MRI with gadolinium is the best imaging modality.9 CT scan can detect microadenomas of 2 mm diameter and presence 177of any suprasellar extension. MRI has a resolution of 1 mm and is more sensitive than CT. It also avoids radiation. The indications for this test are:
  1. Serum prolactin level > 100 ng/mL.10
  2. Presence of headaches and visual field defects.
  3. Abnormal X-ray cone down view of the sella turcica.
With increasing use of these modalities, incidental discovery of pituitary microadenomas is seen in 10% of individuals having normal prolactin levels. These tumors are called pituitary incidentalomas.7
Macroadenoma by definition is more than 1 cm and imaging techniques now identify suprasellar extensions, compression of optic chiasma and invasion of cavernous sinus.
 
Treatment of Hyperprolactinemia (Fig. 11.2)
 
Aim
  • Elimination of symptoms like galactorrhea and amenorrhea
  • Induction of ovulation
  • Treatment of prolactin secreting macroadenomas.
The management options for hyperprolactinemia are:
  • Expectant
  • Medical
  • Surgical
  • Radiation.
 
Management of Hyperprolactinemia due to Antipsychotic Drugs
The pituitary is imaged to rule out prolactinoma. The drug is changed to an alternative drug such as an atypical neuroleptic or be discontinued. Serum prolactin levels are monitored to ensure they are not rising further. Low dose contraceptive pill may be given if estrogen deficiency is present. The use of dopamine agonist along with an antipsychotic drug may 178lower the prolactin levels but would antagonize the effect of the anti-psycotic drug.
Fig. 11.2: Management of hyperprolactinemia
179
Dopaminergic agents can occasionally induce or worsen psychotic symptoms.
 
Expectant Management
Where no tumor is seen on imaging and there is absence of symptoms like galactorrhea, infertility, menstrual disturbance or hypoestrogenism one can use the expectant line of management with serial monitoring of prolactin levels and a CT scan every 2 years.
 
Medical Management
 
Bromocriptine
It is a derivative of lysergic acid substituted with bromine at position 2. It is a dopamine agonist that binds to dopamine receptors and therefore, induces inhibition of pituitary prolactin secretion.
Dosage
  1. Orally
    1. Tablet of 2.5 mg given in a twice daily dose as half-life is 8 to 12 hours. It can be increased to 10 mg/day.
    2. Slow release oral preparation is also available to be given once a day in a dose of 5 to 15 mg/day and is equally effective.
  2. Long-acting depot intramuscular injection: They are made by embedding glucose initiated polygycolide microspherules and have a maximum degradation time of 3 months. They are given in a dose of 50 to 75 mg/month. Since response to these injections is rapid, they are useful in large tumors with visual field impairment.10 It has the same severity of side effects as the oral preparation.180
  3. Intravaginal: It is given in a similar dose of 5 to 10 mg/day. Since it avoids direct contact with the intestinal mucosa, it has lesser side effects than oral administration. The levels are sustained for a longer time as it escapes the liver first pass effect when given vaginally and therapeutic results are achieved at a lower dose.
Side effects
10% patients show intolerable side effects (Table 11.1).
Measures to reduce side effects are:
  • Building tolerance by slowly increasing the dose at weekly intervals.
  • Taking the drug at bedtime as peak levels are achieved after 2 hours.
  • Individualizing the dosage schedule
  • Using intravaginal route.
Table 11.1   Side effect of bromocriptine
Immediate effects
• Nausea
• Headache
• Fatigue
• Dizziness
• Orthostatic hypotension
• Nasal congestion
• Vomiting
• Abdominal cramps
• Hallucinations
Long-term effects11
• Raynaud's phenomenon
• Constipation
• Psychiatric changes specially aggression
181
Efficacy
  1. Amenorrhea and galactorrhea: 80% of patients with amenorrhea and galactorrhea improved.12 There was 75% reduction in breast secretion by 6 weeks and galactorrhea was suppressed in 60% of patients by 12 weeks.
  2. Infertility: Ovulation was restored within 5 to 6 weeks. Studies have shown successful ovulation induction and pregnancy with bromocriptine in the absence of galactorrhea or hyperprolactinemia in previous nonresponders to clomiphine.13
  3. Pituitary tumor: Bromocriptine causes regression of macroadenomas. In some shrinkage is seen even with low dose (5–7.5 mg/day) whereas in others larger doses and prolonged duration may be required. Usually a dose of more than 10 mg is ineffective. Visual improvement occurs within days and tumor shrinkage occurs rapidly in first 3 months of therapy. Locally invasive tumors with levels of bromocriptine more than 1000 ng/mL show a good response to medical treatment. Problem with treatment of the tumor with this drug is that it has to be taken indefinitely. No adverse effect of bromocriptine has yet been seen as in early pregnancy.14 After 2 years therapy, 75% of microadenomas and 80 to 90% of macroadenomas regress. On discontinuation of drug after 5 years only 25% patients remained normoprolactinemic.15
Problems
  1. Recurrence: Recurrence of symptoms occurred in 75% of patients with prolactinomas within 4 to 6 weeks on stopping of the drug. Hence, these drugs are used only for short-term purpose of achieving pregnancy, curing galactorrhea or reducing tumor mass.16
  2. Resistance: 5 to 18% of patients do not tolerate bromocriptine or are resistant to it because of decreased 182dopamine receptor on lactotroph cell membrane. In these cases other drugs can be tried.
  3. Perivascular fibrosis: It may cause perivascular fibrosis in tumors if given for a long time making surgery difficult.
 
Other Dopamine Agonists
  1. Pergolide: It is more potent, longer-acting, better tolerated and useful in bromocriptine resistant patients. It is given in a dose of 50 to 150 mg/day and is increased slowly to avoid side effects.
  2. Quinagolide: It is a long-acting non-ergot derivative, with higher affinity for dopamine receptors and lesser side effects. It is useful in bromocriptine resistant tumors. It also has antidepressant properties and is given in a dose of 75 to 300 mg/day at bedtime.
  3. Cabergoline: It is useful in bromocriptine resistant cases. It has less side effects compared to bromocriptine, the most common being headache. It is more effective in reducing tumor size and prolactin levels than bromocriptine or quinagolide.17 It is given in a dose of 0.5 to 3 mg once a week orally or vaginally, usually starting with a lower dose. Fetal safety is not yet completely established as there is limited experience. A recent study showed that cabergoline achieved a high pregnancy rate with uneventful outcomes in infertile women with prolactinoma, independent of tumor size and bromocriptine resistance or intolerance. Cabergoline monotherapy could substitute for the conventional combination therapy of pregestational surgery or irradiation plus bromocriptine in macroprolactinomas.18
  4. Hydergine: It is a mixed ergot alkaloid effective only if serum prolactin levels are less than 100 ng/mL. It is well tolerated by patients and should be used in cases where bromocriptine fails.
183
 
Cabergoline vs Bromocriptine in ART
A recent study showed that the cost of treatment was significantly higher with cabergoline than with bromocriptine. However, side effect rate was significantly higher with bromocriptine than with cabergoline (15.3% vs 2.5%). Cabergoline and bromocriptine showed no differences in IVF outcomes and pregnancy results.19
 
Estrogen Replacement Therapy
Where tumor is small but is producing a significant hypoestrogenism estrogen replacement therapy should be given for protection of the bones and vascular system. When contraception is needed, they can be put on low dose contraceptive pills. There is no risk of tumor expansion due to estrogens since the level given is only enough to raise levels up to those in a natural cycle.20
 
Surgical Removal
 
Indications
  1. Patient unwilling for long-term drug therapy.
  2. Drug resistance.
  3. Intolerable side effects of drugs.
  4. Nonfunctioning tumors where prolactin levels are not very high. These tumors may expand with invasion into cavernous sinus, compression of optic chiasma and hemorrhage causing pituitary apoplexy.
  5. Suprasellar extension not regressing with drug therapy.
184
Surgical techniques have advanced to microsurgery using the trans-sphenoidal approach to expose the sella turcica. Normal yellow tissue of the pituitary gland can be distinguished from tumor tissue in small tumors. However, this tumor does not have a capsule and it may be difficult to distinguish from normal tissue in large tumors. Once it grows beyond the sella turcica it cannot be removed totally.
Preoperative medical therapy: This decreases the tumor size in 50 to 70% of cases making tumor more amenable for surgery. However, prolonged administration of drug may cause fibrosis and difficulty in dissecting.
Results: Serum prolactin comes to normal with resumption of normal menstrual cycles in 30% of cases with macroadenoma and 70% with microadenoma. The best results are obtained in patients with serum prolactin less than 500 ng/mL. Cure rate decreases if levels of prolactin are high. Results are better in intrasellar tumors.21 Pregnancy was achieved in 88% of those desiring conception following surgery.22
Recurrence: It may occur for 10% of cases with macroadenoma and 70% of those with microadenoma.23 Reason for recurrence being:
  1. Partial resection of tumor, since it may be difficult to differentiate from normal tissue.
  2. Multifocal origin of prolactinoma.
  3. Persistant abnormal stimulus to lactotrophs.
Follow-up in cases where symptoms persist includes monitoring of serum prolactin every 6 months and imaging of pituitary every year for 2 years. Imaging is thereafter done every year. Dopamine agonists are used for tumor recurrence or when ovulation induction is required.
Repeat surgery has a success rate of 33% only. Irradiation after surgery may cause stroke and other brain tumors.185
 
Complications
  1. Panhypopituitarism: It is seen in up to 10 to 30% of patients.
  2. Cerebrospinal fluid leak.
  3. Meningitis.
  4. Diabetes insipidus: It manifests in 10 to 40% of patients and lasts for about 6 months.
  5. Mortality: Mortality occurs in less than 1% of the total surgeries.
 
Radiation
It is not the primary choice of treatment and may be tried if medical management or surgery fail. It is given using linear cobalt or proton mode.
 
Disadvantage
  1. Results are less satisfactory than with surgery.
  2. Slow response with radiotherapy prolactin levels years to come to normal.
  3. Panhypopituitarism can occur even after many years and may need hormone replacement.
  4. Multiple endocrinopathies.24
  5. Damage to optic nerve.
  6. Diabetes insipidus.
Hyperprolactinemia is a frequent cause of anovulatory infertility and luteal phase defect. Dopaminergic treatment is the first line of treatment and is very effective in both idiopathic hyperprolactinemia and prolactinoma, with a 60 to 80% pregnancy rate.
 
References
  1. Ben-David M, Schenker JG. Transient hyperprolactinemia. A correctable cause of female idiopathic infertility. J Clin Endocrinol Metab. 1983;57:442–4.
  1. Cunha-Filho JS, Gross JL, Lemos NA, Brandelli A, Castillos M, Passos EP. Hyperprolactinemia and luteal insufficiency in infertile patients with mild and minimal endometriosis. Horm Metab Res. 2001;33(4):216–20.
  1. Padilla SL, Person GK, McDonough PG, Reindollar RH. The efficacy of bromocriptine patients with ovulatory dysfunction and normoprolactinemic galactorrhea. 1985;44:695–8.
  1. Strachan MW, Teoh WL, Don-Wauchope AC, Seth J, Stoddart M, Beckett GJ. Clinical and radiological features of patients with macroprolactinaemia. Clin Endocrinol. 2003;59(3):339–46.
  1. Sadideen H, Swaminathan R. Macroprolactin: what is it and what is its importance? Int J Clin Pract. 2006;60(4):457–61.
  1. Cattaneo F, Kappeler D, Müller B. Macroprolactinaemia, the major unknown in the differential diagnosis of hyperprolactinaemia. Swiss Med Wkly. 2001;131(9-10):122–6.
  1. Elster AD. Modern imaging of the pituitary. Radiology. 1993;187:1–14.
  1. Bayrak A, Saadat P, Mor E, Chong L, Paulson RJ, Sokol RZ. Pituitary imaging is indicated for the evaluation of hyperprolactinemia. Fertil Steril. 2005;84(1):181–5.
  1. Hall WA, Luciano MG, Doppman JL, Patronas NJ, Oldfield EH. Pituitary MRI in normal human volunteers: Occult adenomas in general populations. Ann Intern Med. 1994;120:817–20.
  1. Beckers A, Petrossians P, Abs R, Flandroy P, Stadnik T, de Longueville M, et al. Treatment of macroprolactinomas with long acting and repeatable form of bromocriptine: a report of 29 cases. J Clin Endocrinol Metab. 1992;75:275–80.
  1. Soule SG, Jacob HS. Prolactinoma: Present day management. Br J Obstet Gynecol. 1995;102:178–81.
  1. Cuellar FG. Bromocriptine mesylate (Parlodel) in the management of amenorrhea/galactorrhea associated with hyperprolactinemia. Obstet Gynecol. 1980;55:278–84.
  1. Porcile A, Gallardo E, Venegas E. Normoprolactinemic anovulation nonresponsive to clomiphene citrate: Ovulation induction with bromocriptine. Fertil Steril. 1990;53:50–5.
  1. Turkalj I, Braun P, Krupp P. Surveillance of bromocriptine in pregnancy. JAMA. 1982;247:1589–91.
  1. Webster J. Carbogoline and qunagolidine therapy for prolactinomas. Clin Endocrinol. 2000;53:549–50.
  1. Passos VQ, Souza JJ, Musolino NR, Bronstein MD. Long-term follow-up of prolactinomas: normoprolactinemia after bromocriptine withdrawal. Clin Endocrinol Metab. 2002;87:3578–82.
  1. Di Sarno A, Landi ML, Cappabianca P, Di Salle F, Rossi FW, Pivonello R, et al. Resistance to cabergoline as compared to bromocriptine in hyperprolactinemia: prevalence clinical definition and therapeutic strategy. J Clin Endocrinol Metab. 2001;86:5256–61.
  1. Ono M, Miki N, Amano K, Kawamata T, Seki T, Makino R, et al. Individualized high-dose cabergoline therapy for hyperprolactinemic infertility in women with micro- and macroprolactinomas. J Clin Endocrinol Metab. 2010;95(6):2672–9.
  1. Bahceci M, Sismanoglu A, Ulug U. Comparison of cabergoline and bromocriptine in patients with asymptomatic incidental hyperprolactinemia undergoing ICSI-ET. Gynecol Endocrinol. 2010;26(7):505–8.
  1. Correnblum B, Donovan L. The safety of physiological estrogen plus progestin replacement therapy and oral contraceptive therapy in women with pathological hyperprolactinemia. Fertil Steril. 1993;59:671.
  1. Losa M, Mortini P, Barzaghi R, Gioia L, Giovanelli M. Surgical treatment in prolactin secreting adenoma: Early results and long-term outcome. J Clinical Metab. 2002;87:3180–6.
  1. Feigenbaum SL, Downey DE, Wilson CB, Jaffe RB. Transsphenoidal pituitary resection for preoperative diagnosis of prolactin secreting adenoma in women: Long-term follow-up. J Clin Endocrinol Metab. 1996;81:1711–9.
  1. Schlechte JA, Sherman BM, Chapler FK, Van Gilder J. Long-term follow-up of women with surgically treated prolactin secreting tumors. J Clin Endocrinol Metab. 1986;62:1296–301.
  1. Hoybye C, Grenback E, Rahn T, Degerblad M, Thorén M, Hulting AL. Adrenocorticotropic hormone producing pituitary tumor: 12 to 22 year follow up after treatment with sterotactic radiosurgery. Neurosurg. 2001;49:284–91.

Role of Androgens in Ovulation Inductionchapter 12

Shashi Prateek,
Surveen Ghumman
Androgens are intermediates in estrogen biosynthesis and local regulators of ovarian function. The atherogenic nature of androgens has been questioned. It is believed that androgens may amplify the FSH effects on the ovary improving ovarian response and that granulosa cell stimulation by FSH is actually an androgen-modulated process. Androgens contribute to the paracrine regulation of follicular maturation and atresia. Ovarian stroma and granulosa cells of primordial follicles and follicles at more advanced stages of folliculogenesis have androgen receptors. During intermediate stages of follicular development, locally produced androgen acts via granulosa cell androgen receptors (AR) to promote follicle-stimulating hormone (FSH)-induced granulosa cell differentiation through amplifying cAMP-mediated post-receptor signaling.1 Granulosa cell-specific androgen receptors, promote preantral follicle growth and prevent follicle atresia, thus enhancing ovarian function.2
In a recent study, the physiological significance of androgens in female reproduction became clear when female mice with 189global knockout of androgen receptor (AR) expression were found to have reduced fertility with abnormal ovarian function.3 It was concluded that androgen receptors and androgens are essential for follicular recruitment, and normal follicle development and play a critical role in regulating ovarian function and fertility. This theory is supported by the fact that very low levels of testosterone on day 3 decrease success rate of IVF.4
Decreased ovarian reserve is becoming a common entity as more women are postponing childbearing. Many protocols are followed for these poor responders. Therapeutic benefits from supplementation with dehydroepiandrosterone (DHEA) and testosterone in women with diminished ovarian reserve (DOR) have been noted in the form of improved response to ovarian stimulation. Supplemental treatment with these drugs during ovarian stimulation may represent a novel way to maximize ovarian response.5
 
Impact of Androgens on Reproductive Outcome
 
Improvement in Number of Oocytes and Embryo
It was suggested that DHEA supplementation appears to augment ovarian stimulation with gonadotropins in poor responders, resulting in improved oocyte yields. Cumulative DHEA effects may have possible effects on follicle recruitment. In a study after DHEA, a significant decrease in cycle day-3 estradiol levels was found. There was an increased number of >17 mm follicles (3 versus 1.9) and MII oocytes (4 versus 2.1).6
In a recent study, it was observed that transdermal testosterone increased the number of recruited follicles in previously poor responders by over fivefold. The total amount of gonadotropin needed to achieve such improved response 190with testosterone was significantly lower. The antral follicle count increased during testosterone treatment, and the number of follicles available for recruitment and development at the time of starting FSH therapy was higher.7 Androgen treatment promoted FSH action in those recruitable follicles as suggested by increased serum levels of androstenedione and IGF-I (both considered as markers of ovarian responsiveness to gonadotropin stimulation), thus yielding higher numbers of oocytes.8 80% of patients underwent oocyte retrieval and embryo transfer in the testosterone supplemented IVF cycle.7
 
Improvements in Oocytes and Embryo Quality
It has been observed that with DHEA and testosterone, not only significant increase in oocytes and embryo numbers are seen, but also improved embryo quality is available.9
After DHEA supplementation, a significant increase in number of top quality day 2 (2.2 versus 1.3) and day 3 embryos (1.9 versus 0.7) were achieved. Cycle cancelation rates were reduced (5.3% versus 42.1%), and pregnancy rates per patients (47.4% vs 10.5%, P < 0.001) and per embryo transfer (44.4% vs 0.0%, P < 0.01) were improved.6 A randomized, prospective, controlled study showed higher live birthrate compared with controls (23.1% versus 4.0%).5 Pretreatment with transdermal testosterone gel (TTG) was also shown to improve number of mature oocytes, fertilized oocytes, good-quality embryos, embryo implantation rate and clinical pregnancy rate per cycle initiated.8
 
Premature Versus Physiologic Diminished Ovarian Reserve (DOR)
DHEA supplementation was effective in both age dependent DOR and premature ovarian aging (POA), though POA patients did mildly better.10 The beneficial effects of DHEA increased with length of DHEA supplementation.11191
Premature ovarian failure (POF)/primary ovarian insufficiency (POI) patients have conceived spontaneously whereas 50–75 mg of dehydroepiandrosterone supplementation for at least 4 months. It also considerably improved intrauterine insemination and IVF outcome and pregnancy rates in these women. FSH levels in these patients decrease with DHEAS administration. Positive effect has been reported with oocyte and embryo quality, with number of euploid embryos increasing and miscarriage rate decreasing.12
 
Effects on Embryoploidy, Miscarriage Risk and Live Birthrates
It is assumed that ovarian aging is related to aneuploidy and a higher incidence is seen in older women and those with DOR. Hence, miscarriage rate, which is dependent on aneuploidy, is much more in these women. Pregnancy loss rates in women with DOR were 57.1% in women <35 years old, 63.5% in women 35–40 years old, and 90.0% in women >40 years old. These rates of pregnancy loss were significantly higher compared to age-matched patients with normal ovarian reserve.13 Anti-Mullerian hormone (AMH) levels are a direct reflection of ovarian aging. Significantly improved live birthrates at AMH ≥1.06 ng/mL were seen. The chance of live birth per treatment cycle is only 5% if AMH was less than 1.05 ng/mL. Above that, chances are significantly improved. Thus, AMH 1.05 ng/mL represents a distinct point of separation between poorer and better live birth chances.14
In a recent study on carrying out preimplantation genetic screening (PGS), short term DHEAS supplementation of 4–12 weeks resulted in reduction in aneuploidy. Beneficial DHEA effects on DOR patients, at least partially, are the likely consequence of lower embryo aneuploidy.15 Increasing 192aneuploidy with advancing female age occurs and it is difficult to have adequate numbers of PGS to check for embryoploidy. Alternatively, miscarriage rates may be taken as a surrogate for aneuploidy risk. Since at least 60% of spontaneous pregnancy loss is attributable to chromosomal abnormalities, it can be hypothesized that significant reductions in aneuploidy after DHEA supplementation should be reflected in lower miscarriage rates. After DHEA supplementation the miscarriage rate was found to be 15.1%, being significantly lower at all ages but most pronounced above age of 35 years. Miscarriage rates after DHEA not only were lower than in an average IVF population but were also comparable to rates reported in normally fertile populations. This supports the assumption of a DHEA effect on embryoploidy. It was suggested that preconception DHEA supplementation in normal fertile populations above age 35 years may have a positive role.16
 
How Does DHEA Affect Ovarian Reserve?
Many mechanisms have been suggested explaining the beneficial effects of DHEAS and testosterone on ovarian reserve. DHEA may be able to decrease aneuploidy rate by reversing damage to oocytes. Second mechanism may be that oocytes in unrecruited stage do not age but changes start taking place once follicle starts maturing after recruitment. The ovarian environment then varies with age and may provide substandard conditions for the oocyte. As proposed by Hodges et al, the environment affects segregation processes during meiosis, giving rise to increased aneuploidy at older ages. He suggested major disturbances in chromosome alignments on the meiotic spindle of oocytes (congression failure), responsible for aneuploidy, result from the complex interplay of signals regulating folliculogenesis. These changes subtly alter the late stages of oocyte growth, 193increasing the risk of a nondisjunction error. These findings have important implications for human aneuploidy, since they suggest that it may be possible to develop prophylactic treatments for reducing the risk of age-related aneuploidy. This pharmacological maneuvering to reduce aneuploidy and miscarriage rate has been tried with DHEAS.17
DHEA levels peak in humans between ages 20 and 30 years, and then decline by approximately 2% per year, to reach nadirs of 10 to 20% around age 80 years.18
Age-related aneuploidy may actually be a reversible DHEA deficiency. Since follicles are still present in menopausal ovaries if ovarian environment can be reconstituted to that present at young age fertility could be preserved for much longer. DHEA may, therefore, represent a first compound in a new category of pharamacological agents with potential to rejuvenate ovarian environments. Following a similar concept, Bentov et al based on the known loss of mitochondrial functions with advancing age, recently suggested the use of mitochondrial nutrients, like coenzyme Q10 (CoQ10), after demonstrating that CoQ10 increases oocyte numbers in older mice.19 Androgens also positively affect mitochondrial function.20
DHEA was seen to increase IGF-1 and since growth hormone had been suggested to improve oocytes yields via IGF-1, it is hypothesized that DHEA may be able to achieve similar effects.21
 
Predicting the Effectiveness of DHEA
It was seen in a study that AMH concentrations significantly improved after DHEA supplementation longitudinally by 60% over time (P=0.002). Younger women (under age 38 years) demonstrated improved AMH concentrations more than older females.22194
AMH levels are predictable of treatment outcomes after DHEA utilization. Spontaneous pregnancies were, of course, conceived after shorter exposure to DHEA as they conceived spontaneously on DHEA during waiting period for IVF. So, it was concluded that shorter exposure sometimes may be enough to raise fecundity but may not suffice to positively affect ploidy and miscarriage rates.23
 
Treatment Protocols, Side Effects and Complications
Pretreatment with 12.5 mg transdermal testosterone gel (TTG) has been applied daily for 21 days in the cycle preceding COS with GnRH antagonist protocol for IVF in poor responders. It may be given as transdermal 20 μg/kg per day during the 5 days preceding gonadotropin treatment. It takes about three months for a given primordial follicle to reach the preovulatory stage. However, the time of exposure to testosterone in the above studies that showed a positive result was relatively reduced. It can be hypothesized that it affects late events involved in follicular maturation rather than earlier stages. At present, since it has not been determined whether androgens are rescuing follicles or simply increasing the number of recruited ones, further research is needed in this respect. Therefore, the daily dose, timing and duration of androgen supplementation may be critical to adequately stimulate folliculogenesis mainly considering a recent study in rhesus monkeys showing that chronic administration 195(for five days before and continuing throughout FSH and LH treatment) of high dose of androgens is antagonistic to gonadotropin-stimulated ovarian function in primates. It has been postulated that there is a threshold effect of androgens on follicular function such that antagonistic actions of androgens may be manifested at elevated concentrations.24
 
Dose
DHEAS is given as 25 mg of micronized tablet tid, for at least four weeks prior to IVF cycle.11 Short-term supplementation is up to 4 to 12 weeks of DHEA prior to IVF and PGS.11
 
Preparation and Route
Distinct advantages from micronized and orally delivered DHEA was demonstrated in a single-dose study comparing three dehydroepiandrosterone delivery methods (oral crystalline steroid, micronized steroid, and vaginal administration) to ascertain whether physiologic levels of circulating dehydroepiandrosterone can be obtained while increases in testosterone are minimized. Micronization of oral dehydroepiandrosterone diminishes bioconversion to testosterone. Vaginal dehydroepiandrosterone delivers equivalent dehydroepiandrosterone but substantially diminishes dehydroepiandrosterone bioconversion.25
 
Side Effects
Side effects at these dosages are small and rare, and primarily relate to androgen effects.
They include oily skin, acne vulgaris and hair loss. More frequently, patients comment on improved energy levels and better sex drive.196
A recent systemic review and meta-analysis in 2012 on the use of androgens or androgen-modulating agents in poor responders undergoing IVF concluded that transdermal testosterone pretreatment seems to increase clinical pregnancy and live birthrates in poor responders undergoing ovarian stimulation for IVF. However, there is insufficient data to support a beneficial role of rLH, hCG, DHEA or letrozole administration in the probability of pregnancy in poor responders undergoing ovarian stimulation for IVF.26
In conclusion, ovarian environments, but not resting oocytes, age as women grow older. Hence, pretreatment with transdermal testosterone may be a useful approach for women known to be low responders on the basis of a poor response to controlled ovarian stimulation but having normal basal FSH concentrations. DHEA supplementation apparently significantly reduces these age-related increases in aneuploidy, and, therefore, also reduces age-associated increases in miscarriages. A definite beneficial effect of androgens is seen in poor responders but needs further research to determine the best dose and period of administration.
 
References
  1. Hillier SG, Tetsuka M. Role of androgens in follicle maturation and atresia. Baillieres Clin Obstet Gynaecol. 1997;11(2):249–60.
  1. Ware VC. The role of androgens in follicular development in the ovary. I. A quantitative analysis oocytes ovulation. J Exp Zoology. 1982;222:155–67.
  1. Sen A, Hammes SR. Granulosa cell-specific androgen receptors are critical regulators of ovarian development and function. Mol Endocrinol. 2010;24(7):1393–403.
  1. Frattarelli JL, Peterson EH. Effect of androgen levels on in vitro fertilization cycles. Fertil Steril. 2004;81(6):1713–4.
  1. Casson PR, Lindsay MS, Pisarska MD, Carson SA, Buster JE. Dehydroepiandrosterone supplementation augments ovarian stimulation in poor responders: a case series. Hum Reprod. 2000;15(10):2129–32.
  1. Sönmezer M, Ozmen B, Cil AP, Ozkavukçu S, Taşçi T, Olmuş H, et al. Dehydroepiandrosterone supplementation improves ovarian response and cycle outcome in poor responders. Reprod Biomed Online. 2009;19(4):508–13.
  1. Balasch J, Fábregues F, Peáarrubia J, Carmona F, Casamitjana R, Creus M, et al. Pretreatment with transdermal testosterone may improve ovarian response to gonadotrophins in poor-responder IVF patients with normal basal concentrations of FSH. Hum Reprod. 2006;21(7):1884–93.
  1. Kim CH, Howles CM, Lee HA. The effect of transdermal testosterone gel pretreatment on controlled ovarian stimulation and IVF outcome in low responders. Fertil Steril. 2011;95(2):679–83.
  1. Barad D, Gleicher N. Effect of dehydroepiandrosterone on oocyte and embryo yields, embryo grade and cell number in IVF. Hum Reprod. 2006;21(11):2845–9.
  1. Wiser A, Gonen O, Ghetler Y, Shavit T, Berkovitz A, Shulman A. Addition of dehydroepiandrosterone (DHEA) for poor-responder patients before and during IVF treatment improves the pregnancy rate: a randomized prospective study. Hum Reprod. 2010;25(10):2496–500.
  1. Barad D, Brill H, Gleicher N. Update on the use of dehydroepiandrosterone supplementation among women with diminished ovarian function. J Assist Reprod Genet. 2007;24(12):629–34.
  1. Mamas L, Mamas E. Dehydroepiandrosterone supplementation in assisted reproduction: rationale and results. Curr Opin Obstet Gynecol. 2009;21(4):306–8.
  1. Levi AJ, Raynault MF, Bergh PA, Drews MR, Miller BT, Scott RT Jr. Reproductive outcome in patients with diminished ovarian reserve. Fertil Steril. 2001;76(4):666–9.
  1. Gleicher N, Weghofer A, Barad DH. Anti-Müllerian hormone (AMH) defines, independent of age, low versus good live-birth chances in women with severely diminished ovarian reserve. Fertil Steril. 2010;94(7):2824–7.
  1. Gleicher N, Weghofer A, Barad DH. Dehydroepiandrosterone (DHEA) reduces embryo aneuploidy: direct evidence from preimplantation genetic screening (PGS). Reprod Biol Endocrinol. 2010;8:140.
  1. Gleicher N, Ryan E, Weghofer A, Blanco-Mejia S, Barad DH. Miscarriage rates after dehydroepiandrosterone (DHEA) supplementation in women with diminished ovarian reserve: a case control study. Reprod Biol Endocrinol. 2009;7:108.
  1. Hodges CA, Ilagan A, Jennings D, Keri R, Nilson J, Hunt PA. Experimental evidence that changes in oocyte growth influence meiotic chromosome segregation. Hum Reprod. 2002;17(5):1171–80.
  1. Walker ML, Anderson DC, Herndon JG, Walker LC. Ovarian aging in squirrel monkeys (Saimiri sciureus). Reproduction. 2009;138(5):793–9.
  1. Bentov Y, Esfandiari N, Burstein E, Casper RF. The use of mitochondrial nutrients to improve the outcome of infertility treatment in older patients. Fertil Steril. 2010;93(1):272–5.
  1. Pitteloud N, Mootha VK, Dwyer AA, Hardin M, Lee H, Eriksson KF, et al. Relationship between testosterone levels, insulin sensitivity, and mitochondrial function in men. Diabetes Care. 2005;28(7):1636–42.
  1. Casson PR, Santoro N, Elkind-Hirsch K, Carson SA, Hornsby PJ, Abraham G, et al. Postmenopausal dehydroepiandrosterone administration increases free insulin-like growth factor-I and decreases high-density lipoprotein: a six-month trial. Fertil Steril. 1998;70(1):107–10.
  1. Gleicher N, Weghofer A, Barad DH. Improvement in diminished ovarian reserve after dehydroepiandrosterone supplementation. Reprod Biomed Online. 2010;21(3):360–5.
  1. Gleicher N, Barad DH. Dehydroepiandrosterone (DHEA) supplementation in diminished ovarian reserve (DOR). Reprod Biol Endocrinol. 2011;9:67.
  1. Zeleznik AJ, Littler-Ihrig L, Ramasawamy S. Administration of dihydrotestosterone to rhesus monkeys inhibits gonadotropin-stimulated ovarian steroidogenesis. J Clin Endocrinol Metab. 2004;89:860–6.
  1. Casson PR, Straughn AB, Umstot ES, Abraham GE, Carson SA, Buster JE. Delivery of dehydroepiandrosterone to premenopausal women: effects of micronization and nonoral administration. Am J Obstet Gynecol. 1996;174(2):649–53.
  1. Bosdou JK, Venetis CA, Kolibianakis EM, Toulis KA, Goulis DG, Zepiridis L, et al. The use of androgens or androgen-modulating agents in poor responders undergoing in vitro fertilization: a systematic review and meta-analysis. Hum Reprod Update. 2012;18(2):127–45.

Ovarian Reserve and Fertility in Older Womenchapter 13

Neerja Goel,
Surveen Ghumman
In the modern era, women are increasingly delaying childbirth till their thirties and forties when fertility declines and risk of congenital anomalies and miscarriage increases. This is the section of population that is becoming a challenge for the infertility expert. A woman's advancing age is directly related to poor ovarian response to simulation. This situation arises from the natural process of aging and the depletion of the primordial follicular pool and consequent failure in recruitment of follicles. When superovulated these women produce few eggs and are known as poor responders. This may be because of interference with FSH action due to proteins. An autoimmune basis where there could be presence of antibodies against granulosa cells, or defective angiogenesis, autocrine or paracrine alterations leading to decreased quantities of certain intraovarian peptides could be the cause. There could be a genetic basis with FSH-receptor polymorphism. Low diffusion of exogenous gonadotropins has also been proposed. Evaluation of the ovarian reserve forms an integral part of workup of these women.201
 
Age and Fertility
Effect of age is seen on all the reproductive organs affecting fertility.
 
Ovary
  1. Decreasing population of follicles in the ovary that are responsive to stimulation are seen. During embryonic development, the number of oogonia peak by 20 weeks of gestation to approximately 7 million oocytes. At birth this number declines to 1–2 million germ cells and by 20 years only 250,000 to 300,000 remain, decreasing to about 25,000 by age of 30–35 years, and 6,000 by age of 40 years.1
  2. Oocytes show high incidence of chromosomal abnormalities because of advancing maternal age. During IVF the fertilization rate declines with advancing age and an increasing proportion of the residual unfertilized eggs showed abnormal chromosomes. It is this decline in the number and quality of oocytes that hampers fertility.
 
Uterus
Uterine senescence may be partially responsible for the decrease in fertility associated with increasing age. It is clear that uterine factor plays only a small part in this decline. With surrogacy it has been proven that even an older uterus is capable of reproductive functions when stimulated. There are various theories put forward for poor reproductive performance of the uterus with age.
  1. Number of pinopodes in the uterine endothelium may be drastically decreased, thus hindering implantation.
  2. Uterus may become fibrous with poor blood supply and has decreased ability to synthesize prostaglandins.202
  3. Reactive oxidative species produced in the reproductive tract of older women is thought to decrease implantation.
  4. There is an increase in the incidence of fibroids and endometriosis in older women.
 
Tube
  1. Deciliation of the Fallopian tube endothelium occurs leading to decreased efficiency of tubal transport.
  2. Increased incidence of pelvic inflammatory disease in older women may lead to damaged tubes.
 
Management of Older Women with a Fertility Problem
It is advisable to initiate investigations at an early stage. It is important to exclude factors such as fibroids, incipient ovarian failure, and endometrial polyps, which are less likely to affect younger women. Fibroids of significant size or location should generally be removed prior to any form of assisted conception. Smoking appears to expedite the ovarian aging process.
More recent molecular research has indicated a genetic etiology for the age diminished ovarian reserve. There is evidence to show a decline in oocyte quality with increasing maternal age. With DNA fragmentation, the rates of chromosomal abnormalities were found to be significantly higher in older women when their unfertilized oocytes were analyzed following IVF. A significantly higher amount of disassociated chromatids was found in older women. For women less than 34, the rate of genetic aberrations was 24%, between 35 and 39 years the rate was 52%, and women 40 years and above had a rate of 95.8%.2203
 
Assessment of the Ovarian Reserve
With intensified research on reproductive aging, methods to measure the number, quality and reproductive potential of the remaining ovarian follicle pool have evolved and been given the name of ‘Ovarian Reserve Tests’. These tests are indicated in those groups of patients at high risk of ovarian failure (Table 13.1).
No single test for assessment of ovarian reserve is adequate, so a combination of following tests is recommended (Table 13.2).
 
Basal Follicle Stimulating Hormone Levels
Raised early follicular FSH level is one of the earliest indications of reproductive aging (Table 13.3). It has been shown that women with elevated FSH levels have a higher minimum threshold level of FSH required to initiate sustained follicular development.3
Table 13.1   Indications for testing ovarian reserve
1. Age more than 35 years
2. Unexplained infertility regardless of age
3. Family history of early menopause
4. Previous ovarian surgery
  • Cystectomy
  • Ovarian drilling
  • Unilateral oophorectomy
  • Chemotherapy
  • Radiation
5. Smoking
6. Demonstrated poor response to exogenous gonadotropin stimulation
204
Table 13.2   Ovarian reserve tests
1. Clinical variables--age, history of canceled cycles
2. Blood tests
  • Basal FSH
  • Day 2 or 3 serum estradiol
  • Basal inhibin B
  • Mullarian inhibiting substance
3. Dynamic tests
  • Clomiphene citrate challenge test
  • Dynamic assay of estradiol and inhibin B after GnRHa stimulation test (GAST)
  • Exogenous FSH ovarian reserve test (EFORT)
4. Ultrasound
  • Antral follicle count
  • Ovarian volume
  • Ovarian stromal peak systolic velocity, including waveform and pulsatility index
Table 13.3   Basal follicle stimulating hormone level
FSH levels IU/L
< 9
Reassure
9–10
Suboptimal
10–12
Decreased ovarian reserve
12–17
Markedly reduced ovarian reserve
17–20
Poor prognosis
> 20
No pregnancy
205
 
Early Follicular Phase Estradiol Levels
Elevated estradiol level in early follicular phase is an indicator of diminished ovarian reserve and has been associated with poor results. Early elevation of serum estradiol level reflects advanced follicular development and early selection of dominant follicle seen in older women due to rising FSH levels. A premature elevation of estradiol may suppress FSH causing masking of an elevated day 3 FSH. Hence, it is better if both FSH and estradiol are measured, to eliminate false negative results. Basal E2 level of more than 80 pg/mL is associated with poor follicular response and development.
 
Early Follicular Phase Inhibin B Levels
Inhibin A is secreted predominantly in the luteal phase and inhibin B in the follicular phase by granulosa cells. Inhibin B may, therefore, be a direct marker of ovarian reserve.4
Inhibin B levels decline with increasing age due to decreased number of follicles and decreased secretion by the granulosa cells. Since inhibin is decreased, there is poor negative feedback and FSH level is increased. Low levels of inhibin B on day 3 are associated with a poor outcome of IVF. Its role is limited because of less reliability and difficult serum estimation. A value of more than 45 pg/mL is taken as normal.
 
Anti-Müllerian Hormone
Like inhibin levels of anti-Müllerian hormone reflect the health of the granulosa cell. The anti-Müllerian hormone and antral follicle count correlates well with age even in women under 40 years whereas FSH and inhibin B predominantly changes in women more than 40 years of age.5
It has been seen that <1 pmol/L shows a extremely poor response and cycle should be canceled. 1-5 pmol/L are still at risk of poor response and short stimulation protocols should 206be used. Response is found to be adequate when values are between 5–15 pmol/L. Between 15 to 25 pmol/L there is risk of hyperstimulation and caution should be used. More than 25 pmol/L there is high risk of hyperstimulation6 (Table 13.4).
Using a cut-off value of 8.1 pmol/L, plasma AMH assessment could predict poor ovarian reserve on a subsequent IVF cycle with a sensitivity of 80% and a specificity of 85%. Plasma AMH assessments are superior to FSH in identifying women with reduced ovarian reserve. Anti-müllerian hormone assessment should be considered as a useful adjunct to FSH/oestradiol levels and antral follicle count when estimating ovarian reserve.7
 
Ovarian Sensitivity Index
Ovarian sensitivity index (OSI) is calculated dividing the total administered FSH dose by the number of retrieved oocytes. AMH and OSI show a highly significant negative correlation that is stronger than the one between AMH and the total number of retrieved oocytes and between AMH and the total FSH dose. OSI reflects quite satisfactory the AMH level and may be proposed as a surrogate of AMH assay in predicting ovarian responsiveness to FSH in IVF.
Table 13.4   AMH values predicting ovarian reserve
Value of AMH (pmol/L)
Ovarian response
<1
Extremely poor response and cycle should be canceled
1–5
At risk of poor response and short stimulation protocols should be used
5–15
Response adequate
15–25
Chances of hyperstimulation. Caution to be applied
>25
High chances of severe OHSS
207
Being very easy to calculate and costless, its use could be proposed where AMH measurement is not available or in developing countries where limiting costs is of primary importance.8
 
Ovarian Volume and Antral Follicle Count
There is growing evidence that both ovarian volume and number of antral follicles within the ovaries are useful indicators of ovarian reserve and response to superovulation (Table 13.5).9 Decreased ovarian volume is a sign of aging and may be observed earlier than rise in FSH level. Small ovaries are associated with poor response to superovulation.
Antral follicle count may be helpful in determining stimulation protocol, as it is the most reliable determinant of oocytes retrieved per starting FSH dose. Antral follicle count predicts ovarian response, but not embryo quality, implantation or pregnancy rate.10
 
Basal Ovarian Stromal Blood Flow
Undetectable basal ovarian stromal blood flow in at least one ovary is related to low ovarian reserve in infertile women and is linked to the pathophysiology of ovarian aging.11
On 3D ultrsonography, the vascularization index, flow index, and vascularization flow index were significantly lower in ovarian stroma (P<.05) in the poor responder compared with the women with a normal response.12
Table 13.5   Antral follicle count
< 4
Poor
4–7
Low count—high dose of FSH needs to be given
8–12
Slightly reduced
> 12
Normal
208
 
Clomiphene Challenge Test (CCT)
It is a provocative and sensitive test of ovarian reserve that takes into account the endocrine dynamics of the cycle after stimulation with clomiphene. In normal women, there is a transient increase of FSH and LH with clomiphene. LH increases more than FSH. In women with a decreased ovarian reserve, FSH increases much more because a smaller follicle cohort in aging women produce less inhibin resulting in decreased negative feedback. Baseline FSH and LH levels are evaluated and then the patient is given clomiphene citrate from day 5 to day 9 in a dose of 100 mg/day. The FSH and LH levels are evaluated again on day 10. The value of FSH should not be more than 26 IU. Elevated levels are strongly suggestive of a decreased reserve and should be taken into account during controlled ovarian hyperstimulation protocols.13 It has been seen that women who have a normal day 3 FSH and an elevated day 10 level have the same prognosis as one with an elevated day 3 FSH level. An abnormal clomiphene challenge test is seen in 10% of all infertile women, being higher in older women and those with unexplained infertility.14 Positive predictive value is 90% for both day 3 FSH and clomiphene challenge test. Both tests have a poor sensitivity of 7 to 26% and good specificity of 98%.15 Basal FSH and the CCT are similar in predicting the ability to achieve a clinical pregnancy in women undergoing infertility treatment. With either test, a normal result is not useful, but an abnormal result virtually confirms that pregnancy will not occur with treatment. When compared with basal FSH, CCT has hardly any additional value.16,17
 
GnRHa Stimulation Test (GAST)
This provides a more reliable assessment than mere basal assays. GnRHa produces an initial rise in LH, FSH, inhibin 209B and estradiol. This initial increase in estradiol is a better predictor than basal FSH, and FSH:LH ratio.18 The test evaluates the change in serum E2 levels between cycle day 2 and 3 after administering 1 mg of subcutaneous leuprolide acetate. Four different patterns of E2 levels were noted. Patients with E2 elevations by day 2 and decline by day 3 had better implantation and pregnancy rates than those with either no rise in E2, or persistently elevated E2 levels. The GAST has a rather good ability to predict poor response in IVF. However, comparing the predictive accuracy and clinical value of the GAST with a day 3 antral follicle count (AFC) and inhibin B, it appeared that neither a single nor a repeated GAST performed better. In addition, the predictive ability toward ongoing pregnancy is poor. Therefore, the use of the GAST as a predictor of outcome in IVF should not be advocated.19,20
 
Exogenous FSH Ovarian Reserve Test (EFORT)
This is a dynamic test. Originally, the test was developed to improve the predictive value of day 3 FSH values in controled ovarian hyperstimulation for IVF. The E2 level is recorded on cycle day 3 before the administration of 300 IU of purified FSH. Another level of E2 is done 24 hours after giving FSH. It was postulated that the dynamic increase in E2 of more than 30 pg/mL would be predictive of a good response in a subsequent IVF cycle.
 
Advantages of Ovarian Reserve Testing
  1. In high-risk populations, it affects treatment decisions.
  2. Frankly abnormal tests may persuade women to look for other alternatives.
  3. Borderline values may persuade them to take advantage of a rapidly decreasing opportunity.210
  4. Abnormal ovarian reserve test may signify an aging ovum even in women of younger age making them a population at increased risk of fetal aneuploidy, justifying prenatal diagnostic studies.
In the recent years, these tests are occupying an increasingly important place in the workup of infertile women. Although they are reliable, rigid interpretation is discouraged as each treatment cycle may be different from the next.
 
Treatment of Patients with Reduced Ovarian Reserve
A number of options are there for these women. They are often termed as poor responder as they do not respond to the regular stimulation protocols (See chapter 14).
  1. Superovulation with IUI: It may be tried if patients do not demonstrate marked decrease in ovarian reserve.
  2. Pretreatment with oral contraceptives followed by gonadotropins: In patients who are estrogen deficient with a high gonadotropin level, ovulation can be accomplished by first suppressing pituitary gonadotropins with estrogen and then giving hMG for ovarian stimulation. The idea of giving ethynylestradiol 0.05 mg/day was to reduce the level of FSH and LH thus withdrawing the downregulatory effect of elevated gonadotropins on LH and FSH receptors and restoring sensitivity of the remaining follicles. When FSH and LH reach normal values gonadotropins stimulation is started with 150 IU of FSH. If E2 after 5 days is less than 35 pg/mL the dosage of FSH is increased up to 375 IU. If E2 does not rise beyond 50 pg/mL despite maximum dose of 5 ampules the cycle should be canceled.
  3. High dose of gonadotropins: Since these patients are poor responders, they are started with a gonadotropin dose of 211150 IU/day and may be increased to 450 IU/day for an effect.21
  4. Short GnRH agonist protocol: It is useful as its flare effect gives high levels of FSH that these patients require.22
  5. Oocyte donation: It is an important option in cases of infertility with advanced age. In 1984, the first live birth with oocyte donation and IVF was reported. Synchronization of recipient and donor is to be achieved in oocyte donation protocol. In women who are menstruating daily oral estradiol of 4 to 8 mg is given till day of transfer when it is increased to 6 mg/day. Length of estrogenic exposure can vary from 6 to 38 days as seen in various studies. Usually a 14 day period is sufficient. Patients are down-regulated with GnRH agonists prior to estradiol administration. Optimal time of embryo transfer is 2 to 4 days after progesterone administration that is given as 400 to 600 mg intravaginally or 100 mg intramuscularly. After pregnancy test is positive estradiol is increased to 8 mg/day. Replacement therapy is carried out till 60 days. If the woman is amenorrheic then 3 to 6 months of estrogen and progesterone are given till minimum 3 months of bleeding, an uterocervical length of 5 to 6 cm and an endometrial thickness of 8 to 9 mm is achieved.
  6. Cryopreservation of oocyte or embryo: It is an important option for women at risk of decreasing ovarian reserve. Embryo cryopreservation gives better results than oocyte preservation with a pregnancy rate of 20 to 30% per transfer of 2 to 3 embryos. Because of fragility of the meiotic spindle and formation of ice crystals, the success of oocyte preservation is limited, but is steadily improving.
  7. Ovarian transplant: The ovarian transplant can be orthotopic (at ovarian site) or heterotopic (another site). The main obstacle to the success of this technique is 212poor oocyte viability. It is a treatment that will become accessible in the years to come.
Fertility in older women is a challenge being faced with increasing frequency ovarian reserve testing can only help in guiding the therapist on the lines of treatment and its success rate. A combination of test should be done to improve accuracy. Cryopreservation of embryo or oocyte is becoming an important option for women at risk of decreased ovarian reserve as those who undergo treatment for cancer.
 
References
  1. Faddy MJ, Gosden RG, Gougeon A, Richardson SJ, Nelson JF. Accelerated disappearance of ovarian follicle in midlife: implications for forecasting menopause. Hum Reprod. 1992;7:1342–6.
  1. Demario MA, Davis OK, Rosenwaks Z. The role of maternal age in assisted reproductive technologies. Reprod Med Review. 1999;7:141–60.
  1. Seifer DB, Scott RT, Bergh PA, Abrogast LK, Friedman CI, Mack CK, et al. Women with declining ovarian reserve may demonstrate a decrease in day 3 serum inhibin B before a rise in day 3 FSH. Fertil Steril. 1999:72:63-5.
  1. Balasch J, Creus M, Fabregues, et al. Inhibin, FSH, and age as predictors of ovarian response in IVF cycles stimulated with GnRHa treatment. Am J Obstet Gynaecol. 1996;175:1226–30.
  1. van Rooij IA, Broekmans FJ, Scheffer GJ, Looman CW, Habbema JD, de Jong FH, et al. Serum anti-Mullerian hormone levels best reflect the reproductive decline with age in normal women with proven fertility: A longitudinal study. Fertil Steril. 2005;83(4):979–87.
  1. Nelson SM, Yates RW, Flemming R. Serum anti-Müllerian hormone and FSH: prediction of live birth and extremes of response in stimulated cycles—implications for individualization of therapy. Hum Reprod. 2007;22(9):2414–21.
  1. Tremellen KP, Kolo M, Gilmore A, Lekamge DM. ANZJOG. 2005;45:20–4.
  1. Biasoni V, Patriarca A, Dalmasso P, Bertagna A, Manieri C, Benedetto C, et al. Ovarian sensitivity index is strongly related to circulating AMH and may be used to predict ovarian response to exogenous gonadotropins in IVF. Reprod Biol Endocrinol. 2011;9:112.
  1. Lass A, Brinsden P. The role of ovarian volume in reproductive medicine. Hum Reprod. 1999;5:256–66.
  1. Hsu A, Arny M, Knee AB, Bell C, Cook E, Novak AL, et al. Antral follicle count in clinical practice: analyzing clinical relevance. Fertil Steril. 2011;95(2):474–9.
  1. Younis JS, Haddad S, Matilsky M, Radin O, Ben-Ami M. Undetectable basal ovarian stromal blood flow in infertile women is related to low ovarian reserve. Gynecol Endocrinol. 2007;23(5):284–9.
  1. Pan HA, Wu MH, Cheng YC, Wu LH, Chang FM. Quantification of ovarian stromal Doppler signals in poor responders undergoing in vitro fertilization with three-dimensional power Doppler ultrasonography. Am J Obstet Gynecol. 2004;190(2):338–44.
  1. Vanushpolsky EH, Hurwitz S, Tikh E, Racowsky C. Predicting usefulness of cycle day 10 follicle stimulating hormone level in clomiphene citrate challenge test for in vitro fertilization outcome in women younger than 40 years of age. Fertil Steril. 2003;80:111.
  1. Scott RT, Leonardi MR, Hofmann GE, Illions EH, Neal GS, Navot D. A prospective evaluation of clomiphene citrate challenge test screening of the general infertility population. Obstet Gynecol. 1993;82:539–44.
  1. Jain T, Soules MR, Collins JA. Comparison of basal follicle stimulating hormone versus clomiphene citrate challenge test for ovarian reserve screening. Fertil Steril. 2004;82:180–6.
  1. Hendriks DJ, Mol BW, Bancsi LF, te Velde ER, Broekmans FJ. The clomiphene citrate challenge test for the prediction of poor ovarian response and nonpregnancy in patients undergoing in vitro fertilization: a systematic review. Fertil Steril. 2006;86(4):807–18.
  1. Jain T, Soules MR, Collins JA. Comparison of basal follicle-stimulating hormone versus the clomiphene citrate challenge test for ovarian reserve screening. Fertil Steril. 2004;82(1):180–5.
  1. Amir R, Stuart L, Sappho M, Mandy D, Raul M, Geoff T, et al. Dynamic assays of inhibin B and oestradiol following buserelin acetate administration as predictors of ovarian response in IVF. Hum Reprod. 2000;15:2297–301.
  1. Hendriks DJ, Broekmans FJ, Bancsi LF, Looman CW, de Jong FH, te Velde ER. Single and repeated GnRH agonist stimulation tests compared with basal markers of ovarian reserve in the prediction of outcome in IVF. J Assist Reprod Genet. 2005 Feb;22(2):65–73.
  1. Maheshwari A, Gibreel A, Bhattacharya S, Johnson NP. Dynamic tests of ovarian reserve: a systematic review of diagnostic accuracy. Reprod Biomed Online. 2009 May;18(5):717–34.
  1. Surrey ES, Schoolcraft WB. Evaluating strategies for improving ovarian response of the poor responder undergoing assisted reproductive techniques. Fertil Steril. 2000;73:667–76.
  1. Sharara FI, McClormick HD. Use of microdose GnRH-agonist protocol in women with low ovarian volumes undergoing IVF. Hum Reprod. 2001;16:500–3.

Ovarian Stimulation for the Poor Responderchapter 14

Surveen Ghumman
About 10 to 15% of women have a poor ovarian response during ovarian stimulation. This could be because of advanced ovarian age, decreased ovarian function due to chemotherapy, radiotherapy or surgery or a premature ovarian failure (Table 14.1).
Table 14.1   Causes of poor ovarian response
1. Diminished ovarian reserve
2. Decreased number of FSH receptors in granulosa cells
3. Inappropriate local vascular network for the distribution of gonadotropins
4. Presence of autoantibodies against granulosa cells
5. Defective signal transduction after FSH-receptor binding
6. Presence of a special FSH receptor-binding inhibitor in the follicular fluid
7. Lowered circulating gonadotropin surge-attenuating factor (GnSAF) bioactivity
216
 
 
How Do we Define a Poor Responder?
No exact definition has been selected so far. Low ovarian response is confirmed only after the patient has failed ovarian stimulation following an accepted ‘standard’ ovarian stimulation regimen.
Different authors have applied different criterion. A poor responder could be one with any of the following1:
  1. No. of follicles after a standard-dose ovarian stimulation protocol: Varies with different authors < 3 to <5 on the day of hCG administration.
  2. No. of oocytes after a standard-dose ovarian stimulation protocol: Varies with different authors < 3 to <5.
  3. Peak estradiol level at time of hCG also correlated with follicular development: < 500 pg/ml.
  4. Excessive requirements of gonadotropins: > 450 IU1.
  5. Prolonged stimulation.
  6. Basal FSH more than 10.
 
Prediction
There are a number of tests that predict a poor ovarian reserve (See chapter on poor ovarian reserve).
  1. Basal day 3 FSH.
  2. CC challenge test.
  3. Inhibin B.
  4. AMH.
  5. Ovarian volume.
  6. Antral follicle count.
 
Treatment
A number of therapeutic alternatives have been tried mostly alteration of stimulation protocol and addition of adjuvants. 217Complementary alternative therapy and options like oocyte donation have also been considered (Table 14.2).
Table 14.2   Protocols for poor responders
1. Gonadotropins
  • CC and hMG
  • High dose FSH/hMG
  • Use of recombinant vs purified urinary FSH
  • Luteal initiation of FSH
  • Low dose minimal stimulation protocols
2. GnRH agonist protocol
  • Flare GnRH agonist protocols – short and ultrashort
  • Minidose GnRH agonists
3. Antagonist
  • Only antagonist
  • Antagonist with agonist
4. Adjuncts
  • Use of LH
  • Growth hormone or GH-releasing factor or pyridostigmine
  • Oral contraceptives
  • Adjunctive use of nitric oxide (NO)-donors (L-arginine)
  • Glucocorticoids
  • Testosterone and DHEAS
  • Letrozole
  • Luteal estradiol
  • Aspirin
5. Natural cycle
6. CAM therapy (Complementary and alternative medicine)
7. Donor oocytes
218
 
Stimulation Protocols
 
Long Protocol
In a study on comparison of long agonist protocol with antagonist protocol, there were no significant differences in the cycle cancelation rates, duration of stimulation, consumption of gonadotropins, and mean numbers of mature follicles, oocytes and embryos obtained in poor responders. The implantation rates were similar, but the number of embryos transferred and pregnancy rate were significantly higher for the antagonist group.2
 
High Doses of Gonadotropins
High doses of gonadotropins up to 600 IU/day have been used but clear advantage with these high doses has not been seen in any study. A study conducted to assess optimal dose of gonadotropin after microdose gonadotropin-releasing hormone analog (GnRHa) flare cycles in poor responders showed a clinical pregnancy rate of 13.1%, 15.3%, and 16.1% for 300, 450 and 600 IU, respectively of FSH. There were no significant differences in the age, peak serum E2 concentration, days of stimulation with rFSH, total number of M2 oocytes retrieved, number of embryos transferred, clinical pregnancy rates, and cancelation rates of stimulation and embryo transfer between the three groups except for total rFSH dosage. The study concluded that there is no need to use doses above 300 IU of rFSH to increase the pregnancy rate in microdose cycles.3
On comparing 375 IU/day gonadotropin to the 450 IU/day in poor responders, the additional 75 IU/day does not give any improvement in either embryology or pregnancy outcomes.4219
 
Use of Recombinant FSH Versus Purified Urinary FSH
Small studies indicate better results with recombinant rather than purified FSH with a pregnancy rate of 33% versus 6%. However, larger randomized trials are needed to give definitive proof.5
 
Use of Recombinant FSH Versus Human Menopausal Gonadotropin
A recent study showed that recombinant follicle-stimulating hormone (rFSH) has no advantage over urinary human menopausal gonadotropin (hMG) on ovarian performance or the outcome of IVF–ET in poor responders’ IVF cycles. There were no statistical differences in numbers of follicles, oocytes recovered, cycle characteristics, or pregnancy rates between hMG and rFSH-stimulated cycles.6
 
Luteal Initiation of FSH
The initiation of FSH in luteal phase may increase the number of recruited follicles by opening the recruitment window earlier, in the preceding cycle. However, no study showed an advantage.
 
Stop Agonist Regime
These regimens are characterized by the use of relatively low doses of GnRH agonists commencing in the mid-luteal phase of the cycle and usually ending at the time of menses or shortly thereafter, in combination with high doses of gonadotropins. A clinical pregnancy rate per transfer of 32% was achieved.7
 
Microdose Protocol
Microdose of GnRH agonist are given with the idea of suppressing raised LH but not downregulating severely. 220GnRH agonist is given from the midluteal phase of the previous cycle to the onset of menstruation in the next cycle. Then high doses of gonadotropins (hMG/FSH) were given. The low dose and half duration of GnRHa therapy lessened the suppression of the response of the ovaries to COH compared with the regular long protocol of GnRHa downregulation therapy leading to better results.
 
Flare Protocol
GnRH agonists are given on the first day of period and stopped after 3 days.
Advantages:
  1. Ovarian suppression is not excessive as GnRH agonist is stopped before down regulation; hence, a better response to gonadotropin stimulation could be achieved.
  2. The initial stimulation of the GnRH receptors releases the FSH and this enhances the effect of exogenous FSH that is started after 3 days.
Standard Flare Protocol
Buserelin 0.5 mg is given for the first 3 days. A low cancelation rate (11.3%) and a good pregnancy rate (29%) was found.8 On comparing the flare-up versus the luteal GnRH agonist regimen, higher pregnancy rates (20.4% versus 11.7%) were observed with flare protocol.9 Results of flare protocol were similar to antagonist protocol.10
Microdose Flare Protocol
Several microdoses have been tried in an attempt to identify a dose, which causes endogenous FSH release with no increase in LH, progesterone and androgen secretion seen with classical flare protocol. Leuprolide 40 μg was given for 3 days followed 221by FSH. Smaller doses up to 20 μg has been tried. These studies show a trend toward improved results but larger studies are needed to support this issue.11 Microdose leuprolide acetate (LA) (50 μg twice daily) starting on the second day of withdrawal bleeding is also given. Stimulation protocol with decapeptyl involves giving oral contraceptives (OC) from day 21. On third day of OC cessation 0.005 mg decapeptyl is started twice daily. FSH with hMG on 2nd day of decapeptyl 450 to 750 IU/day is given. Microdose GnRHa flare-up protocol and multiple dose GnRH antagonist protocol seem to have similar efficacy in improving treatment outcomes of poor responder patients.12
 
GnRH Antagonist
In a study stimulation started with clomiphene citrate (100 mg/daily, from cycle days 2 to 5) combined with the appropriate dose of gonadotropins (mean 375 IU/day). The GnRH antagonist cetrorelix was started on cycle day 6 at a dose of 0.25 mg/day. There were increased number of retrieved oocytes per cycle and increased pregnancy rates per transfer (23.5% versus 10%).13
 
GnRH Antagonist Along with Agonists
Protocol:
  • Decapeptyl on first day of period for 3 days in a dose of 0.1 mg. Start maximal FSH dose from day 3 with flexible antagonist protocol.
  • Decapeptyl 0.1 mg from day 21 to period and then stop it. Start FSH maximal dose on day 2 along with antagonist when lead follicle more than 13 mm.
The available limited data, derived from small or preliminary studies, do not show any advantage from the use of GnRH antagonists.222
 
Mild Stimulation Protocols
The women with mild ovarian stimulation had lower duration of stimulation, total doses of gonadotropins used, serum E2 level on hCG day, the number of retrieved oocytes, and the number of mature oocytes than conventional stimulation. However, there is no significant difference in the number of good embryos, the number of transferred embryos, the cancelation rate, or the clinical pregnancy rate. In older women over 37 years old, the clinical pregnancy rate and live birthrate were higher when compared with the conventional protocol. In poor responder groups, mild ovarian stimulation is more cost-effective and patient friendly than conventional IVF and should be considered for poor responders over 37 years old.1,14
 
Modified Natural Cycle
Modified natural cycle (MNC) treatment consists of giving human menopausal gonadotropin (hMG) 150 IU/day IM if serum estradiol was ≤ 50 pg/mL on day 2 or 3 of the menstrual cycle.15 GnRH antagonists prevent LH surge and therefore, improve the results of natural IVF cycles. The GnRH antagonist should be started at day 8, at the daily dose of 0.25 mg. The natural cycle is a successful option in treatment of poor responders and implantation failure.16
 
Adjuvants
 
Androgens in Poor Responder
It is believed that androgens may amplify the FSH effects on the ovary improving ovarian response and that granulosa cell stimulation by FSH is actually an androgen-modulated process.223
Testosterone: Pretreatment with 12.5 mg transdermal testosterone gel (TTG) has been applied daily for 21 days in the cycle preceding COS with GnRH antagonist protocol for IVF in poor responders. It may be given as transdermal 20 μg/kg per day during the 5 days preceding gonadotropin treatment. It was associated with an increase in clinical pregnancy and live birthrates in poor responders undergoing ovarian stimulation for IVF.17
DHEA: It was suggested that DHEA supplementation appears to augment ovarian stimulation with gonadotropins in poor responders, resulting in improved oocyte yields. Cumulative DHEA effects may have possible effects on follicle recruitment. In a study with DHEA, a significant decrease in cycle day 3 estradiol levels was found. There was an increased number of >17 mm follicles (3 versus 1.9) and MII oocytes (4 versus 2.1).18 DHEAS is given as 25 mg of micronized tablet tid, for at least four to twelve weeks prior to IVF cycle.
A recent systemic review concluded that transdermal testosterone pretreatment seems to increase clinical pregnancy and live birthrates in poor responders undergoing ovarian stimulation for IVF. However, there is insufficient data to support a beneficial role of DHEA in the probability of pregnancy in poor responders undergoing ovarian stimulation for IVF.19
 
Growth Hormone (GH) and GH-releasing Factor or Pyridostigmine
GH stimulates ovarian steroidogenesis, leading to follicular development and enhances the ovarian response to FSH mediated via the IGF-1.20 Dose is 4 to 12 IU. GH is administered SC, starting on the day of ovarian stimulation with gonadotropins.
Pyridostigmine is an acetylcholinesterase inhibitor leading to increase in acetylcholine, which increases GH secretion. 224This approach was evaluated in a randomized study in that poor responders were given 120 mg/day pyridostigmine orally, from the day of downregulation until the day of hCG, along with a long luteal GnRH agonist regimen (triptorelin 0.1 mg on day 21, hMG/FSH 300 IU/day, IM). Compared with placebo, pyridostigmine was associated with a improved pregnancy rates (25.7% versus 11.4%).21
A recent Cochrane review 2010 showed a statistically significant difference in both live birthrates and pregnancy rates favoring the use of adjuvant growth hormone in in vitro fertilization protocols in women who are considered poor responders without increasing adverse events. The result needs to be interpreted with caution, the included trials were few in number and there was a small sample size. Therefore, before recommending growth hormone adjuvant in in vitro fertilization further research is necessary to fully define its role.22,23
 
Glucocorticoids
Dexamethasone may directly influence follicular development and oocyte maturation or may increase serum GH. It may also affect endometrium by immunosuppression. Only study conducted was on normal responders and showed lower cancelation rate. More RCT are needed before a conclusion can be drawn of its advantage in poor responders.
 
Adjunctive use of Nitric Oxide (NO)-Donors (L-arginine)
NO may be involved in follicular selection and maturation, possibly due to its contribution in periovulatory vasodilatory modulation. Better results were seen in a study with GnRH agonist flare-up regimen, high doses of purified FSH and orally administered L-arginine.24225
 
LH
In a recent study, exogenous LH supplementation was consistently associated with higher peak estradiol concentrations. The use of hMG in long GnRH agonist cycles was associated with a 3–4% increase in live birth- rate. There was insufficient evidence to make definitive conclusions on the need for exogenous LH activity in GnRH antagonist cycles or the benefit of recombinant LH and hCG protocols. Poor responders and patients 35 years of age and older may benefit from exogenous LH.25 Additional exogenous LH activity in the form of either recombinant luteinizing hormone or low-dose recombinant hCG is unnecessary in microdose cycles to increase pregnancy rates.26
 
Combined Oral Contraceptive (COC) Pills
COC administration prior to the GnRH agonist protocol was associated with higher pregnancy rates and lower cancelation rates. It suppresses endogenous gonadotropins, and at the same time (through its estrogen component), produces and sensitizes more estrogen receptors.27
 
Luteal Estradiol
Poor responders aged <35 years may be treated with the aggressive luteal estradiol/gonadotropin-releasing hormone antagonist (E2/ANT protocol to improve cycle.28
 
Aspirin
No improvement secondary to aspirin intake was found in IVF outcome in poor responders.29226
 
Letrozole
The GnRH antagonist with letrozole protocol is an effective protocol that may be used in poor ovarian responders. Study has shown it has better results than flare protocol.30
 
Complementary and Alternative Medicine
Acupuncture: Currently available literature does not provide sufficient evidence that adjuvant acupuncture improves IVF clinical pregnancy rate.31
 
Other Options
Oocyte Donation
Oocyte donation forms an important option for these women as they are unable to yield oocytes but endometrium can be prepared.
Cryopreservation of Oocyte or Embryo
Those with decreasing ovarian reserve embryo or oocyte cryopreservation can be offered.
There is insufficient evidence to support the routine use of any particular intervention either for pituitary downregulation, ovarian stimulation or adjuvant therapy in the management of poor responders to controlled ovarian stimulation in IVF. More robust data from good quality RCTs with relevant outcomes are needed.32
 
References
  1. Yoo JH, Cha SH, Park CW, Kim JY, Yang KM, Song IO, et al. Comparison of mild ovarian stimulation with conventional ovarian stimulation in poor responders. Clin Exp Reprod Med. 2011;38(3):159–63.
  1. Cheung LP, Lam PM, Lok IH, Chiu TT, Yeung SY, Tier CC, et al. GnRH antagonist versus long GnRH agonist protocol in poor responders undergoing IVF: a randomized controlled trial. Hum Reprod. 2005;20(3):616–21.
  1. Berkkanoglu M, Ozgur K. What is the optimum maximal gonadotropin dosage used in microdose flare-up cycles in poor responders? Fertil Steril. 2010;94(2):662–5.
  1. Dilbaz S, Demir B, Cinar O, Dede S, Aydin S, Beydilli G, et al. Does 75 IU difference improve the cycle performance in poor responders? Comparison of daily 375 versus 450 IU gonadotropin doses. Gynecol Endocrinol. 2011;27(12):1001–6.
  1. Raga F, Bonilla-Musoles F, Casan EM, Bonilla F. Recombinant follicle stimulating hormone stimulation in poor responders with normal basal concentrations of follicle-stimulating hormone and estradiol: improved reproductive outcome. Hum Reprod. 1999;14:1431–4.
  1. Eskandar M, Jaroudi K, Jambi A, Archibong EI, Coskun S, Sobande AA. Is recombinant follicle-stimulating hormone more effective in IVF poor responders than human menopausal gonadotrophins? Med Sci Monit. 2004;10(1):PI6–9.
  1. Faber BM, Mayer J, Cox B, Jones D, Toner JP, Oehninger S, et al. Cessation of gonadotropin-releasing hormone agonist therapy combined with high-dose gonadotropin stimulation yields favorable pregnancy results in low responders. Fertil Steril. 1998;69:826–30.
  1. Padilla S, Dugan K, Maruschak V, Shalika S, Smith R. Use of the flare-up protocol with high dose follicle stimulating hormone and human menopausal gonadotropins for in vitro fertilization in poor responders. Fertil Steril. 1996;65:796–9.
  1. Toth TL, Awwad JT, Veeck L, Jones HWJr, Muasher SJ. Suppression and fare regimens of gonadotropin-releasing hormone agonist: use in women with different basal gonadotropin values in an in vitro fertilization program. J Reprod Med. 1996;41:321–6.
  1. Akman MA, Erden HF, Tosun SB, Bayazit N, Aksoy E, Bahceci M. Comparison of agonistic flare-up protocol and antagonistic multiple dose protocol in ovarian stimulation of poor responders: results of a prospective randomized trial. Hum Reprod. 2001;16(5):868–70.
  1. Schoolcraft W, Schlenker T, Gee M, Stevens J, Wagley L. Improved controlled ovarian hyperstimulation in poor responder in vitro low ovarian response to stimulation for IVF. Fertil Steril. 1997;67:93–7.
  1. Kahraman K, Berker B, Atabekoglu CS, Sonmezer M, Cetinkaya E, Aytac R, et al. Microdose gonadotropin-releasing hormone agonist flare-up protocol versus multiple dose gonadotropin-releasing hormone antagonist protocol in poor responders undergoing intracytoplasmic sperm injection-embryo transfer cycle. Fertil Steril. 2009;91(6):2437–44.
  1. Craft I, Gorgy A, Hill J, Menon D, Podsiadly B. Will GnRH antagonists provide new hope for patients considered difficult responders’ to GnRH agonist protocols? Hum Reprod. 1999;14:2959–62.
  1. Mohsen IA, El Din RE. Minimal stimulation protocol using letrozole versus microdose flare up GnRH agonist protocol in women with poor ovarian response undergoing ICSI. Gynecol Endocrinol. 2013;29(2):105–8.
  1. Lou HY, Huang XY. Modified natural cycle for in vitro fertilization and embryo transfer in normal ovarian responders. J Int Med Res. 2010;38(6):2070–6.
  1. Elizur SE, Aslan D, Shulman A, Weisz B, Bider D, Dor J. Modified natural cycle using GnRH antagonist can be an optional treatment in poor responders undergoing IVF. J Assist Reprod Genet. 2005;22(2):75–9.
  1. Kim CH, Howles CM, Lee HA. The effect of transdermal testosterone gel pretreatment on controlled ovarian stimulation and IVF outcome in low responders. Fertil Steril. 2011;95(2):679–83.
  1. Sönmezer M, Ozmen B, Cil AP, Ozkavukçu S, Taşçi T, Olmuş H, et al. Dehydroepiandrosterone supplementation improves ovarian response and cycle outcome in poor responders. Reprod Biomed Online. 2009;19(4):508–13.
  1. Bosdou JK, Venetis CA, Kolibianakis EM, Toulis KA, Goulis DG, Zepiridis L, et al. The use of androgens or androgen-modulating agents in poor responders undergoing in vitro fertilization: a systematic review and meta-analysis. Hum Reprod Update. 2012;18(2):127–45.
  1. Jia N, Kalmijn J, Hseuh A. Growth hormone enhances FSH induced differentiation of cultured rat granulosa cells. Endocrinology. 1986;118:1401–9.
  1. Chung-Hoon K, Hee-Dong C, Yoon-Seok C. Pyridostigmine co-treatment for controlled ovarian hyperstimulation in low responders undergoing in vitro fertilization ± embryo transfer. Fertil Steril. 1999;71:652–7.
  1. Duffy JM, Ahmad G, Mohiyiddeen L, Nardo LG, Watson A. Growth hormone for in vitro fertilization. Cochrane Database Syst Rev. 2010 Jan 20;(1):CD000099.
  1. Kolibianakis EM, Venetis CA, Diedrich K, Tarlatzis BC, Griesinger G. Addition of growth hormone to gonadotropins in ovarian stimulation of poor responders treated by in-vitro fertilization: a systematic review and meta-analysis. Hum Reprod Update. 2009;15(6):613–22.
  1. Battaglia C, Salvatori M, Maxia N, Petraglia F, Facchinotti F, Volpe A. Adjuvant L-arginine treatment for in-vitro fertilization in poor responder patients. Hum Reprod. 1999;14:1690–7.
  1. Hill MJ, Levy G, Levens ED. Does exogenous LH in ovarian stimulation improve assisted reproduction success? An appraisal of the literature. Reprod Biomed Online. 2012;24(3):261–71.
  1. Berkkanoglu M, Isikoglu M, Aydin D, Ozgur K. Clinical effects of ovulation induction with recombinant follicle-stimulating hormone supplemented with recombinant luteinizing hormone or low-dose recombinant human chorionic gonadotropin in the midfollicular phase in microdose cycles in poor responders. Fertil Steril. 2007;88(3):665–9.
  1. Lindheim S, Barad D, Witt B, Ditkoff E, Sauer M. Short-term gonadotrophin suppression with oral contraceptives benefits poor responders prior to controlled ovarian hyperstimulation. J Assist Reprod Genet. 1996;16:745–7.
  1. Shastri SM, Barbieri E, Kligman I, Schoyer KD, Davis OK, Rosenwaks Z. Stimulation of the young poor responder: comparison of the luteal estradiol/gonadotropin-releasing hormone antagonist priming protocol versus oral contraceptive microdose leuprolide. Fertil Steril. 2011;95(2):592–5.
  1. Frattarelli JL, McWilliams GD, Hill MJ, Miller KA, Scott RT Jr. Low-dose aspirin use does not improve in vitro fertilization outcomes in poor responders. Fertil Steril. 2008;89(5):1113–7.
  1. Yarali H, Esinler I, Polat M, Bozdag G, Tiras B. Antagonist/letrozole protocol in poor ovarian responders for intracytoplasmic sperm injection: a comparative study with the microdose flare-up protocol. Fertil Steril. 2009;92(1):231–5.
  1. El-Toukhy T, Sunkara SK, Khairy M, Dyer R, Khalaf Y, Coomarasamy A. A systematic review and meta-analysis of acupuncture in vitro fertilisation. BJOG. 2008;115(10):1203–13.
  1. Shanbhag S, Aucott L, Bhattacharya S, Hamilton MA, McTavish AR. Interventions for ‘poor responders’ to controlled ovarian hyperstimulation (COH) in in-vitro fertilisation (IVF). Cochrane Database Syst Rev. 2007 Jan 24;(1):CD004379.

Endometrial Receptivity—A Vital Rolechapter 15

Surveen Ghumman
Despite progress and research in the field of infertility, the rate of live birth rate with IVF has not gone beyond 40%. In 75% of the failed cases, there is no cause for failure other than implantation forming a major obstacle to ART. The complexities of embryo apposition to invasion of the epithelium are only partly understood at the molecular level and still present a challenge to the infertility specialist. Both partners, the mother as well as the embryo, play equal roles in the embryo-maternal dialog.
Definition: Endometrial receptivity can be defined as the histological and molecular changes occurring in a temporal and spatial manner in the endometrium so as to facilitate embryonic implantation.
Endometrial receptivity (ER) involves molecular as well as histological changes occurring in a temporal and spatial manner in the endometrium so as to facilitate embryonic implantation. Under the influence of estrogen and progesterone, the endometrium undergoes these important 232changes so as to make it receptive to the implanting embryo and this period is known as the ‘Window of Receptivity’. It lasts for approximately 4 days usually from day 20 to 24 in women with a 28 day menstrual cycle. The key factor for implantation is the synchrony between embryo development and endometrial receptivity. The ability of the decidua to respond optimally to the invading trophoblasts is determined by endocrine and end organ interactions that long precede ovulation. There are many causes of poor endometrial response (Table 15.1).
The implantation process, as described by Enders, consists of the three important phases.1
Table 15.1   Causes of poor endometrial response
Poor endometrial response
Causes
Poor hormonal environment
Suboptimal estrogens
Suboptimal progesterones
Out of phase development
Excessive estrogen levels
Hyperprolactinemia
Hyperandrogenemia
Infections
Anatomical
Endometritis
Tuberculosis
Chlamydia
Uterine septum
Foreign bodies
Leiomyomas
• Submucous
• Multiple/Large Intramural
Endometrial polyps
Lost intrauterine devices
Iatrogenic
Vigorous curettage
Adhesions
Intrauterine synechiae
Drugs
Clomiphene citrate
Endometrial
Squamous metaplasia
Calcification/Ossification
233
  1. Apposition: The blastocyst at this phase ceases to move freely in the uterine cavity and comes in close proximity to the uterine epithelium. This is facilitated not only by absorption of uterine fluid but also by elevating the endometrial surface toward the blastocyst, by pinopode formation.
  2. Adhesion: It is believed that the sulfhydryl groups of the endometrium are chemotactic for the embryo.2 This approximation leads to an interaction between the trophoectoderm and the endometrial epithelial cells leading to the attachment of the blastocysts to the uterine lining. The attachment can only occur if the embryo has successfully hatched out of the zona pellucida. In cases of zona hardening and improper hatching, the embryo will be unable to adhere to the endometrium. Hence, Assisted Hatching procedures are useful in a select group of patients. Blastocyst culture has taught us that the blastocyst is sticky and will readily adhere, a quality that makes it survive in the endometrium.
    A number of adhesion molecules have been identified that are secreted by the endometrial cells and facilitate adhesion. There are:
    • Integrins: Integrins are maximally expressed in the endometrium during the implantation window. The integrins α1β1 and α4β1 are co-expressed in the endometrial epithelial cells in varying amounts. However, α5β3 integrin has been maximally studied as a potential marker for endometrial receptivity. It is secreted under the influence of progesterone during the implantation window. Low levels of this integrin were found in women with unexplained infertility, attending for IVF, ICSI compared to fertile controls.3
    • Osteopontin: Osteopontin is a glycoprotein whose expression is increased in the endometrium during the implantation window, i.e. 7 days after the LH peak. 234Osteopontin and its receptor alpha (v) beta (3) integrin have recently been proposed as a major complex to promote embryo attachment, and thus, they would be useful as markers of endometrial receptivity.4
    • Mucin 1(MUC 1): This cell surface mucin is abundantly expressed during the implantation window.5 Glandular MUC 1 levels were found to be diminished in patients with recurrent miscarriages.
  3. Invasion: This involves epithelial penetration and placentation. The trophoectoderm cells, taking advantage of the loss of polarity and tight junctions between the epithelial cells that occurs during this period, invades into the endometrium. The trophoblasts produce serine proteases and matrix metalloproteinases that help to digest the matrix.
    Several factors affect the process of implantation (Table 15.2).
Table 15.2   Factors that affect implantation
1. General
  • Age
  • Hormonal control of endometrial preparation
  • Endometriosis
  • Hydrosalpinx
2. Uterine factors
  • Endometrial receptivity
  • Congenital uterine abnormalities
  • Fibroids/Polyps
  • Endometritis
  • Poor uterine artery blood flow
3. Immunological function
  • Antiphospholipid syndrome
  • Lupus erythematosis
  • Rheumatoid arthritis
  • Hashimoto's thyroiditis
4. Embryological
  • Aneuploidy in embryos
  • Spindle damage
235
 
Histological Changes
The most significant morphological change observed during this period is the development of pinopodes, which are closely associated with the development of endometrial receptivity and could be an important histological marker.6 Pinopodes are large cytoplasmic protrusions of the apical membranes of the secretary cells after they lose their microvilli. They develop 4–5 days after ovulation, i.e. day 18 of a 28 day cycle and are fully developed by day 20 after that they start regressing, largely disappearing by day 22. They are thought to have a pinocytotic function and are involved in the apico-basal transport of fluids and macromolecules toward the stroma.
The other important change seen at this time is the decrease in cell polarity and tight junctions between cells thus assisting in trophoblastic invasion.
 
Biochemical and Molecular Changes
The various phases of implantation involve synchronized interplay between a variety of molecules. These molecules, expressed at the three stages of implantation, are in accordance with the needs of that phase (Table 15.3).
 
Biochemical Markers of Endometrial Receptivity
Mucins: These are highly glycosylated large molecules found on the apical site of the endometrial epithelial cells and are believed to play a role in attachment. Mucin named MAG's (mouse ascites Golgi) use as a marker for ER has been suggested and 60% of women with unexplained infertility have an abnormal MAG expression.7 236
Table 15.3   Molecules at three stages of implantation
1. Apposition – Chemokines
  • Interleukin-8 (IL-8)
  • Monocyte chemoattractant protein-1 (MCP-1)
  • Regulated on activation, normal T-cell expressed and secreted (RANTES)
2. Adhesion
  • Cytokines LIF, IL-1
  • Trophonin, tastin, osteopontin
  • Uteroglobin
  • Glycodelin A
  • Growth factors IGFBP1, HBEGF
  • Integrins alpha V beta 3
  • HOXA10 gene
3. Invasion – Proteolytic enzymes
  • Serine proteases
  • Metalloproteases
  • Collagenases
Integrins: These are transmembrane glycoprotiens belonging to the family of cell-adhesions molecules and promote cell-cell binding. Defects of integrin expression leads to disordered endometrial function The alpha5 beta3 integrins are emerging as important markers of endometrial receptivity and are absent in cases of unexplained infertility,8 luteal phase defect, endometriosis and hydrosalpinx.9
Trophinin: It is an intrinsic membrane protein important for cell adhesion whereas tastin (trophinin assisting protein) is a cytoplasmic protein.10 It has been observed that trophinin concentrates on the endometrial surface at possible sites for blastocyst attachment.
Growth Factors: Epidermal growth factor (EGF) and HBEGF (heparin binding epidermal growth factor) have a role, both in attachment and penetration during the implantation process.11 Insulin like growth factor binding protein 1 (IGFBP1) also plays an important role in embryo implantation. 237Increased levels of IGFBP1 are seen at maternal-fetal interface in severe preeclampsia that is characterized by inadequate placentation.
Cytokines: Cytokines are low molecular weight soluble proteins involved in regulating cellular activity.12 They act as messengers within the immune system and between the immune system and other systems of the body. The presence of cytokines is sensed by the cell by means of specific cytokine receptors. Cytokines may be pro-inflammatory or anti-inflammatory in nature (Table 15.4). The cytokines that play an important role in the cascade of events that lead to implantation are leukemia inhibiting factor (LIF) that is decreased in women with unexplained infertility,13 and colony-stimulating factor (CSF) that interacts with the receptors on trophoectoderm and promotes blastocyst attachment. The most potent anti-inflammatory cytokine is interleukin (IL).10
Calcitonin: It is secreted under the influence of progesterone in the uterine lumen 4 days post ovulation. It causes changes in the calcium signaling leading to redistribution of critical cell adhesion molecules or junctional complexes at the site of implantation, preparing it for contact with the trophoblast.
HOXA 10 is a hormone-regulated endometrial transcription factor gene that appears during the window of implantation. Homeobox A10 is a protein that is encoded by Hoxa-10 gene.
Table 15.4   Pro- and anti-inflammatory cytokines
Pro-inflammatory Cytokines
Anti-inflammatory cytokines
Interleukin-2 (IL2)
Interleukin-4 (IL4)
Interferon gamma (IFN)
Transforming growth factor beta (TGFβ)
Interleukin-1 (IL1)
Interleukin 10 (IL10)
Tumor necrosis factor alpha (TNFα)
Interleukin 6 (IL6)
Leukemia inhibitory factor (LIF)
238
It was discovered that maternal expression of Hoxa-10 transcription factor is necessary for female fertility and diminished Hoxa-10 expression has been demonstrated in women with infertility.14
Cyclo-oxygenase-2 (COX-2): It is the rate limiting enzyme in prostaglandin biosynthesis. Prostaglandins are involved in increase vascular permeability during implantation and decidualization. Targeted disruption of COX-2 in mice produces reproductive failures in the female.15
Fibronectin: During implantation contact between fibronectin and peri-implantation blastocyst elevates intracellular calcium that strengthens trophoblast adhesion through protein redistribution.16
Corticotropin-releasing hormone (CRH): The hypothalamic neuropeptide CRH as well as its receptors have been identified in decidualized endometrial stroma as well as the trophoblast cells. It is believed that it may play a role in local immune phenomenon associated with embryo implantation.17
 
Immunological Aspects of Endometrial Implantation
The conceptus is a foreign allograft to the mother as it contains paternal antigens. The appropriate interaction between the preimplantation embryo and maternal endometrium is controlled by cytokines and growth factors and its receptors. Both the cytokines and growth factors may react through ‘embryo endometrial dialog’ by helpful or harmful responses. According to these responses either pregnancy is maintained or rejected. During blastocyst implantation, the maternal endometrial response to the invading embryo has characteristics of an aseptic acute inflammatory response yet once implanted the embryo suppresses this response and prevents its rejection.239
 
Th1 vs Th2 Response of Endometrium
In pregnancy, the Th1 subset of cytokines are downregulated and the Th2 subset are upregulated. It is well known that the reproductive steroid hormones, particularly progesterone, in addition to its widely recognized effects on endometrial epithelial and stromal cells and spiral arteries, affect the activities of T cells and natural killer cells in the decidua, thus inducing active immune tolerance against the fetal antigens.18 It immunomodulates and makes Th2 response predominate for pregnancy continuation.
Decidual antigen-presenting cells including dendritic cells (DCs) and CD14(+) macrophages, as mediators of the first encounter with fetal antigens, appear to be critically involved in the initiation of primary immune response by regulating innate and adaptive immunity.19 Interleukin-15, produced by them, permits the proliferation and differentiation of CD3(–) CD16(–) CD94(+) NKG2A(+) CD56(+bright) and decidual NK cells that identify trophoblast cells. Large granular lymphocytes (LGL) comprise 70 – 80% of the endometrial leukocyte population and play a role in implantation and maintenance of pregnancy.
 
Natural Killer Cells
The role of natural killer (NK) cells in human implantation has recently attracted attention. Women with unexplained recurrent abortion and infertile women in whom multiple attempts at embryo transfer have failed, show elevated levels of peripheral and endometrial CD56 (+), CD16 (+), NK cells. Daily administration of prednisolone for 3 days has been reported to reduce the percentage of peripheral blood NK cells. It is possible that steroid had beneficial effect on IVF outcome through reduction of NK cells.20240
 
T Regulatory (T Reg) Cells
These cells are essential for maternal tolerance of the conceptus. Inadequate number of T reg cells or their functional deficiency is linked with infertility miscarriage and preeclampsia. There is potential to use these cells as therapeutic agents for reproductive pathologies.21
Several lines of evidence suggest that the human leukocyte antigen (HLA)-G plays a key role in the regulation of human pregnancy.22
 
The Fas – FasL System and Immune Tolerance
Jiaang and Vacchio showed that placental trophoblast can induce fas mediated cell death of T cells that recognize fetal antigens as foreign.23 This would protect the fetus from harmful effects of maternal cell-mediated immunity. FasL is present both in the villous cytotrophoblast and in the extravillous trophoblast.
 
Newer Molecules Identified in Endometrial Implantation
Search is on through the field of proteomics and metabolomics for newer molecules that play a key role in implantation. The ones that are currently being investigated are:
  • Glycodelin A is a progesterone-induced endometrial glycoprotein that has been amply documented to play a role in down-modulation of the maternal immune response to fetal allo-antigens and to be indispensable for the maintenance and progression of pregnancy.24
  • Uteroglobin is a progesterone binding protein, a member of the anti-inflammatory gene family and possibly a novel cytokine. Secretory uteroglobin is found in endometrial tissue homogenates in highest levels of expression during 241the early and mid luteal phase strongly suggesting an involvement of uteroglobin in endometrial preparations for implantation.25
  • Osteoprotegerin (OPG), a soluble receptor of the tumor necrosis factor family, and its ligand, the receptor activator of nuclear factor-B ligand (RANKL), are emerging as important regulators of vascular pathophysiology. OPG is a positive regulator of microvessel formation in vivo that is required for successful embryo nidation.26
 
Endometrial Vascular Changes
  • There are marked changes in the vascularity of the endometrium during the menstrual cycle as demonstrated by Doppler. The lowest impedance to blood flow is during the secretory phase around implantation. This increase in vascularity leads to endometrial (epithelial and stromal) growth and leads to efficient distribution as well as expression of molecular biomarkers required for preparation of the endometrium for implantation. Levels of vascular endothelial growth factor increase in the endometrium to facilitate implantation.
 
Role of Chronic Endometritis in Endometrial Receptivity
Chronic endometritis may be:
  1. Tubercular endometritis.
  2. Nonspecific chronic endometritis.
Tuberculosis and chronic nonspecific endometritis has a high incidence in unexplained infertility, recurrent implantation failure and abortions.242
 
Tubercular Endometritis
Tuberculosis is an infection that can lie latent and be reactivated at any time. Balance of Th2 to Th1 may be disturbed by tubercular bacilli leading to implantation failure or pregnancy loss. Mycobacterium may inhibit basal production of progesterone and stimulatory effect of hCG. Hence, it may cause a luteal phase effect and lead to failure of implantation. A direct antigonadotropin effect has been seen, and women with tuberculosis have needed higher doses of gonadotropins for response. They have a higher basal FSH showing a poor ovarian reserve.27
A high incidence of implantation failure has been seen in women with genital tuberculosis and it is suggested that latent tuberculosis should be considered in young Indian patients presenting with unexplained infertility with apparently normal pelvis and non-endometrial tubal factors or those with repeated IVF failure.28
 
Chronic Nonspecific Endometritis
Chronic endometritis is a persistent inflammation of endometrial lining that may be asymptomatic or accompanied by mild symptoms like pelvic pain, abnormal uterine bleeding, dyspareunea and vaginal discharge. It can be caused by infection, intrauterine contraceptive device, submucosal leiomyoma and endometrial polyp. Chronic endometritis occurred in 22% of IVF program, 14% unexplained infertility, 23.6% women with history of first trimester miscarriage and 30.3% of recurrent implantation failure.29 Cases with bacterial vaginosis have a high incidence of chronic nonspecific endometritis (45%vs 5.2%) and should be screened.30
Pathogenesis of implantation failure with chronic endometritis is shown in Figure 15.1. Endometrial lymphocytes play a critical role in endometrial receptivity. There were 243lower % of CD56(+), CD16, CD56 whereas CD3 was significantly higher in cases of chronic endometritis.
Fig. 15.1: Pathogenesis of implantation failure with chronic endometritis
 
Diagnosis of Chronic Nonspecific Endometritis
It can be diagnosed by histopathology, culture or hysteroscopy. Histopathological features include plasma cells, lymphocytes, macrophages, eosinophils, spindle stroma, epithelial changes, vasculitis, hyperemia, mucosal edema, micropolyps and adhesions.
Problems with diagnosing chronic endometritis (CE):
  1. No typical findings at clinical examination or ultrasound
  2. Diagnosis relies on histology that has the following problems
    • Number of plasma cells does not correlate with symptoms
    • Intensity of inflammation showed no association with patient's duration of symptoms244
    • 16% pathological underdiagnosis mostly because of non-recognition of plasma cells as they are obscured by mononuclear cell infiltrate, plasmacytoid stromal cell and abundant stromal mitosis.
      Immunohistochemistry with syndecan 1: Immunohistochemistry with syndecan 1 (a proteoglycan found on the surface of plasma cells) would detect these cells.31,32 It should be used for detection, only in high-risk groups like those with recurrent implantation failure and abortions, unexplained infertility, abnormal uterine bleeding where no cause found and those with characteristic milieu of CE in that plasma cells not found on hematoxylin and eosin (H&E) stain.
  3. Endometrial cultures—There can be contamination from vaginal and endocervical contents. Demonstration of organism does not imply its pathogenic significance. A poor concordance is seen between endocervical, vaginal and endometrial cultures.33 Hence, an endometrial culture should be sent to confirm micro-organism responsible for endometritis. Antimicrobial therapy should be initiated in chronic non- specific endometritis only after organism is identified since, besides chlamydia and gonococcus, other organisms like E. coli, Staphylococcus, Streptococcus and U. urealyticum have a high incidence.
  4. Hysteroscopy: The findings on hysteroscopy are:
    • Focal or diffuse hyperemia
    • Mucosal edema
    • Adhesions
    • Micropolyps.
A high degree of inflammation is necessary for development of micropolyps. Hysteroscopy is not a useful screening test for chronic endometritis in asymptomatic women - low prevalence population.245
Chromohysteroscopy—On introduction of methylene blue dye, areas stained with dye can detect chronic endometritis as dye is taken up by damaged endometrium. These areas may be sent for biopsy. Detection rate of chronic endometritis is improved with a targeted biopsy. Observation of diffuse light blue staining without dark areas strongly suggests a normal endometrium free of endometritis. In a study the power of dark staining for detection of endometritis was calculated as follows—sensitivity 69.2%, specificity 74%, positive predictive value 40.9% and negative predictive value 90.2%.34
 
Current Strategies to Assess Endometrial Receptivity
The implantation rates even today are not as yet efficient as those in a natural conception cycle. Unfortunately, there are no universally accepted markers of endometrial receptivity and currently a working definition of a receptive versus non- receptive endometrium is incomplete.
Tests that assess receptivity can be grouped under two major headings (Table 15.5) (Fig. 15.2).
  1. Tests that assess endometrial changes:
    1. Endometrial aspiration/biopsy:
      1. Endometrial Histopathology: This is done by endometrial biopsy taken during the late luteal phase. This was the gold standard to assess endometrial maturity. However, there are limitations to this procedure as it is preformed in the late luteal phase and hence may give information only of invasion phase. It is representative of the endometrial changes within the cycle and cycle variations are well known. However, it cannot be carried out in the cycle 246in which the patient is undergoing ART. It does not reflect the endometrium as a whole, regional variations being frequently encountered.
        Table 15.5   Methods of endometrial evaluation
        I. Tests to assess endometrial changes
        1. Endometrial aspiration/biopsy
          • Endometrial histopathology
          • Study of pinopodes
          • Endometrial culture-AFB/Bactec
          • PCR for tuberculosis
        2. Ultrasonography & Doppler, MRI
        3. Hormonal evaluation
        4. Hysteroscopy for anatomical and infectious cause
        II. Markers of the embryo-endometrial dialogue
        • Biochemical markers – endometrial proteins
        • Uteroglobin & Glycodelin A
      2. Study of Pinopodes: Pinopodes are studied by scanning electron microscopy (SEM). Though a better indicator of endometrial receptivity, it is expensive, not universally available and cannot be carried out during an IVF cycle.
      3. Culture: Culture should be sent for AFB or Bactec and for other organisms like Chlamydia, gonococcus, E. coli, Staphylococcus, Streptococcus and U. urealyticum
      4. PCR: PCR for tuberculosis should be sent.
  2. Ultrasound and Doppler: Transvaginal sonography is a simple noninvasive modality that is being increasingly used to assess endometrial receptivity. Both endometrial thickness and echogenic pattern have been studied as potential markers of endometrial receptivity. Calculating total endometrial volume by a new 3D software is more objective than endometrial thickness alone. Using Doppler the impedance to 247blood flow in the uterine artery is expressed as the pulsatility index (PI) and is the lowest at the time of implantation.
    Fig. 15.2: Evaluation and management of poor endometrium
    1. Endometrial thickness: It is generally accepted that if the thickness is < 7 mm on ultrasound the implantation is poor. Similarly endometrial 248volume of <2.5 cc is associated with a poor pregnancy rate.
    2. Echogenicity: Endometrial character is hypoechoic compared to the surrounding myometrium in the proliferative phase. As thickness increases a distinct triple line/multilayered pattern is seen and is considered to be predictive of implantation. Further under the influence of progesterone, the endometrium undergoes secretory changes and becomes more isoechoic and then hyperechoic. A non-multilayered endometrium is associated with poor implantation.
    3. Endometrial vascularity: Sub- and intra-endometrial vascularity is a prognostic factor for implantation if endometrium is more than 7 mm irrespective of the morphological index. Uterine perfusion is maximum during the mid-luteal phase. PI < 3 is associated with increased pregnancy rate. Absence of sub-endometrial blood flow on the day of LH surge is related to implantation failure.
      Spiral artery perfusion is evaluated by color/pulse Doppler and the endometrium has been divided into 4 zones.
      Zone 1—Only myometrial vessels surrounding endometrium are seen
      Zone 2—Vessels penetrate through the hyperechogenic endometrial edge
      Zone 3—Vessels reach the internal endometrial hypoechogenic zone
      Zone 4—Vessels reach up to the endometrial cavity
      A good vascularity in zone 3 and 4 relates to the surface of the endometrium suggesting a good endometrial receptivity.249
    4. Hormonal levels: are of not much use for assessment of endometrial receptivity.
    5. Hysteroscopy: Hysteroscopy may detect anatomical lesions missed earlier. Hysteroscopy may show evidence of endometritis in the form of focal or diffuse hyperemia, white spots, micropolyps and intrauterine adhesions.
  3. Markers of the Embryo-endometrial dialog:
    1. Evaluation of the various biomarkers are mostly research tools. Kits for evaluation of integrins and mucins are now commercially available. (Table 15.3)
    2. Peripheral NK cells: The pNK cell levels reflect changes in dNK cell levels. This implicates that pNK cell level is a clinically useful marker to predict pregnancy outcome.35
 
Treatment of Poor Uterine Receptivity
Treatment of poor endometrial receptivity must be done according to the cause (Fig. 15.2).
 
Estrogens
Where endometrial response is suboptimal as often occurs with clomiphene citrate, treatment is given by supplementing estrogen from day 7–21 of cycle or till plasma estradiol is 400–700 pg/mL. Premarin 0.625 mg/day was used earlier; however, with the advent of natural estrogens, the drug of choice is estradiol valerate 2–8 mg per day orally from 8th day of cycle. Vaginal estradiol gel can also be administered.36 However, recent studies have not supported the use of estrogen.250
 
Change of Ovulation Induction Regime
Patients on clomiphene may show persistently poor endometrial response due to its antiestrogenic effect on endometrium. In such cases drug may be changed to letrozole, tamoxifen or gonadotropins.
  • Letrozole: Letrozole does not have an antiestrogenic effect on the endometrium because of its short half-life and absence of estrogen receptor blockage.
  • Tamoxifen: It is a selective estrogen receptor modulator. It causes raised estrogen levels not only due to multi-follicular development but also due to a direct action on the endometrial estrogen receptors, hence leading to a favorable response on the cervical mucus and endometrium. It is an alternative to clomiphene where there is persistently poor endometrial response.
  • Gonadotropins: Use of gonadotropins will enhance the estrogenic effect on the endometrium.
 
Drugs to Improve the Endometrial Blood Flow
  • Aspirin: Low dose aspirin, 75 mg/day reduces platelet aggregation and may enhance endometrial blood flow.37
  • Sildenafil: It is given in a dose of 25 mg four times a day intravaginally for 3–10 day to improve endometrial vascularity.38 It is a type 5 specific phosphodiesterase inhibitor. Pulsatility index decreases from 3 to 2.1 whereas there is no effect in the placebo group. This, however, is an observational study and is not supported by randomized control trials. It should be used with caution as it has side effects like headache, hypertension and occasional death.
  • Nitroglycerin: It was thought to be useful because of its vasodilating effect and was given in a dose of 800 μg 251sublingually 3 minutes before embryo transfer in IVF or 5 mg daily patch prior to the day of embryo transfer. However, a double blind prospective randomized placebo controlled trial showed that NTG treatment on the day before embryo transfer was no more effective than placebo in improving the implantation rates.39
  • L-arginine: Nitric oxide (NO) is formed from L-arginine and leads to increased vascularity and improves blood flow in the ovarian follicle and endometrium. It is given in a dose of 16 g/day in poor responders and implantation failures. However, randomized placebo controlled trials are lacking on this subject except one in poor responders.40
 
Immune Suppression/Potentiation
Administration of intravenous immunoglobulins (IVIG) has been tried in patients with high titers of antiphospholipid antibodies to mitigate its harmful effects on nidation. It has also been tried in patients with prior failed IVF cycles to improve endometrial receptivity. In one observational study early IVIG therapy was associated with improved success of IVF.41 Elevated NKT cells in recurrent pregnancy loss or implantation failure can be ameliorated with IVIG treatment, and result in successful pregnancy.42
Leukocyte immunotherapy (LIT) by paternal leukocytes was reported to be successful in a paper published from Germany.43 In 20 patients with a history of unsuccessful sterility treatment, paternal leukocytes are injected into the mother in an attempt to alter the maternal immune response, thus making it favorable for implantation. However, a Cochrane review on the use of paternal leukocytes and intravenous immunoglobulins, analyzed 28 trials and showed no benefits of such treatment over placebo.44252
 
Reducing Uterine Contractility
Reducing uterine contractility will give a better contact of the embryo with the endometrium:
  1. Ritodrine: Administration of this drug has shown better pregnancy rate in randomized controlled trials.
  2. Piroxicam: In a randomized controlled trial 10 mg of piroxicam 1 to 2 hour before embryo transfer showed 20% improvement in pregnancy rate. Recent studies, however, have shown no improvement in success rate.45
 
Luteal Phase Support
Treatment consists of giving micronized progesterone per vaginum or per rectal in a dose of 200–400 mg in two divided doses. However, the intramuscular route in a dose of 50–100 mg daily gives more sustained levels of progesterone. Dydrogesterone can also be given orally in a dose of 20–30 mg/day in divided doses. hCG administration in a dose of 2500 IU every 3 days would provide luteal support. Progesterone production is improved by hCG in normally functioning corpus luteum whereas its effect is minimal if there is a malfunctioning corpus luteum.
 
Surgical Evaluation and Treatment
Lysis of uterine synechiae: In cases of adhesion following curettage or endometritis, hysteroscopic lysis of adhesions should be done. Patients are treated with oral estrogens for 1 to 3 cycles following this to allow the endometrium to regenerate.
Drainage of hydrosalpinx: It is seen that fluid from hydrosalpinx impairs implantation. Hence, drainage of hydrosalpinx or salpingectomy should be performed laparoscopically prior to ART procedures.46 In cases with dense 253peritubal adhesions, e.g. genital kochs, tubal delinking at the cornual end will prevent the backflow of hydrosalpinx fluid.
 
Medical Treatment of Endometritis
Treatment of endometritis is a must. If tubercular etiology is present, antitubercular treatment is given. In case of chronic nonspecific endometritis, it is imperative to identify the organism by an endometrial culture and give antimicrobial therapy accordingly. In case biopsy shows chronic non- specific endometritis but culture is negative, a course of metronidazole and azithromycin is given. Often these infections are difficult to eradicate and antibiotics may be needed for a longer period. The response to treatment is seen in 50%.
 
Treatment of other Hormonal Pathologies
Hyperprolactinemia is treated with dopamine agonists. Hyperandrogenemia is treated with antiandrogenic drugs. This may improve endometrial receptivity.
 
Endometrial Preparation for Frozen Embryo Transfer
Many regimes have been used for endometrial preparation in donor oocytes or frozen ET. Estradiol supplementation is started with 2 mg/twice a day and increased up to 2 mg/thrice a day depending on endometrial response. 17 beta-estradiol transdermal patches at steadily increasing dosage from 100 to 300 μg have been given for at least 12 days. This was increased by 100 μg after 7 days.
A recent Cochrane review showed no difference in pregnancy rate when no treatment was compared to 254aspirin, steroids, ovarian stimulation, or human chorionic gonadotropin (hCG) prior to embryo transfer. No significant benefit for using GnRH agonists was found. Starting progesterone on the day of oocyte pick-up (OPU) or the day after OPU produced a significantly higher pregnancy rate than when recipients started progesterone the day prior to OPU. So there is insufficient evidence to recommend any one particular protocol for endometrial preparation over another with regard to pregnancy rates after embryo transfers.47
It is important for a successful pregnancy to have a good endometrium. Preparation of the endometrium involves a systematic interplay of hormones, cytokines, growth factors and many as yet unidentified molecules. Diagnostic tests for endometrial quality have limitations (Fig. 15.2). There are a number of biomarkers suggested but none of them is an undisputed biomarker. Improving endometrial quality can be challenging. Many suggested therapies are still empirical. Considerable research is ongoing in this area.
 
References
  1. Enders AC. Contributions of comparative studies to understanding mechanisms of implantation. In: Glasser SR, Mulholland J, Psychoyos A (Eds). Endocrinology of Embryo-Endometrium Interactions. Raven Press.  and London: Plenum Press.  1994;pp11-6.
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  1. Wang J, Mayernik L, Armant DR. Intergrin signaling regulates blastocyst adhesion to fibronectin at implantation: intracellular calcium transients and vesicle trafficking in primary trophoblast cells. Dev Bio. 2002;245:270–9.
  1. Sophia N Klantaridou, Antonis Makrigiannakis, Emmanouil Zoumakis, George P Chrousos. The Role of corticotropin-releasing Hormone (CRH) on Implantation and Immunotolerance of the fetus in Immunology of Pregnancy 2006 Landes Bioscience ISBN 0-387-30612-9.
  1. Kyurkchiev D, Ivanova-Todorova E, Kyurkchiev SD. New target cells of the immunomodulatory effects of progesterone. Reprod Biomed Online. 2010;21(3):304–11.
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  1. Tuckerman E, Mariee N, Prakash A, Li TC, Laird S. Uterine natural killer cells in peri-implantation endometrium from women with repeated implantation failure after IVF. J Reprod Immunol. 2010;87(1-2):60–6.
  1. Guerin LR, Prins JR, Robertson SA. Regulatory T cells and immune tolerance in pregnancy: a new target for infertility treatment. Hum Reprod Update. 2009;15(5):517–35.
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  1. Jiang SP, Vacchio MS. Multiple mechanisms of peripheral T cell tolerance to the fetal “allograft”. J Immunol. 1998;160:3086–90.
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  1. Benslimane-Ahmim Z, Heymann D, Dizier B, Lokajczyk A, Brion R, Laurendeau I, et al. Osteoprotegerin, a new factor in vasculogenesis, stimulates endothelial colony-forming cells properties. J Thromb Haemost. 2011;10:1538–7836.
  1. Kumar A, Rattan A. Antigonadotrophic effect of Mycobacterium tuberculosis. Horm Metab Res. 1997;29(10):501–3.
  1. Dam P, Shirazee HH, Goswami SK, Ghosh S, Ganesh A, Chaudhury K, et al. Role of latent genital tuberculosis in repeated IVF failure in the Indian clinical setting. Gynecol Obstet Invest. 2006;61(4):223–7.
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  1. Korn AP, Bolan G, Padian N, Ohm-Smith M, Schachter J, Landers DV. Plasma cell endometritis in women with symptomatic bacterial vaginosis. Obstet Gynecol. 1995;85(3):387–90.
  1. Matteo M, Cicinelli E, Greco P, Massenzio F, Baldini D, Falagario T, et al. Abnormal pattern of lymphocyte subpopulations in the endometrium of infertile women with chronic endometritis. Am J Reprod Immunol. 2009;61(5):322–9.
  1. Smith M, Hagerty KA, Skipper B, Bocklage T. Chronic endometritis: a combined histopathologic and clinical review of cases from 2002 to 2007. Int J Gynecol Pathol. 2010;29(1):44–50.
  1. Cicinelli E, De Ziegler D, Nicoletti R, Tinelli R, Saliani N, Resta L, et al. Poor reliability of vaginal and endocervical cultures for evaluating microbiology of endometrial cavity in women with chronic endometritis. Gynecol Obstet Invest. 2009;68(2):108–15.
  1. Küçük T, Safali M. ”Chromohysteroscopy” for evaluation of endometrium in recurrent in vitro fertilization failure. J Assist Reprod Genet. 2008;25(2-3):79–82.
  1. Park DW, Lee HJ, Park CW, Hong SR, Kwak-Kim J, Yang KM. Peripheral blood NK cells reflect changes in decidual NK cells in women with recurrent miscarriages. Am J Reprod Immunol. 2010;63(2):173–80.
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  1. Zhao M, Chang C, Liu Z, Chen LM, Chen Q. Treatment with low-dose aspirin increased the level LIF and integrin b3 expression in mice during the implantation window. Placenta. 2010;31(12):1101–5.
  1. Sher G, Fisch JD. Effect of vaginal sildenafil on the outcome of in vitro fertilization (IVF) after multiple IVF failures attributed to poor endometrial development Fertil Steril. 2002;78(5):1073–6.
  1. Ohl J, Lafebvre-Maunoury C, Wittemer C, Nisand G, Laurent MC, Hoffman P. Nitric oxide donors for patients undergoing IVF: A prospective, double blind, randomised, placebo controlled trial. Hum Reprod. 2002;17(10):2615–20.
  1. Battaglia C, Salvatori M, Maxia N, Petraglia F, Facchinetti F, Volpe A. Adjuvant L-arginine treatment for in vitro fertilisation in poor responder patients. Hum Reprod. 1999;14(7):1690–7.
  1. Sher J, Salazer C. Clinical experience with IVIg treatment in patients with failed IVF pregnancies:report of 30 consecutive patients. Am J Reprod Immunol. 2000;44(2):121–4.
  1. van den Heuvel MJ, Peralta CG, Hatta K, Han VK, Clark DA. Decline in number of elevated blood CD3(+) CD56(+) NKT cells in response to intravenous immunoglobulin treatment correlates with successful pregnancy. Am J Reprod Immunol. 2007;58(5):447–59.
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Luteal Phase Defectchapter 16

Surveen Ghumman,
Neerja Goel
Luteal phase defect has remained a disorder of controversy since its description as a clinical entity by Jones in 1949.1 Luteal phase defect is characterized by inadequate endometrial maturation due to a qualitative or quantitative disorder in corpus luteum function.
Progesterone secreted by the corpus luteum is essential for the initiation and maintenance of normal gestation. Luteal support remains essential till about the seventh week of gestation, by that time the trophoblast acquires sufficient steroidogenic capacity to support the pregnancy. When pregnancy occurs chorionic gonadotropins are responsible for the prolongation of corpus luteum function. Normal formation and function of the corpus luteum and optimal endometrial preparation is a prerequisite for both nidation and normal progress of early pregnancy. This is dependent on normal follicular and ovulatory phase endocrine events.
 
Pathophysiologic Mechanism
During folliculogenesis, there is a complex interplay between GnRH pulsatile patterns, FSH release and activity within the 261growing follicle, and peripheral steroid feedback. Disturbance in any of these factors leads to possible mechanisms for development of luteal phase defect (Fig. 16.1).
It was observed that there was a significantly low progesterone receptor content on endometrial glandular nucleus in luteal phase defect group. This resulted in a deficient endometrial response to progesterone stimulus. The result is a poorly prepared endometrium either due to inadequate progesterone receptor induction during the follicular phase or insufficient peripheral progesterone levels reaching the endometrium from the ovary leading to abnormal implantation or early pregnancy wastage.2
 
Why Luteal Support is Needed in ART Cycles?
  1. Supraphysiological estrogen levels seen in controlled ovarian hyperstimulation protocols may induce premature luteolysis.
    Fig. 16.1: Pathophysiology of luteal phase defect
    262
  2. Follicular phase downregulation may impair luteal phase luteinizing hormone release.
  3. Some protocols may give only pure FSH thus, leading to a relatively low LH value.
  4. Ovarian aspiration may cause disruption of granulosa cells leading to aberrant steroidogenesis.
  5. Controlled ovarian stimulation accelerates endometrial maturation hindering implantation.
The ability of the endometrium to respond to progesterone is an acquired property depending on the induction of adequate progesterone receptors by estradiol during the follicular phase of the cycle.
Hence, there is a concern in IVF/ICSI cycles of luteal phase defect and luteal phase is supported by progesterone, hCG, sometimes estradiol as a routine. Recently, single dose of GnRH agonist has also been tried.
 
Diagnosis of Luteal Phase Inadequacy
In general, a deficient luteal phase will be found in less than 10% of women who seek evaluation for infertility. It is mostly seen in:
  1. Hyperprolactinemia.3
  2. Elevated circulating androgens.
  3. Oligo-ovulation.
  4. Extremes of reproductive age.
  5. Treatment with ovulation inducing agents or ovarian suppressive agents.
  6. Patients with history of recurrent abortion.
  7. Endometriosis.4
  8. Following discontinuation of suppressive medical therapies.
  9. Strenuous exercise.
263
Diagnosis is based on endometrial histopathology, basal body temperature, low luteal progesterone levels and transvaginal sonography. There are other tests like decidual prolactin, steroid receptor studies and endometrial biochemical markers that can be done (Table 16.1).
 
Endometrial Biopsy
Two endometrial biopsies out of phase by 2 days obtained in 2 consecutive cycles are diagnostic of luteal phase defect. However, endometrial biopsy has a large interobserver error. Biopsy sites can also cause inconsistencies in findings.
 
Basal Body Temperature
It was seen that if a suspicious but ovulatory basal body temperature was present, a luteal phase defect was found in 80% cases on biopsy.
Table 16.1   Techniques for diagnosis of luteal phase defects
1.
Basal body temperature charts—monophasic or rise of temprature for less than 11 days
2.
Luteal phase progesterone levels—less than 10 ng/mL
3.
Transvaginal ultrasonography and Doppler studies
Sonographic evidence of aberrant luteolysis
  1. Persistent perifollicular reaction
  2. Rupture of follicle of <17 mm
  3. Poorly formed or ill-defined dominant follicle
  4. Luteinized unruptured follicle
  5. Lutein cyst formation
  6. Absence of corpus luteum
  7. Lack of endometrial echogenicity on 7th postovulation day
4.
Endometrial biopsy and histopathology—lag of 2 days
5.
Serum prolactin measurement
6.
Decidual prolactin measurement4
7.
Steroid hormone receptor analysis
8.
Biochemical markers for endometrial receptivity
264
If luteal phase is shorter than 11 days it correlates with a 6.25 days endometrial histological lag.5 Ovulation occurring on day 18 to 19 correlates with poor follicular progression, premature LH surge, equivocal progesterone levels and biopsy specimens with glands and stroma out of phase.
 
Low Progesterone Levels in Luteal Phase
More than 3 ng/mL of progesterone signifies ovulation. However, a level of more than 10 ng/mL is needed for adequate luteal support. Since there is pulsatatility of progesterone release these values cannot be depended upon.
 
Color and Pulsed Doppler Ultrasound
It is being used for diagnosis of luteal phase defect. Significantly lower resistance indices were seen in the uterine, arcuate, radial, and spiral arteries of the ovulatory group in the mid-luteal phase, which was inversely related to the progesterone level.6 Blood flow impedance in the corpus luteum and spiral arterioles is used to assess luteal adequacy. Sonographic criteria for aberrant luteolysis is shown in Table 16.1 (see Chapter 14).
 
Treatment of Luteal Phase Defect
Treatment is considered by allaying the factors responsible like hyperprolactinemia, uterine septum, etc and addition of progesterone (Fig. 16.2). The therapy is broadly categorized into two:
  1. Increasing progesterone levels
    1. Progesterone supplementation
    2. hCG265
    Fig. 16.2: Management of luteal phase defect
    266
  2. Improving folliculogenesis
    1. Clomiphene
    2. Human menopausal gonadotropins
 
Progesterone Supplementation
Progesterone or its derivative is the treatment of choice because of effective endometrial decidualization with no teratogenicity. Initiation of progesterone therapy after missing menses is not adequate, because the nidation site has not been properly prepared. Hence, in patients of luteal inadequacy progesterone supplementation should commence after ovulation so as to avoid early abortion. Progesterone supplementation is continued till 10 weeks of gestation as at this time placenta takes over the role of progesterone production.
 
Micronized Progesterone
Micronized progesterone can be given orally, vaginally or parenterally.
Oral Administration: Though progesterone is absorbed orally, more than 90% is metabolized during the first hepatic pass limiting its efficacy. Many micronized forms have become available to overcome this problem. Micronization in combination with lipophilic vehicles enhances absorption. Metabolites of orally administered progesterone may produce a hypnotic effect.7
Vaginal Administration: There is increased bioavailability and reduced variability when progesterone is given vaginally or rectally compared to oral route. This sustained level produces a more physiologic endometrial response. Micronized progesterone may exert a direct effect on the uterus by blocking the rejection of the embryo. It does not cause drowsiness or sleepiness but is inconvenient because of 267vaginal discharge. Patient is advised the use of progesterone vaginal suppositories after ovulation is confirmed. In a dose of 200 to 400 mg per day in two divided doses. This produces concentration similar to the luteal phase that is maximal within 1 to 8 hours and decrease over 24 hours. Vaginal gel also produces endometrial response as good or better than the intramuscular route.8 Pregnancy outcomes were comparable for progesterone replacement with vaginal gel and intramuscular progesterone in an oocyte donation program.9 A polysyloxane vaginal ring containing 1 gram of natural progesterone has been tried in IVF patients. It provides a continuous release of 10 to 20 nmol/L for 90 days.10 There has been increasing evidence of preferential drug distribution to the uterus after vaginal application. This is called the ‘first uterine pass effect’. The mechanism of uterine tropism of vaginal progesterone can be explained by various theories.11
  1. Passive diffusion through the tissues.
  2. Passage through the cervical canal.
  3. Absorption through venous or lymphatic systems.
  4. Countercurrent transport between fluids flowing in opposite directions.
Advantages of Vaginal Administration: Convenience and acceptability and a high tissue level making it unnecessary to monitor serum progesterone levels.
Intramuscular Administration: It is the most reliable route to achieve desired concentration of progesterone. It is rapidly absorbed and peak level is reached in 8 hours. Serum progesterone levels remain sustained compared to other routes as it is administered in an oil vehicle. It has the disadvantage of inconvenience of daily injections and pain or abscess formation at injection site. Allergic reactions may be seen. Intramuscular dose is 50 to 100 mg per day.268
 
Dehydrogesterone
It is a retroprogesterone or stereoisomer of progesterone. It is closest to native progesterone. It was found that pregnancy and implantation rate was similar whether progesterone or dehydrogesterone was given in the luteal phase following IVF and ET. It is given in a dose of 20 to 30 mg/day.12
 
Synthetic Progestational Agents
These should not be used to treat luteal phase inadequacy as they may have a luteolytic effect on the corpus luteum and can produce glandular stromal disparity, worsening the situation.13 Use of 19-nor-progestins is contraindicated in early pregnancy as it can masculinize a female fetus and may cause cardiovascular and limb defects.
 
Results
Supplementation has proven effective in correcting abnormal endometrial histology in more than 80% of these cases. Pregnancy rates in treated infertile patients with luteal phase inadequacy have ranged from 50 to 80%.14
 
Human Chorionic Gonadotropin (hCG)
Administration of hCG stimulates the corpus luteum to produce progesterone. It is ineffective in the presence of inadequate number of LH receptors or a malfunctioning corpus luteum, which is hyporesponsive to hCG. hCG is effective if there is a specific defect in postovulatory LH secretion or in trophoblastic hCG production. The steroidogenic response differs with timing of hCG. If given at LH surge there is no increase in E2 or progesterone concentration. 269Given on luteal day 5 it produces a marked steroidogenic response that remains throughout luteal phase. In mid-luteal phase on administration of hCG, progesterone levels reach early pregnancy levels but are not sustained. In late luteal phase progesterone response is shorter. This uncertainty of response, injectable mode of administration and difficulty in interpreting a positive pregnancy test prevent it from being the first choice of therapy. As hCG has a longer half-life than LH, it is advantageous in treating inadequate luteal function. In order to achieve complete luteinization of the preovulatory follicle, 10,000 IU of hCG should be administered at the time of ovulation followed by a dose of 2500 IU every 3 to 4 days. Treatment is stopped after the 12th postovulatory day to avoid a high incidence of psuedopregnancy and with the assumption that if pregnancy is achieved exogenous hCG should no longer be necessary. The long half-life of hCG renders pregnancy testing invalid for 7 days after the last hCG injection.
 
Estradiol Supplementation in Luteal Phase
Supplementation in the luteal phase with different doses of estradiol is being used in doses of 2 mg, 4 mg and 6 mg/day. In initial studies conducted significantly higher implantation rate and pregnancy rate were recorded in those who received low dose E2 supplementation compared with no substitution (PR 23.1% vs 32.8%). The best implantation and pregnancy results were found in the group with high dose E2 supplementation (PR 51.3%).15 However, more recent studies showed no difference in terms of number of oocytes retrieved, embryos transferred, pregnancy and implantation rates.16270
 
GnRH Agonist in the Luteal Phase
Single dose: The exact mechanism is still not known. It was suggested that GnRH agonist can help in the maintenance of the corpus luteum, acting directly on the endometrium via local receptors, a direct effect on the embryos or by some combination of these possibilities. A single dose of GnRH agonist (0.5 mg leuprolide acetate) was administered subcutaneously on day 6 after ICSI. A meta-analysis showed that the luteal-phase single-dose GnRH-agonist administration can increase implantation rate in all cycles and clinical pregnancy rate and ongoing pregnancy rate in cycles with GnRH antagonist ovarian stimulation protocol.17 GnRH agonist addition during the luteal phase significantly increases the probability of live birthrates.18
Multiple dose: 200 μg intranasal buserelin followed by 100 μg every day or alternate day up to day 14 of the luteal phase is given in multiple dose protocol. Intranasal administration of buserelin could be effective in triggering ovulation and in providing luteal support. This treatment was associated with a good pregnancy rate (28%) with IUI.19
 
How Long Should Luteal Phase Support Continue?
A study showed that after FSH/GnRH antagonist cycles, the withdrawal of progesterone supplementation in early pregnancy, with normally increasing β-hCG levels on the 16th day postembryo transfer, had no significant clinical impact in terms of ongoing pregnancy rates beyond 12 weeks. When comparing the groups where progesterone was continued till 7 weeks to that when it was stopped at 16th day, the ongoing pregnancy rate beyond 12 weeks was 82% versus 73%, abortion before or after 7 weeks of gestation 9% versus 12%, 271and 8% versus 10%, and bleeding episodes were14% versus 19%.20
 
Which is the Most Appropriate Luteal Support?
A recent Cochrane review 2011 showed a significant effect in favor of progesterone for luteal phase support. Overall, the addition of other substances such as estrogen or hCG did not seem to improve outcomes. No evidence favoring a specific route or duration of administration of progesterone was identified. hCG, or hCG with progesterone, was associated with a higher risk of OHSS and should therefore be avoided. There were significant results showing a benefit from addition of GnRH agonist to progesterone for the outcomes of live birth, clinical pregnancy and ongoing pregnancy. The review concluded that progesterone seems to be the best option as luteal phase support.21
 
Stimulation of Folliculogenesis
The ability of the endometrium to respond to progesterone is an acquired property depending on the induction of adequate progesterone receptors by estradiol during the follicular phase of the cycle. Patients who do not respond to progesterone supplementation may be having inadequate progesterone receptors. Therapy aimed at improving folliculogenesis may promote endometrial receptively, thus providing luteal phase adequacy.
 
Clomiphene Citrate
It has been seen that the functional capacity of the corpus luteum is dependent upon the normal growth and maturation of the preovulatory follicle. Emergence of dominant ovarian follicle, proliferation of granulosa layer, induction of LH 272receptors, endometrial proliferation, progesterone receptors induction and finally stimulation of the LH surge itself are all important prerequisites of the follicular phase for subsequent normal luteal function. Downs and Gibson have shown that the therapeutic efficacy of clomiphene appears directly related to the magnitude of the luteal defect.22
Despite reports of its efficacy in the treatment of luteal phase defect some authors regard such use of clomiphene as inappropriate due to the deleterious influence of the drug at the ovarian, endocervical and endometrial levels. On the endometrial level, clomiphene interferes with induction of endometrial progesterone receptors and limits the population of receptors ultimately available for a good endometrial secretory response. This predisposes to delay in endometrial maturation that could be recognized as a luteal phase defect. The higher luteal phase E2 levels that frequently occur in clomiphene treated cycles may also interfere with the process of endometrial decidualization. It has been seen that after repeated administration, a significant amount of clomiphene is retained in the circulation thus interfering with estrogen induced progesterone receptor replenishment during the luteal phase. Thus, the treatment that has been successfully initiated with clomiphene may require further treatment in the luteal phase as there is no guarantee of continued normal luteal function. Thus, it is clear that optimal use of clomiphene citrate requires careful titration to establish the dose that will produce the desired effects.
 
Human Menopausal Gonadotropins (hMG)
hMG can be used in treating luteal phase inadequacy by directly stimulating folliculogenesis. It is expensive and carries risks of multiple pregnancy and ovarian hyperstimulation. It is only used if previous trials with clomiphene and progesterone 273prove unsuccessful. However, Check et al treated infertile patients of luteal phase defect with ultra low dose of 75 IU and showed that this regimen was effective in correcting infertility related to luteal phase defect because of follicular maturation defects.23
Luteal phase defect is an important cause of infertility and recurrent pregnancy loss. Treatment of luteal phase inadequacy is aimed at correcting identifiable disorders (such as hyperprolactinemia or a uterine septum), stimulating folliculogenesis or corpus luteum function, or providing end product (progesterone) replacement. Natural progesterone is the drug of choice for supplementation with intramuscular or vaginal route being equally effective.
 
References
  1. Jones GES. Some newer aspects of the management of infertility. JAMA. 1949;141:1123–9.
  1. Jacobs MH, Balasch J, Gonzalez-Merlo JM, Vanrell JA, Wheeler C, Strauss JF 3rd, et al. Endometrial cytosolic and nuclear progesterone receptors in the luteal phase defect. J Clin Endocrinol Metab. 1987;64:472–5.
  1. Garzia E, Borgato S, Cozzi V, Doi P, Bulfamante G, Persani L, et al. Lack of expression of endometrial prolactin in early implantation failure: A pilot study. Hum Reprod. 2004;19(8):1911–6.
  1. Cunha-Filho JS, Gross JL, Bastos de Souza CA, Lemos NA, Giugliani C, Freitas F, et al. Physiopathological aspects of corpus luteum defect in infertile patients with mild/minimal endometriosis. J Assist Reprod Genet. 2003;20(3):117–21.
  1. Downs K, Gibson M. Basal body temperature graph and the luteal phase defect. Fertil Steril. 1983;40:466–8.
  1. Dal J, Vural B, Caliskan E, Ozkan S, Yucesoy I. Power Doppler ultrasound studies of ovarian, uterine, and endometrial blood flow in regularly menstruating women with respect to luteal phase defects. Fertil Steril. 2005;84(1):224–7.
  1. Arafat ES, Hargroove JT Maxson WS, Desiderio DM, Wentz AC, Anderson RN. Sedative and hypnotic effects of oral administration of micronized progesterone may be mediated through its metabolites. Am J Obstet Gynecol. 1988;159:1203–9.
  1. Gibson WE, Toner JP, Hamacher P, Kolm P. Experience with a novel vaginal progesterone preparation in a donor oocyte program. Fertil Steril. 1998;69:96–101.
  1. Berger BM, Phillips JA. Pregnancy outcomes in oocyte donation recipients: vaginal gel versus intramuscular injection progesterone replacement. J Assist Reprod Genet. 2012;29(3):237–42.
  1. Zegers-Hochschild F, Balmaceda JP, Fabres C, Alam V, Mackenna A, Fernandez E, et al. Prospective randomised trial to evaluate the efficiency of a vaginal ring releasing progesterone for IVF and oocyte donation. Hum Reprod. 2000;15(10):2093–7.
  1. Cicinelli E, Borraccino V, Petruzzi D, et al. Pharmacodynamics and endometrial effects of the vaginal administration of unmodified progesterone in an oil based solution to postmenopausal women. Fertil Steril. 1996;65:860–2.
  1. Ganesh A, Chakravorty N, Mukherjee R, Goswami S, Chaudhury K, Chakravarty B. Comparison of oral dydrogestrone with progesterone gel and micronized progesterone for luteal support in 1,373 women undergoing in vitro fertilization: a randomized clinical study. Fertil Steril. 2011;95(6):1961–5.
  1. Allenbach M, Hellnig G. The endometrium in natural and artificial luteal phase. Human Reprod. 1988;3:165–68.
  1. Wentz AC, Herbert CM, Maxon WS, Gernier CH. Outcome of progesterone treatment of luteal phase inadequacy. Fertil steril. 1984;41(6):856–62.
  1. Lukaszuk K, Liss J, Lukaszuk M, Maj B. Optimization of estradiol supplementation during the luteal phase improves the pregnancy rate in women undergoing in vitro fertilization-embryo transfer cycles. Fertil Steril. 2005;83(5):1372–6.
  1. Tonguc E, Var T, Ozyer S, Citil A, Dogan M. Estradiol supplementation during the luteal phase of in vitro fertilization cycles: a prospective randomised study. Eur J Obstet Gynecol Reprod Biol. 2011;154(2):172–6.
  1. Oliveira JB A, Baruffi R, Petersen CG, Mauri AL, Cavagna M, Franco JG Jr. Administration of single-dose GnRH agonist in the luteal phase in ICSI cycles: a meta-analysis. Reprod Biol Endocrinol. 2010;8:107.
  1. Kyrou D, Kolibianakis EM, Fatemi HM, Tarlatzi TB, Devroey P, Tarlatzis BC. Increased live birthrates with GnRH agonist addition for luteal support in ICSI/IVF cycles: a systematic review and meta-analysis. Hum Reprod Update. 2011;17(6):734–40.
  1. Pirard C, Donnez J, Loumaye E. GnRH agonist as novel luteal support: results of a randomized, parallel group, feasibility study using intranasal administration of buserelin. Hum Reprod. 2005;20(7):1798–804.
  1. Kyrou D, Fatemi HM, Zepiridis L, Riva A, Papanikolaou EG, Tarlatzis BC, Devroey P. Does cessation of progesterone supplementation during early pregnancy in patients treated with recFSH/GnRH antagonist affect ongoing pregnancy rates? A randomized controlled trial. Hum Reprod. 2011;26(5):1020–4.
  1. van der Linden M, Buckingham K, Farquhar C, Kremer JA, Metwally M. Luteal phase support for assisted reproduction cycles. Cochrane Database Syst Rev. 2011 Oct 5;(10):CD009154.
  1. Downs KA, Gibson M. Clomiphene citrate therapy for luteal phase defects. Fertil Steril. 1983;39(1):34–8.
  1. Check JH, Fine W. Similar pregnancy and abortion rates after treatment with low dose hMG versus pure FSH in women with luteal phase defect. Clin Exp Obstet Gynecol. 1997;24(1):5–7.

Complications of Ovulation Inductionchapter 17

Surveen Ghumman
With increasing progress in ovulation induction strategies the indication for their use have expanded. As the result of this we are dealing with an increasingly growing population facing the complications of these therapies. Adverse effects of ovarian stimulation may be divided into immediate and delayed effects.
Immediate Effects
  1. Drug specific side effects.
  2. Effects of ovarian overstimulation
    • Multiple pregnancy
    • Ovarian hyperstimulation syndrome.
Long-term Problems
  1. Risk of ovarian cancer.
The drug specific side effects have been dealt with in previous chapters. This chapter deals with ovarian hyperstimulation syndrome, multiple pregnancy and risk of ovarian cancer.277
 
Ovarian Hyperstimulation Syndrome
Ovarian hyperstimulation syndrome (OHSS) is one of the known complications of controlled ovarian stimulation. It is a syndrome with a wide spectrum of clinical and laboratory symptoms and sign, due to a fluid shift from the intravascular to the third space because of increased intravascular permeability, manifesting as ascites, pleural effusion, hemoconcentration, oliguria, electrolyte imbalance and hypercoagulability. It is accompanied by ovarian enlargement.
 
Incidence
OHSS is classified as mild, moderate and severe with the incidence ranging from 3 – 23% of inductions.1 The incidence varies with the ovarian stimulation protocols and the risk profile of the population being treated.
Mild OHSS
: 8–23%
Moderate OHSS
: 0.005–7%
Severe
: 0.005–2%
It occurred in 0.008 to 23% of hMG/hCG cycles and 0.6 to 14% in GnRH–a/hMG/hCG cycle.
 
Classification
It presents after hCG administration or rise of hCG due to an early pregnancy. It can be ‘early onset’, within 3–7 days of hCG administration, or ‘late onset’ 12 to 17 days after hCG because of early pregnancy.
  1. Golan's Classification: Golan proposed an acceptable classification with greater practical advantages (Table 17.1). It incorporates clinical signs, symptoms, ultrasonographic findings and laboratory findings to yield three stages and five grades of OHSS severity.2278
    Table 17.1   Golan's classification of OHSS2
    Grade
    Mild
    Moderate
    Severe
    1.
    Abdominal
    discomfort/distension
    2.
    Features of grade 1 along with nausea, vomiting and/or diarrhea. Ovaries enlarged 5–12 cm
    3.
    Features of mild OHSS and USG evidence of ascites
    4.
    Feature of moderate OHSS plus clinical evidence of ascites and/or hydrothorax with/or difficulty in breathing
    5.
    All of the above plus change in blood volume, increased blood viscosity due to hemoconcentration, coagulation disturbances and diminished renal perfusion and function
  2. Navot's Classification (Table 17.2): This classification further defined the severest degree of OHSS as given by Golan into severe and critical life-threatening stage based on a multitude of clinical and biochemical findings.3 In this classification generalized edema and liver dysfunction are considered additional signs of severe OHSS whereas adult respiratory distress syndrome, a tense ascitis, severe 279hemoconcentration (>55%) and profound leukocytosis (>25,000) are signs of the severest life-threatening form and need aggressive medical and surgical intervention.
Table 17.2   Clinical signs and laboratory criteria of ovarian hyperstimulation syndrome3
Mild to moderate
Severe
Critical
Ovarian enlargement
5–12 cm
>12 cm
Variable
Abdominal distension
Moderate
Severe
Tense
Clinical ascitis
None
Yes
Tense
Hydrothorax
None
Possible
Yes
Pericardial effusion
None
Infrequent
Infrequent
Decreased renal function
None
Infrequent
Frequent
Renal failure
None
None
Possible
Thromboembolism
None
None
Possible
ARDS
None
None
Possible
Hemoconcentration (hematocrit)
<45%
45–55%
>55%
WBC count
<15,000
15,000–25,000
>25,000
Liver enzymes
Normal
Elevated
Elevated
Creatinine (ng/mL)
<1.0
1.0–1.5
>1.6
Creatinine clearance (mL/min)
>100
50–100
<50
 
Pathophysiology of Ovarian Hyperstimulation Syndrome
The complicated pathophysiology of OHSS has still yet not been completely elucidated.280
Two major events are, however, recognized.
  1. Neovascularization: Neovascularization leads to increased vascularity.
  2. Increased vascular permeability of mesothelial surfaces: The increased capillary permeability of the ovarian vessels and other mesothelial surfaces leads to acute fluid shift to the third space (Fig. 17.1). This is triggered by release of vasoactive substances secreted by the ovary under the influence of hCG. These are prorenin and active renins, interleukins, nitric oxide, and vascular endothelial growth factor.
Fig. 17.1: Pathophysiology of OHSS
281
 
Clinical Features
The clinical features associated with OHSS are due to the shift of fluid into the third space by vascular permeability (Tables 17.1 and 17.2). The patient may have clinical features like:
  • Lower abdominal pain and distension.
  • Symptoms of nausea, vomiting and diarrhea.
  • Progressive lethargy
  • History of decreased urine output.
  • Increased pulse rate, shortness of breath, fluid collection at the base of lungs.
  • Significant fluid electrolyte imbalance.
  • Dehydration in severe cases.
  • Hypercoagulability of blood causing thrombosis.
 
Fatal Complications
  1. Vascular Complications: Venous compression due to enlarged ovaries and ascitis, immobility and a state of hypercoagubility causes deep vein thrombosis. Cerebrovascular complications subsequent to thromboembolic phenomenon may lead to hemiplegia and carotid artery embolism.
  2. Liver Dysfunction: The increased permeability in hepatic vasculature leads to edema, damage to hepatic cells and altered hepatic function. These changes may persist for 60 days.
  3. Respiratory Complications: Ascitis, pleural effusion and ARDS are due to fluid shift into the third space.
  4. Renal Complications: Prerenal failure occurs due to hypovolemia secondary to fluid transudate.
  5. Gastrointestinal Complications: Gastrointestinal symptoms may be the initial symptoms a patient presents with, and these help to diagnose the syndrome early.
  6. Adnexal Torsion: The enlarged ovaries can undergo torsion leading to an acute abdomen.
282
 
Management of OHSS
The most effective treatment of OHSS is precise prediction and active prevention. This can be done effectively with the combined use of ultrasonography and serum estradiol levels.
 
Prevention
 
Identify Patients who are at High Risk
This is the first step in prevention (Table 17.3). Monitoring of induction of ovulation is done to identify high-risk cases. A number of factors are related to increased risk of OHSS:
  1. Size and number of follicles: Women with a large number of follicles (>15), decreased fraction of large follicles and a high proportion of small and intermediate size follicles, are more prone to OHSS.
    Table 17.3   Risk factors for OHSS
    Predicting factors
    High risk
    Low risk
    Age
    Young (<35 years)
    Older (>36 years)
    Cause of anovulation
    Polycystic ovarian disease
    Hypogonadotropic hypogonadism
    Build
    Asthenic habitus
    Heavy build
    Number of follicles
    Multiple follicles (>35)
    Fewer follicles (<20)
    Ultrasonography of ovary
    “Necklace” sign present
    Absent
    Outcome of IVF cycle
    Pregnancy
    No pregnancy
    Luteal supplementation
    hCG luteal supplementation
    Progesterone/no supplementation
    Ovulation induction protocol
    GnRH agonist protocol
    GnRH antagonist protocol
    History of OHSS
    Present
    Absent
    283
  2. Serum estradiol: At serum estradiol levels of 4000 pg/mL or above hCG is withheld though studies have even quoted values above 3500 pg/mL.
  3. Age: Young patients are more prone to develop OHSS.
  4. Built: Thin patients are at higher risk of OHSS.
  5. PCOS: At the start of the cycle PCOS patients have a large number of small follicles that are all likely to respond to the dose of gonadotropins once the FSH threshold is reached. This would lead to hyperstimulation.
  6. OHSS in previous cycle: A history of OHSS in previous cycle, increases risk of recurrence in next cycle.
  7. Protocol of ovarian stimulation: GnRH agonist protocol has higher risk of OHSS compared with an antagonist protocol.
  8. Pregnancy: Patient who conceived, and more so with multiple pregnancy, were more prone to OHSS.
  9. Trigger for inducing follicular rupture: If hCG was used as a trigger for follicular rupture there were higher chances of OHSS.
  10. Luteal support: With hCG as a luteal support, the chances of development of OHSS were higher.
  11. Basal anti-Mullerian hormone (AMH): The basal serum AMH level predicted OHSS with a sensitivity of 90.5% and specificity of 81.3%.4
 
Withholding hCG
The criterion for withholding hCG varies in different centers. It is mostly based on more than one parameter like number and size of follicles, estradiol levels, slope of rise of estradiol, history of OHSS in previous cycle and presence of PCOS.
  1. Level of estradiol: hCG is withheld when estradiol levels are more than 3000 pg/mL. Incidence of severe OHSS was 1% if serum estradiol levels are 3000 – 3999 pg/mL and it increases to 5.97% if the levels are more 284than 4000 pg/mL. Hence, many prefer to take a cut-off value of 4000 pg/mL. However, many cases of OHSS can occur in normal estradiol levels and often, high estradiol levels may lead to no overstimulation.5
    Slope of rise of the plasma estradiol level: If values are more than doubling during 2–3 days (steep slope) then it should be regarded as a serious warning sign, and hCG should be withheld in that cycle.
  2. Number of follicles on ultrasonography: When there is an increase in fraction of the small and intermediate size follicles there was greater chance of OHSS developing. Presence of 15–20 follicles that are mainly immature (9 mm) should be taken as a cut-off. The final decision must see multiple parameters.
 
Delaying hCG (Coasting)
hCG can be delayed and a GnRH agonist or an antagonist can be started.
Agonist Coast: Withholding gonadotropins causes decreased FSH that causes downregulation of LH receptors reducing number of granulosa cell available for luteinization and a concomitant decrease in vasoactive substances causing OHSS. The optimum time to start coasting is when the lead follicle reaches 16 mm in diameter and estradiol levels are high. hCG is delayed, while GnRH agonist is continued witholding hMG till E2 falls below 3,000 pg/mL. At this level hCG is given. Longer period of coasting beyond 5 days, is associated with lower pregnancy rates.6 However, coasting for 1 or 2 days can be used successfully to prevent OHSS without compromising IVF cycle outcome.7
Antagonist Coast: When a patient undergoing an IVF cycle with long protocol is at high risk of severe OHSS, rescuing the cycle by withdrawing the agonist and replacing it with an antagonist and triggering ovulation with an agonist bolus 285could be considered without jeopardizing the safety of the patient while retaining the opportunity for success of the cycle.9 Administration of daily GnRH antagonist in high-risk patients for OHSS who were down-regulated by GnRHa resulted in rapid drop of E2 and decrease in incidence of OHSS.10
 
Decreasing Dose of hCG
Lower dosage may avoid hyperstimulation by exerting shorter periods of stimulation. 5,000 IU of hCG is given instead of 10,000 IU as an ovulation trigger.
 
Use of GnRH Agonist as a Trigger
Since period of stimulation is lesser with GnRH agonist surge, there is no hyperstimulation. The pregnancy rates are similar after an agonist or hCG trigger.11 However, some studies have reported a lower ongoing pregnancy rate after GnRHa trigger.12
The excellent conception rates reported in recipients receiving embryos originating from donor cycles or in women receiving frozen embryos originating from fresh cycles during which GnRHa was used to induce oocyte maturation suggest that it does not adversely affect the quality of the oocyte or embryo. A defective corpus luteum function resulting from the relatively short endogenous luteinizing hormone surge may be causing detrimental effects on endometrial receptivity. Aggressive luteal phase support and monitoring is, therefore, essential in view of the overwhelming evidence suggestive of abnormal luteal phase steroid profile. This may be achieved by the use of adequate estradiol and progesterone 286supplementation in the luteal phase and the first trimester.13 After modified luteal support there is now a non-significant difference of 6% in delivery rate in favor of hCG triggering.14
A recent Cochrane review (2011) recommends use on GnRH agonist as ovulation trigger in patients at high risk of OHSS.15
 
Luteal Phase Support
Progesterone, intravaginally or intramuscularly, is given for luteal support instead of hCG when patients are at high risk of OHSS.
 
Follicle Aspiration
Follicle aspiration was found to decrease the incidence of OHSS.16 Hence, if women are showing signs of being at risk of hyperstimulation, follicles should be aspirated.
 
Post-oocyte retrieval Albumin or Hydroxyethyl Starch Administration
Albumin helps by increasing serum oncotic pressure and reversing the leakage of fluid into the third space. Albumin has a half-life of 10–15 days, and needs timely administration at oocyte recovery, in a dose of 50 –100 g. The disadvantage is its oncotic action lasts for less than 36 hours, following which it moves into the interstitial compartment drawing fluid out of the intravascular space. A recent Cochrane review (2011) states there is limited benefit from intravenous albumin but better results with hydroxyethyl starch in preventing OHSS.17
 
Cryopreservation of Embryo and Subsequent Replacement
OHSS decreases by the tenth day if no pregnancy occurs, but continues for a longer time with viable pregnancy. 287Cryopreservation of embryos helps decrease chances of OHSS due to pregnancy. However, a recent Cochrane review did not support this.18
 
Steroids
Methyl prednisolone has been tried in cases of OHSS.19 However, most studies have not shown a protective effect.
 
Step-up Low Dose Regime of Gonadotropins
Low dose of gonadotropins are given in cases at high risk for hyperstimulation like PCOS and gradually stepped up.
 
Metformin
Addition of metformin to ovulation induction regimen in polycystic ovarian disease results in decreased incidence of OHSS (Cochrane review 2009).20
 
Dopamine Agonists
The dopamine receptor 2 agonists cabergoline and bromocriptine inactivate VEGF receptor-2 and prevent increased vascular permeability.
Carbagoline: It is given in a dose of 0.5 mg/d administered from the day of human chorionic gonadotropin for 8 days.21 It is considered a safe and effective medication.
Bromocriptine: Bromocriptine also decreased incidence of OHSS.22
Quinagolide: Quinagolide appears to prevent moderate/severe early OHSS while not affecting treatment outcome. A study showed an incidence of moderate/severe early OHSS of 23% in the placebo group and 12%, 13% and 4%in the quinagolide 50, 100 and 200 μg/day groups, respectively.23288
 
Avoidance of Excessive Gonadotropin Stimulation
The only reliable way to eliminate the risk of ovarian hyperstimulation syndrome (OHSS) is complete avoidance of gonadotropin ovarian stimulation
  1. Individualizing dose and low ovarian stimulation protocols: The CONSORT (CONsistency in rFSH Starting dOses for individualized tReatmenT) dosing algorithm individualizes recombinant human follicle-stimulating hormone doses for assisted reproduction technologies, assigning 37.5 IU increments according to easily available patient characteristics (basal follicle-stimulating hormone, body mass index, age, and antral follicle count) that have been proven to accurately predict ovarian response to ovarian stimulation.24
  2. Natural cycle ART: IUI or IVF without ovarian stimulation can lead to complete elimination of OHSS in high-risk cases.
  3. IVM: IVM is a treatment option in many centres. Although IVM may not replace standard IVF, it plays an increasingly important role in assisted reproductive technology, especially in the settings of high responders and those patients at risk of OHSS.25
 
Investigation and Monitoring of an OHSS Patient
  1. General Condition: General condition is monitored by regular charting of vital signs, weight charts, abdominal girth measurement and a strict fluid balance record (Table 17.4).
  2. Biochemical tests: A complete biochemical assessment includes hematocrit, electrolytes, liver function tests, kidney function tests and coagulation profile. Blood gases and acid-base balance is required if there is a respiratory 289or renal compromise.
    Table 17.4   Investigation and monitoring of OHSS patient
    1. General condition: It is monitored by regular charting of:
    1. Vital signs
    2. Weight charts
    3. Abdominal girth measurement
    4. Strict intake output chart
    2. Biochemical tests:
    1. Hematocrit
    2. Electrolytes
    3. Liver function tests
    4. Kidney function tests
    5. Coagulation profile
    6. Blood gases and acid-base balance
    7. Serum βHCG to rule out pregnancy
    3. Ultrasonographic examination: It is done to evaluate
    1. Ovarian size
    2. Amount of ascites
    3. Presence of hydrothorax
    4. Pregnancy, whether single or multiple
    Serum βhCG is done to rule out pregnancy. Serum and urinary osmolarity and urinary electrolytes may be needed in more severe forms of the disease. The frequency of these tests is guided by the severity of the disease.
  3. Ultrasonographic examination: Ultrasound gives important information on ovarian size, amount of ascites, presence of hydrothorax or pericardial effusion, and detection of pregnancy, whether single or multiple.
  4. Chest X-ray: A chest X-ray can rule out pleural effusion.
  5. Serun β-hCG: It is done to confirm pregnancy making the women at a high risk for developing severe disease.
  6. Invasive hemodynamic monitoring: When OHSS becomes critical monitoring of pulmonary artery pressure and central venous pressure may be required.
290
 
Treatment
The condition usually resolves within 10–14 days. Treatment is based on severity of the disease.
 
Mild OHSS
In mild cases the treatment is usually conservative and is done at outpatient level with close follow-up.
Grade I
  1. Reassure
  2. Plenty of fluids
  3. Avoid exertion
  4. Counsel on warning signs.
Grade II
  1. Serum electrolytes, hematocrit and ultrasonography should be done.
  2. Minimize physical activity and take plenty of fluids.
  3. Analgesics and antiemetics may be used if required.
  4. Intake output monitoring.
  5. Drug Therapy:
    1. Role of GnRH antagonists: If given on day 6 after oocyte retrieval in women with OHSS 4 days, combined with luteal phase support using exogenous estradiol and progesterone OHSS regressed.26 In women on antagonist regime, antagonist administration was re-initiated if OHSS developed and continued daily for a week, while all embryos were cryopreserved.
    2. Role of GnRH agonists. This resolved the OHSS. A marked decrease of hematocrit (Ht), WBC count, ovarian volume and ascitic fluid has been observed during 1 week of follow-up.27291
    3. Carbogoline: Carbogoline is given as 0.5 mg/day. It reduces hemoconcentration and ascites in hyperstimulated women undergoing assisted reproduction.28
Reassess if:
  1. Increase in weight more than 2 kg.
  2. Worsening of symptoms.
Indication of Hospitalization
Hospitalization should be considered in higher grades of the disease or if condition worsens and patient is not responding to treatment.
  1. In cases of grade II or III admission is required, if there is:
    1. Intolerable nausea and vomiting
    2. Hypotension
    3. Signs of pleural effusion
    4. Ascitis
    5. Hematocrit >48%
    6. Potassium level >5.0 mg/L
    7. Serum creatinine >1.2 mg
  2. All cases of grade IV and V should be hospitalized.
 
Severe OHSS
Aim of therapy after admission:
  1. Correction of circulatory volume electrolyte imbalance
  2. Maintenance of renal function
  3. Prevention of thrombosis.
  1. Maintenance of intravascular volume and electrolyte imbalance: The aim must be to restore normal intravascular volume and preserve adequate renal function. Colloid expander may be used for this purpose, but they have the disadvantage that after a short while they redistribute into the extravascular space worsening the ascitis. Low salt albumin is the expander of choice and is given in a dose 292of 50–100 gram every 2 to12 hours. It reverses hematocrit changes, improves renal function and is safe from viral contamination. Other options tried are mannitol, dextran and fresh frozen plasma. Dextran can cause ARDS. Only if there is hyponatremia, normal saline with or without glucose is the crystalloid used for replacement. Up to 1.5 to 3 liters may be needed. Other electrolyte imbalances like hyperkalemia are corrected.
  2. Prevention of thrombosis: Low dose heparin should be given, as prophylaxis, in cases where there is an altered coagulation profile.
  3. Diuretics: These drugs are usually not used but can be given after hemodilution is achieved if oliguria is persisting or in cases of pulmonary edema.
  4. Dopamine: Dopamine may help to avoid fluid and salt retention by improving the renal blood flow in oliguric patient.
  5. Management of ascitis: Paracentesis under ultrasound guidance is done where there is severe discomfort, compromise of venous return leading to a decreased cardiac output and hypotension, renal compromise, respiratory distress or hemoconcentration unresponsive to medical therapy. Repeat aspiration may be required.
  6. Paracentesis of hydrothorax: This should be done if dyspnea is present because of severe pleural effusion.
 
Critical OHSS
Critical OHSS causes multisystem failure and requires multidisciplinary intensive care.
  1. Renal failure: Dopamine central venous pressure line and hemodialysis may be required in severe cases.
  2. Pulmonary compromise: Arterial blood gas monitoring, thoracocentesis or assisted ventilation is required if they do not respond to basic treatment.293
  3. Thromboembolic events: Patients with thromboembolic episodes require therapeutic anticoagulation with heparin.
  4. Termination of pregnancy: If critical condition does not improve one may consider termination of pregnancy.
  5. Laparotomy: Laparotomy is required if the cysts undergo torsion, hemorrhage or rupture. Laparoscopic unwinding can be done in cases of torsion.
OHSS is an iatrogenic complication of controlled ovarian stimulation and may sometimes lead to life-threatening complications. Prevention is the best way to manage OHSS. Proper monitoring is essential and a balance between a conservative and aggressive approach is ideal to prevent unnecessary cycle cancelation.
 
Multiple Pregnancy
Multiple pregnancy may occur when ovulation induction is done with clomiphene, GnRH agonists and gonadotropins with an incidence of 5 to 10%, 7 to 10% and 16 to 40%, respectively. The greater relative increase in incidence is more with triplets and quadruplets (58%) compared to twins (18%).29 Multiple pregnancy causes increased incidence of preterm delivery, preeclampsia and abnormal bleeding. Cerebral palsy rates are 0.2% in singleton, 1.2% in twins and 4.5% in triplets. Besides this there may be a social burden on the family in bringing up twins. Fetal reduction is offered if there are triplets or more. Transvaginal sonography guided reduction is done at 8–9 weeks and transabdominally at 11 to 12 weeks with aspiration of the gestational sac or injection of cardiotoxic drug (KCl) into fetus. (see Chapter 13). The contribution of superovulation and ovulation induction to the multiple pregnancy epidemic is substantial.30 Strict guidelines should be followed so as to minimize these multiple births. Close monitoring is essential and presence of no more than 3 mature 294follicles should be there for administration of hCG. Transfer of more than 3 embryos in IVF cycle should be discouraged.
 
Ovarian Cancer and Ovulation Induction
Earlier studies suggested a three times increased risk of ovarian cancer in women who had used ovulation inducing drugs.31 There were reports of increased incidence of ovarian epithelial dysplasia in relation to intake of ovulation induction drugs in women who had later undergone hysterectomy and bilateral oophorectomy.32 It is thought that epithelial inclusion cysts formed at each ovulation are stimulated to undergo malignant transformation by gonadotropins that normally become elevated at menopause. This predicts that agents which provoke multiple ovulation by gonadotropin stimulation will increase the risk of ovarian cancer. An alternative theory is that gonadotropins may not be mutagenic but mitogenic (provoke a pre-existing tumor). It need not be a causal relationship but a stimulation of an already existing lesion. However, reports of large increases in ovarian cancer risk associated with fertility medications have not been replicated by more recent investigations.33 Some studies do report an increased incidence of borderline ovarian tumors.34 This was seen particularly with hMG. The association was not demonstrated with invasive tumors. No significant excess risk was associated with treatment with ovulation induction.35
It should also be kept in mind that cancers are over diagnosed in infertile women because of the close medical surveillance, which may also contribute to the early detection of cancers. However, it has been recommended that these drugs should not be used for more than 6 cycles consecutively and not more than a total of 12 cycles. It is therefore appropriate to use the smallest doses of ovarian stimulation for the shortest duration needed for clinical effectiveness.295
Ovulation induction should be individualized to prevent problems of OHSS and multiple pregnancy. High-risk women should be kept under close surveillance. Women should be counseled about these problems before starting treatment.
 
References
  1. Schenker IG, Weinsyein D. Ovarian overstimulation syndrome: a current survey. Fertil Steril. 1978;30:255–68.
  1. Golan A, Ron-elR, Herman A, Soffer Y, Weinraub Z, Caspi E. Ovarian hyperstimulation syndrome: an update review. Obstet Gynecol Survey. 1989;44:430–40.
  1. Navot D, Bergh PA, Lanfer N. Ovarian hyperstimulation syndrome in novel reproductive technologies: prevention and treatment. Fertil Steril. 1992;58:249–61.
  1. Lee TH, Liu CH, Huang CC, Wu YL, Shih YT, Ho HN, et al. Serum anti-mullerian hormone and estradiol levels as predictors of ovarian hyperstimulation syndrome in assisted reproduction technology cycles. Hum Reprod. 2008;23:160–7.
  1. Levy T, Orvieto R, Homberg R, Dekel A, Peleg D, Ben-Rafael Z. Severe hyperstimulation syndrome despite low plasma estrogen levels in hypogonadotropic hypogonadal patient. Hum Reprod. 1996;11:1177–9.
  1. Cheema P, Gelbaya TA, Horne G, Fitzgerald CT, Pease EH, Brison DR, et al. The optimal length of ‘coasting protocol’ in women at risk of ovarian hyperstimulation syndrome undergoing in vitro fertilization. Hum Fertil (Camb). 2006;9(3):175–80.
  1. Moon HS, Joo BS, Moon SE, Lee SK, Kim KS, Koo JS. Short coasting of 1 or 2 days by withholding both gonadotropins and gonadotropin-releasing hormone agonist prevents ovarian hyperstimulation syndrome without compromising the outcome. Fertil Steril. 2008;90(6):2172–8.
  1. D'Angelo A, Brown J, Amso NN. Coasting (withholding gonadotrophins) for preventing ovarian hyperstimulation syndrome. Cochrane Database Syst Rev. 2011;(6):CD002811.
  1. Martínez F, Rodríguez DB, Buxaderas R, Tur R, Mancini F, Coroleu B. GnRH antagonist rescue of a long-protocol IVF cycle and GnRH agonist trigger to avoid ovarian hyperstimulation syndrome: three case reports. Fertil Steril. 2011;95(7):2432.e17-9.
  1. Aboulghar M. Agonist and antagonist coast. Fertil Steril. 2012 Mar;97(3):523–6.
  1. Shapiro BS, Daneshmand ST, Garner FC, Aguirre M, Ross R. Comparison of human chorionic gonadotropin and gonadotropin-releasing hormone agonist for final oocyte maturation in oocyte donor cycles. Fertil Steril. 2007;88(1):237–9.
  1. Griesinger G, Diedrich K, Devroey P, Kolibianakis EM. GnRH agonist for triggering final oocyte maturation in the GnRH antagonist ovarian hyperstimulation protocol: a systematic review and meta-analysis. Hum Reprod Update. 2006;12(2):159–68.
  1. Engmann L, Benadiva C. Ovarian hyperstimulation syndrome prevention strategies: Luteal support strategies to optimize pregnancy success in cycles with gonadotropin-releasing hormone agonist ovulatory trigger. Semin Reprod Med. 2010;28(6):506–12.
  1. Humaidan P, Kol S, Papanikolaou EG. Copenhagen GnRH Agonist Triggering Workshop Group. Collaborators (14). GnRH agonist for triggering of final oocyte maturation: time for a change of practice? Hum Reprod Update. 2011;17(4):510–24.
  1. Youssef MA, Van der Veen F, Al-Inany HG, Griesinger G, Mochtar MH, Aboulfoutouh I, et al. Gonadotropin-releasing hormone agonist versus hCG for oocyte triggering in antagonist assisted reproductive technology cycles. Cochrane Database Syst Rev. 2011;(1):CD008046.
  1. Zhu WJ, Li XM, Chen XM, Zhang L. Follicular aspiration during the selection phase prevents severe ovarian hyperstimulation in patients with polycystic ovary syndrome who are undergoing in vitro fertilization. Eur J Obstet Gynecol Reprod Biol. 2005;122(1):79–84.
  1. Youssef MA, Al-Inany HG, Evers JL, Aboulghar M. Intravenous fluids for the prevention of severe ovarian hyperstimulation syndrome. Cochrane Database Syst Rev. 2011;(2):CD001302.
  1. D'Angelo A, Amso N. Embryo freezing for preventing ovarian hyperstimulation syndrome. Cochrane Database Syst Rev. 2007;(3):CD002806.
  1. Lainas T, Petsas G, Stavropoulou G, Alexopoulou E, lliadis G, Minaretzis D. Administration of methylprednisolone to prevent severe ovarian hyperstimulation syndrome in patients undergoing in vitro fertilization. Fertil Steril. 2002;78:529–33.
  1. Tso LO, Costello MF, Albuquerque LE, Andriolo RB, Freitas V. Metformin treatment before and during IVF or ICSI in women with polycystic ovary syndrome. Cochrane Database Syst Rev. 2009;(2):CD006105.
  1. Youssef MA, van Wely M, Hassan MA, Al-Inany HG, Mochtar M, Khattab S, et al. Can dopamine agonists reduce the incidence and severity of OHSS in IVF/ICSI treatment cycles? A systematic review and meta-analysis. Hum Reprod Update. 2010;16(5):459–66.
  1. Sherwal V, Malik S, Bhatia V. Effect of bromocriptine on the severity of ovarian hyperstimulation syndrome and outcome in high responders undergoing assisted reproduction. J Hum Reprod Sci. 2010;3(2):85–90.
  1. Busso C, Fernández-Sánchez M, García-Velasco JA, Landeras J, Ballesteros A, Muáoz E, et al. The non-ergot derived dopamine agonist quinagolide in prevention of early ovarian hyperstimulation syndrome in IVF patients: a randomized, double-blind, placebo-controlled trial. Hum Reprod. 2010;25(4):995–1004.
  1. Olivennes F. Ovarian hyperstimulation syndrome prevention strategies: individualizing gonadotropin dose. Semin Reprod Med. 2010;28(6):463–7.
  1. Huang JY, Chian RC, Tan SL. Ovarian hyperstimulation syndrome prevention strategies: in vitro maturation. Semin Reprod Med. 2010;28(6):519–31.
  1. Lainas TG, Sfontouris IA, Zorzovilis IZ, Petsas GK, Lainas GT, Alexopoulou E, et al. Live births after management of severe OHSS by GnRH antagonist administration in the luteal phase. Reprod Biomed Online. 2009;19(6):789–95.
  1. Lainas TG, Sfontouris IA, Zorzovilis IZ, Petsas GK, Lainas GT, Kolibianakis EM. Management of severe early ovarian hyperstimulation syndrome by re-initiation of GnRH antagonist. Reprod Biomed Online. 2007;15(4):408–12.
  1. Alvarez C, Martí-Bonmatí L, Novella-Maestre E, Sanz R, Gómez R, Fernández-Sánchez M, et al. Dopamine agonist cabergoline reduces hemoconcentration and ascites in hyperstimulated women undergoing assisted reproduction. J Clin Endocrinol Metab. 2007;92(8):2931–7.
  1. Hecht BR. The impact of assisted reproductive technology on incidence of multiple gestation. In Keith LG, Papiernik E'Keith DM, Luke B (Eds). Multiple Pregnancy London: Parthenon.  1995;175–90.
  1. Legro RS. Superovulation and multiple birth: in search of kryptonite. Fertil Steril. 2012;97(4):793–4.
  1. Whittemore AS, Harris R, Itnyre J, Halpern J. Characteristics related to ovarian cancer risk: collaborative analysis of 12 US case controlled studies. I Methods. Collaborative Ovarian Cancer Group. Am J Epidemiol. 1992;136:1175–83.
  1. Nieto JJ, Crow J, Sundaresan M, Constantinovici N, Perrett CW, MacLean AB, et al. Ovarian epithelial dysplasia in relation to ovulation induction and nulliparity. Gynecol Oncol. 2001;82(2):344–9.
  1. Brinton LA, Moghissi KS, Scoccia B, Westhoff CL, Lamb EJ. Ovulation induction and cancer risk. Fertil Steril. 2005;83(2):261–74.
  1. Ayhan A, Salman MC, Celik H, Dursun P, Ozyuncu O, Gultekin M. Association between fertility drugs and gynaecologic cancers, breast cancer, and childhood cancers. Acta Obstet Gynecol Scand. 2004;83(12):1104–11.
  1. Calderon-Margalit R, Friedlander Y, Yanetz R, Kleinhaus K, Perrin MC, Manor O, et al. Cancer risk after exposure to treatments for ovulation induction. Am J Epidemiol. 2009;169(3):365–75.

Selective Multifetal Pregnancy Reductionchapter 18

Shweta Mittal,
Deepak Chawla,
Abha Majumdar
The incidence of multifetal pregnancies has increased dramatically over the past two decades, mainly because of the widespread use of ovulation induction agents and assisted reproduction techniques.1 These techniques have been a matter of concern since twin and higher order pregnancies have long been associated with an increased risk of maternal complications as well as a high prevalence of perinatal and neonatal morbidity and mortality.
The most common complication is preterm delivery, with twins having an average gestational age at delivery of 36 to 37 weeks; triplets, 34 weeks; quadruplets, 29 to 31 weeks, and quintuplets, even earlier. In addition, researchers have shown an increased incidence of low birth weight, gestational diabetes mellitus, pregnancy-induced hypertension and greater requirement of neonatal hospital admission with multifetal pregnancy.
A couple has several options when faced with a multifetal pregnancy.300
  1. They can electively terminate the multifetal pregnancy with the intent to conceive again. Since the pregnancy is most likely wanted, achieved at great psychological and economic cost and with no guarantee of future conceptions, this option is usually the least desirable.
  2. The couple can attempt to proceed with the pregnancy. Even though there are reports of survival of some or all quadruplets and quintuplets, there is still significant risk of long-term morbidity. Survival with six or seven fetuses, although reported, is extremely rare. There are no reports of any fetal survivals with eight or more fetuses.
  3. The couple can choose multifetal pregnancy reduction. Selective fetal reduction in triplets is still controversial.
The procedure of multifetal pregnancy reduction (MFPR) has, in recent years, become both clinically and ethically accepted as a therapeutic option in pregnancies with four or more fetuses, and in multifetal pregnancies in which one or more of the fetuses has congenital abnormalities.2 MFPR results in better pregnancy outcome, regardless of the initial number of fetuses.3 In a study of IVF-conceived triplets, selective reduction of the pair to a singleton pregnancy was associated with a significantly greater likelihood of delivery at ≥34 weeks. On average, reduction of the pair was associated with 52 days longer gestation.4 The pregnancy loss subsequent to fetal reduction has been reported as ranging from 0 to 40%.
 
Methods of Multifetal Pregnancy Reduction5
  • Transcervical aspiration of the gestational sac
  • Transvaginal puncture and embryo aspiration
  • Intrathoracic injection of potassium chloride, by both transabdominal and transvaginal approaches.301
 
Transcervical Aspiration
Some authors have used transcervical aspiration of the gestational sac. This method, however, was thought to be associated with an increased incidence of fetal loss due to infection caused by introduction of bacteria from the cervix, or due to cervical incompetence brought about by cervical dilatation.
 
Transvaginal Puncture and Embryo Aspiration
Fetal reduction very early in gestation (6 to 8 weeks) by the transvaginal puncture and embryo aspiration has also been reported with fairly good pregnancy outcome.6 However, this method might have some theoretical limitations, such as:
  1. Use of general anesthesia.
  2. Possibility of spontaneous fetal reduction at this stage of gestation.
  3. Inability to perform early fetal screening, such as nuchal translucency test which is done at 10 to 12 weeks of gestation.
  4. Possibility of introducing infections.
 
Intrathoracic Injection of Potassium Chloride by Transabdominal and Transvaginal Approach
Multifetal pregnancy reduction using intrathoracic injection of potassium chloride, by both the transabdominal and the transvaginal approaches, has been reported. No method has yet been proven to be superior to the others.
Although several techniques of multifetal pregnancy reduction have been reported, the most popular is however the intrathoracic injection of potassium chloride by the transabdominal approach at 10 to 12 weeks gestation. It is logical to perform a detailed ultrasonographic fetal anomaly 302scan prior to the reduction (Fig. 18.1). This will allow the reduction to be performed more selectively and will decrease the chance of delivery of a chromosomal or structurally abnormal fetus.
 
Intracranial Injection of Potassium Chloride
In certain cases of MFPR, where difficulty is encountered in reaching the thorax due to the fetal position as well as the location of membranes and placenta, an alternative approach may be the insertion of the needle to the fetal cranium. This approach enables a technically easier procedure than the intrathoracic approach. However, the use of this technique should be reserved for selected cases of MFPR only by experienced operators and centers.7
 
Pre-procedural Preparation
  1. Counseling of the couple regarding the procedure and its possible complications.
  2. Informed written consent.
  3. Prophylactic antibiotic administration.
  4. Patient may be admitted for a day in the hospital.
Fig. 18.1: USG showing triplets
303
 
Transvaginal Procedure of Fetal Reduction8
This procedure is done between 8 and 9 weeks of gestational age under general anesthesia. Strict aseptic conditions should be maintained throughout the procedure. Patient is placed in dorsal lithotomy position. Cleaning of vagina is done with povidone iodine solution. Needle guide is attached to the transvaginal probe. Begin with transvaginal ultrasound examination of all the fetuses. Choose the correct path of the needle avoiding the path of the blood vessels. A 35 cm 18 gauge needle with a stylet is introduced through a guide and advanced through the vaginal wall, uterine wall into the fetal sac. The stylet is removed and a 21 gauge needle 40 cm long is introduced into the fetal thorax. 2 ml of 2 mEq of potassium chloride is injected. Fetal asystole is observed and needle is removed.
 
Transabdominal Procedure of Fetal Reduction5
This procedure is performed between 10 and 12 weeks of gestational age, under local anesthesia. Prior to the procedure ultrasound examination of all the fetuses is performed (Figs 18.2 and 18.3; See accompanying interactive CD-ROM).
Fig. 18.2: Ultrasound machine
304
Fig. 18.3: Ultrasound showing triplets before the procedure
Abdomen is prepared with povidone iodine solution. Fetus nearest to the ultrasound probe is selected (Fig. 18.4).
Spinal needle no. 21 with stylet is advanced through the abdominal and uterine wall into the fetal sac. Stylet is removed (Fig. 18.5).
Syringe is loaded with 2 mL of 2 mEq potassium chloride (Fig. 18.6).
Fig. 18.4: Selection of fetus nearest the ultrasound probe
305
Fig. 18.5: Insertion of needle
Fig. 18.6: Loading of syringe
The needle is visualizing on ultrasound and advanced into the fetal thorax (Fig. 18.7).
After the needle is advanced in the fetal thorax potassium chloride is injected. Needle is removed after confirming fetal cardiac asystole (Fig. 18.8). Cardiac activity of other fetus is confirmed.
Post-procedural second look ultrasound is done after few hours and another scan a few days later.306
Fig. 18.7: Visualization of needle tip
Fig. 18.8: Injection of intrathoracic potassium chloride
 
Complications
  1. Leaking per vaginum.
  2. Bleeding per vaginum.
  3. Abortion or loss of remaining fetuses.
  4. Infection.
307
 
Advantages of Transvaginal Procedure
Feasibility of the procedure at an earlier gestational age. However, the physician should be familiar with the procedure before applying it for routine use.
 
Advantages of Transabdominal Route
  1. A more detailed USG of the fetuses can be performed and nuchal thickness can be assessd as it is measured between 10 and 12 week gestational age.
  2. Chances of spontaneous reduction of multifetal pregnancy is ruled out.
  3. Lower risk of infection.
No decision in a high-order-multiple pregnancy is easy, and parents may understandably review their choices for years afterward, wondering if they should have chosen differently.
 
References
  1. Gonen R, Heyman E, Asztalos EV, Ohlsson A, Pitson LC, Shennan AT, et al. The outcome of triplet, quadruplet and quintuplet pregnancies managed in a perinatal unit: obstetric neonatal and follow-up data. Am J Obstet Gynecol. 1990;162:454–9.
  1. Berkowitz RL, Lynch, L, Chitkara U, Wilkins IA, Mehalek KE, Alvarez E. Selective reduction of multifetal pregnancies in the first trimester. N Engl J Med. 1988;318:1043–7.
  1. Antsaklis A, Anastasakis E. Selective reduction in twins and multiple pregnancies. J Perinat Med. 2011;39(1):15–21.
  1. Skiadas CC, Missmer SA, Benson CB, Acker D, Racowsky C. Impact of selective reduction of the monochorionic pair in in vitro fertilization triplet pregnancies on gestational length. Fertil Steril. 2010;94(7):2930–1.
  1. Wapner RJ, Davis GH, Johnson A, Weinblatt VJ, Fischer RL, Jackson LG, et al. Selective reduction of multifetal pregnancies. Lancet. 1990;335:90–3.
  1. Mansour RT, Aboulghar MA, Serour GI, Sattar MA, Kamal A, Amin YM. Multifetal pregnancy reduction: modification of the technique and analysis of the outcome. Fertil Steril. 1999;71(2):380–4.
  1. Lembet A, Selam B, Bodur H, Ergin T, Demirel C. Intracranial injection with KCl: an alternative method in selected cases of multifetal pregnancy reduction. Fetal Diagn Ther. 2009;26(3):134–6.
  1. Shalev J, Frenkel Y, Goldenberg M, Shalev E, Lipitz S, Barkai G, et al. Selective reduction in multiple gestations: pregnancy outcome after transvaginal and transabdominal needle-guided procedures. Fertil Steril. 1989;52:416–20.

Ultrasonography and Color Doppler Imaging in Ovulation Inductionchapter 19

Reeti Sahani
Reproductive organs of a woman during her fertile years shows daily changes which are very diverse. They can easily be viewed and assessed with modern imaging techniques such as sonography and color Doppler imaging.
 
Examination Technique
Sonographic examination of the female pelvic organs is the most commonly performed using the following approaches:
  1. Transabdominal (TAS).
  2. Transvaginal (TVS).
  3. Transperineal (less frequently).
A thorough ultrasound examination of the pelvis should include both complete transabdominal and transvaginal studies. The techniques are complementary, not mutually exclusive unless limited information is needed (e.g. follicle size) or extenuating circumstances dictate otherwise (e.g. patient refusal).310
 
Ovaries
The ovaries are generally situated on either side of the uterus. A search along the internal iliac artery would most often find the ovary located anterior to the vascular bifurcation into anterior and posterior branches. The blood supply is from the ovarian artery and branches of uterine artery (Figs 19.1A and B).
Fig. 19.1A and B: Ovarian artery
311
 
Uterus
Uterus receives its supply via the uterine artery, a branch of the internal iliac artery. From the uterine artery arise perforating branches, which extend through the serosa. Endometrium in midcycle has a triple layered appearance (Figs 19.2A and B). It derives its supply from arcuate branches of the uterine arteries. Radial arteries, branch of arcuate arteries, extend through the myometrium to just outside the endometrium (Fig. 19.3) where they form terminal branches of two types: straight and coiled. The straight branches (basal arteries) supply the basalis layer of the endometrium. The coiled branches (spiral arteries) traverse the endometrium and supply the functionalis layer.
Fig. 19.2A and B: Typical triple layer endometrium
312
Fig. 19.3: Endometrial vascularity
 
Color Doppler Imaging (CDI)
 
Purpose of CDI
  • Identify cyclical endometrial and follicular neovascularity followed by regression in neovascularity.
  • Determine changes in vascularity.
  • Quantify blood flow.
 
Indices used for Quantifying in CDI
  • Resistive index.
  • Pulsatility index.
  • SD ratio.
 
CDI in Ovarian Physiology and Cycle Changes
The ovarian arterial supply exhibits different flow characteristics during the different phases of a normal menstrual cycle (Fig. 19.4). These phases are:
  • Early follicular phase index values of arteries are relatively high (days 5–7).313
    Fig. 19.4: Resistance ovarian waveform pattern
  • Late follicular phase index values (days 11–13) are high.
  • Early luteal phase index values (days 15–17) are low.
  • Late luteal phase index values (days 26–28) rise.
The variations are believed to be hormone related and reflect changes in vascular compliance.1
Ovarian artery blood flow is detectable when the dominant follicle reaches a size of 12 to 15 mm. The resistance index (RI) is 0.54 ± 0.04 and declines the day before ovulation (Fig. 19.5). Moderate RI value of 0.55 and increased flow velocity in subendometrial vessels indicate favorable uterine receptivity (Fig. 19.6).
Fig. 19.5: Low resistance pattern
314
Fig. 19.6: Blood flow velocity waveforms of the sub-endometrial vessels on the day of ET
During ovarian stimulation, the waveform and index value differences normally noted between the two ovaries may be absent. Bilaterally, the ovarian arterial blood supply may demonstrate pulsatility waveforms typical of low impedance.
During the stimulation process, ultrasound has its greatest contribution in monitoring follicular development and guiding the oocyte harvesting procedure.
Gray-scale evaluation of ovarian follicles can help distinguish physiologic from insufficient or abnormal cycles according to the growth of the follicle. Transvaginal color Doppler can be employed to assess the physiologic development of the follicles through depiction of flow parameters2 (Figs 19.7 to 19.9). Four grades of perifollicular flow on color Doppler are seen (Table 19.1).
  1. Normal blood flow surrounding a corpus luteum around entire periphery. In real-time imaging, virtually the entire corpus luteum displayed color flow (Fig. 19.7A).
  2. Normal blood flow surrounding a corpus luteum (90%) (Fig. 19.7B).
  3. Moderate 50% perifollicular flow (Fig. 19.7C).
  4. More than 75% perifollicular flow (Fig. 19.7D).315
:
316
Fig. 19.7A to D: Corpus luteal flow
Fig. 19.8: Normal robust flow
Optimal stromal artery flow in the ovary has also been assessed. The peak systolic velocity should be more than 10 cm/sec for a good pregnancy rate (Table 19.2).
3D ultrasound is much more accurate for volume assessment of the follicle. Presence of cumulus increases the surety of the presence of a mature ovum in the follicle.317
Fig. 19.9: Abnormal anemic flow
Table 19.1   Optimal perifollicular flow
CDI/power Doppler to assess follicle circumference vascularization
1.
Grade 1
< 25%
2.
Grade 2
25–50%
3.
Grade 3
50–75%
4.
Grade 4
> 75%
Table 19.2   Optimal stromal artery flow
• Peak systolic velocity > 10 cm/sec
• Pulsatility index—is not indicative
• Resistive index is not indicative except for hyperstimulation (RI < 0.48)
3D US and 3D PD when used with 2D US and color Doppler for pre-hCG follicular assessment would definitely improve pregnancy rates in IUI cycles.3318
 
CDI in Uterine Physiology
The uterine vessels examined during the cycle are the uterine artery, spiral artery and vessels at the endomyometrial junction to note the following:4
  • Proliferation of spiral arteries
  • Growth of spiral arteries toward the endometrium
  • Increased vascularity in the endometrium
  • Increased flow in the main uterine arteries.
 
Uterine Artery
The general pattern of uterine blood flow throughout the menstrual cycle is that perfusion increases in response to rising plasma estrogen and progesterone and decreases with the periovulatory fall in estrogen.5 The lowest pulsatility index (PI) values are seen around days 8 and 21, while the highest values are seen around days 1, 14 and 17. Significant changes in diastolic blood flow at the different times of the cycle may not be noted. In general; the index values for the uterine artery ipsilateral to the ovary containing the dominant follicle are lower than the contralateral artery (Figs 19.10 and 19.11).
Other patterns of uterine artery blood flow have been described. When the uterine arteries were interrogated at the level of the uterine cornua, the PI reached its peak by day 11 and remained relatively constant until day 16. The lowest values were generally seen around days 1 and 21. At this anatomic level, end-diastolic flow was commonly absent during the early follicular phase (Fig. 19.12) but it was demonstrable by the luteal phase.
The cyclical changes reflected by the flow velocity waveforms and index values appear to be mediated by the reproductive hormones. The baseline evaluation (pre-treatment) demonstrated a narrow systolic spectral flow pattern with a mean PI of 5.2 ± 0.4. Evaluations performed 319on days 13–14, showed a spectral tracing that was broader with an uninterrupted diastolic component.
Fig. 19.10: Doppler waveform of uterine artery ipsilateral to dominant follicle ovary
Fig. 19.11: Doppler waveform of uterine artery contralateral to dominant follicle ovary
320
Fig. 19.12: Near absent end-diastolic flow during follicular phase
The mean PI was 1.5 ± 0.2. On days 26 to 27, no significant differences were noted (mean PI = 1.7 ± 0.3).6,7 Uterine artery RI is given a score of 0–48 (Table 19.3).
 
Endometrium
Endometrial morphology: In preparation for implantation, the endometrium undergoes transformations by increased blood flow and uterine oxygen consumption.
Table 19.3   Optimal uterine artery RI score
RI
Score
< 0.7
4
0.7–0.8
2
> 0.8
0
321
The cells in the stroma and epithelium increase and there is a generalized edema.
The endometrium and periendometrial area is divided into 4 zones (Table 19.4). These zones give it the typical preovulatory triple line appearance that is an indicator of good uterine receptivity (Fig. 19.13).
Endometrial vascularity: Endometrial zonal neovascularity is of prime importance for embryo transfer and is determined at the following levels:
  • Subendometrial
  • Basal
  • Mid zone
  • Inner layer.
The spiral arteries, like the endometrium, are remarkably responsive to the hormonal changes occurring in the menstrual cycle. These include:9
Table 19.4   The endometrial and periendometrial areas have the following four zones
• Zone 1
2 mm thick area surrounding the hyperechoic outer layer of the endometrium
• Zone 2
The hyperechoic outer layer of the endometrium
• Zone 3
The hypoechoic inner layer of the endometrium
• Zone 4
The endometrial cavity
Fig. 19.13: Triple layer endometrial thickness
322
  • Endothelial proliferation
  • Wall thickening and coiling.
These vessels play an important role in implantation. The chances for a normal implantation may be reduced if the spiral arterioles are inadequately developed.
It is possible to see variations in the depth of vascular penetration before, during and after the mid-cycle. In patients with uterine artery PIs of more than 3.0, preliminary results have not revealed any successful pregnancies in IVF patients unless there is vascularity demonstrated either within zone 3 or within zones 3 and 4 prior to transfer10 (Figs 19.14A and B, and 19.15).
Fig. 19.14A and B: Normal zone 3 blood flow is demonstrated on this endovaginal image
323
Fig. 19.15: Vascular penetration on power Doppler to zone 4
An optimal score based on imaging vascularity taking into consideration number of vessels and endometrial power Doppler area, is used for assessment (Table 19.5 and Fig. 19.16).
The color Doppler findings in unsuccessful cycles may relate to the histologic findings. A majority demonstrated an immature endometrium at the time of embryo transfer. The abnormalities included a variety of patterns, all indicating a lack of secretory transformation, suggesting poor endometrial receptivity for implantation.11
Endometrial thickness (ET): Thickness of endometrium is also important for implantation. Less than 7 mm gives a poor pregnancy rate (Table 19.6).
Table 19.5   Optimal endometrial score and vascularity quantification
Characteristic
Scoring
• Number of vessels
   3 and above
Score of 3
   1–3
Score of 2
   Absent
Score of 0
• Endometrial power
Score of 4
   Doppler area of > 5 mm sq
• PSV, RI and PI not reliable
324
Fig. 19.16: Imaging in optimal endometrial score
Table 19.6   Optimal endometrial score
ET
Score
• < 7 mm
0
• 10–11 mm
2
• 12 mm and above
3
In cycles resulting in pregnancy, mean endometrial thickness was higher compared to cycles with negative outcomes. Higher serum estradiol is associated with higher endometrial thickness and pregnancy rates. Women achieving pregnancy and pregnant women with endometrium thicker than 9 mm were younger. Follicle stimulation was better with higher endometrial thickness. After adjustments for age, no statistical difference was found in endometrial thickness between agonist and antagonist protocols.12
Endometrial volume on 3D ultrasound: With the 3D ultrasound being used to assess endometrial receptivity, the volume estimation of endometrium is done. The endometrial volume was measured by area tracing from the fundus to the internal cervical os in a number of parallel slices 1 to 2 mm apart. An ideal volume is 2–7 mL (Fig. 19.17).13325
Fig. 19.17: Endometrial volume on 3D ultrasound
 
Ultrasonographic Scoring of Uterine Receptivity
A scoring system for endometrial receptivity has been done taking into account endometrial thickness, pattern and vascularity. It also includes the appearance of myometrium (Table 19.7).
Another score was devised for patients undergoing IVF to assess the uterine receptivity (Table 19.8). The maximum score was 7 and the cut-off value was 5.
Table 19.7   Scoring system for uterine receptibity
Uterus
Nature
Score
Nature
Score
ET
> 7
4
< 7
0
ET pattern
Triple layer
3
Others
0
Myometrium
Homogenous
2
0
Uterine artery
PI < 3
4
PI > 3
0
End-diastolic flow
Present
4
Absent
0
Endometrial
vascularity
Present
3
Absent
0
Total score
20
0
326
Table 19.8   Two-dimensional ultrasonographic scoring system for evaluation of uterine receptivity in patients undergoing IVF-ET procedures11
Characteristic
Score
Endometrial thickness, mm
6–8
0
9–14
2
>15
1
Endometrial morphology
Triple line
0
Homogenous hyperechoic
2
Hypoechogenic with hyperechogenic borders
1
Sub-endometrial blood flow (RI)
Absent
0
>0.55
1
=0.55
3
Maximum score*
7
*The cut-off value was 5.
Scoring on evaluation of 3D ultrasonography of endometrium included endometrial volume, vascularity and morphology. The maximum score was 7 with a cut-off score of 5 (Table 19.9).
 
Comparison of 3D and 2D Ultrasonography for Assessment of Endometrial Receptivity
On comparing the sensitivity, specificity, positive predictive value, negative predictive value and efficiency of the 2D and 3D ultrasound assessment of endometrial receptivity no significant difference was found (Table 19.10).327
Table 19.9   Three-dimensional ultrasonographic scoring system for evaluation of uterine receptivity in patients undergoing IVF-ET procedures11
Characteristic
Score
Endometrial volume, mL
<2
0
2–7
2
>7
1
Endometrial morphology
Triple line
0
Homogenous hyperechogenic
2
Hypoechogenic with hyperechogenic borders
1
Sub-endometrial blood flow (FI)
<11
0
11–15
3
>15
1
Maximum score*
7
*The cut-off value was 5
Table 19.10   Comparison between 2D and 3D ultrasonographic scoring systems in the assessment of endometrial receptivity in patients undergoing IVF-ET procedures11
Ultrasonographic
Sensitivity%
Specificity%
PPV%
NPV%
Efficiency%
2D CD
97.0
41.0
64.0
92.6
70.1
3D PD
97.0
39.3
63.7
92.3
69.5
NPV indicates negative predictive value; PPV, positive predictive value; 2D CD, 2D color Doppler ultrasonography; 3D PD, 3D power Doppler ultrasonography.
Ultrasonography and color Doppler imaging can help to predict the success rates in assisted reproduction (Table 19.11).328
Table 19.11   Factors predicting higher pregnancy rates
• Endometrium—Triple layer appearance and 12 mm thickness
• Endometrial power Doppler area > 5 mm sq
• Myometrium appears homogenous
• Uterine artery PI < 3
• Uterine artery end-diastolic flow present
• Perifollicular circumfrential vascularity > 75%
• Ovarian stromal arteries—PSV > 10 cm/sec
Ultrasound imaging of the female pelvis has helped us to understand, identify, diagnose, treat and manage the infertile patient. The endovaginal scanning and Doppler imaging has enabled us to extend our evaluation further. It is now possible to perform a sonographic physiologic assessment of the structures we visualize.
 
References
  1. Fleischer AC, Kepple DM, Vasquez J. Conventional and color Doppler transvaginal sonography in gynecologic infertility. Radiol Clin North Am. 1992;30:693–702.
  1. Fleischer AC, Daniell JF, Rodier J, Lindsay AM, James AE Jr. Sonographic monitoring of ovarian follicular development. J Clin Ultrasound. 1981;9:275–80.
  1. Panchal S, Nagori CB. Pre-hCG 3D and 3D power Doppler assessment of the follicle for improving pregnancy rates in intrauterine insemination cycles. J Hum Reprod Sci. 2009 Jul;2(2):62–7.
  1. Kurjak A, Kupesic-Urek S, Schulman H, Zalud I. Transvaginal color flow Doppler in the assessment of ovarian and uterine blood flow in infertile women. Fertil Steril. 1991;56:870–73.
  1. Steer CV, Campbell S, Pampiglione JS, Kingsland CR, Mason BA, Collins WP. Transvaginal color flow imaging of the uterine arteries during the ovarian and menstrual cycles. Hum Reprod. 1990;5:391–5.
  1. Steer CV, Campbell S, Tan SL, Crayford T, Mills C, Mason BA, et al. The use of transvaginal color flow imaging after in vitro fertilization to identify optimum uterine conditions before embryo transfer. Fertil Steril. 1992;57:372–6.
  1. Sterzik K, Grab D, Sasse V, Hütter W, Rosenbusch B, Terinde R. Doppler sonographic findings and their correlation with implantation and in an in-vitro fertilization program. Fertil Steril. 1989;52:825–28.
  1. Friedler S, Shenker JG, Herman A, Lewin A. The role of ultrasonography in the evaluation of endometrial receptivity following assisted reproductive treatments: a critical review. Hum Reprod. 1996;2:323–35.
  1. Fleischer AC. Ultrasound imaging—2000: Assessment of utero-ovarian blood flow with transvaginal color Doppler sonography; potential clinical applications in infertility. Fertil Steril. 1991;55:684–91.
  1. Applebaum M, Cadkin AV. Decidual flow—an early sign of pregnancy. Ultrasound Obstet Gynecol. 1992;2:65.
  1. Fleischer AC, Gordon AN, Entman SS, Kepple DM. Transvaginal scanning of the endometrium. J Clin Ultrasound. 1990;18:337–49.
  1. Giannaris D, Zourla A, Chrelias C, Loghis C, Kassanos D. Ultrasound assessment of endometrial thickness: correlation with ovarian stimulation and pregnancy rates in IVF cycles. Clin Exp Obstet Gynecol. 2008;35(3):190–3.
  1. Kurjak A, Zalud I. Transvaginal color Doppler in the study of uterine perfusion. In: Mashiach S, Ben-Rafael Z, Laufer N, Schenker JG (Eds). Advances in Assisted Reproductive Technologies. New York: Plenum Press.  1990;541–4.
331Index
Page numbers followed by f refer to figure and t refer to table
A Abdominal cramps distension girth measurement Abdominopelvic discomfort Abnormal anemic flow Abortion , Absence of corpus luteum Acanthosis nigricans Acarbose , Accuracy of test Acid-base balance Acne Addison's disease Adenoma with stalk compression Adjunctive use of nitric oxide , Adnexal torsion Adrenal function assessment Advantages of GnRH agonists antagonists over GnRH agonist in ART pulsatile therapy ovarian reserve testing recombinant FSH transabdominal route transvaginal procedure vaginal administration Amenorrhea , , , , , Amitryptilene Amount of ascites Amoxapine Anastrazole Androgen receptor expression Anovulation and tests for ovulation Antagonizing androgens Anti-Müllerian hormone , tailored protocols Antiphospholipid syndrome Antipsychotic drugs Antral follicle count , , , Appearance of ovary after drilling Aromatase inhibitors Ascitis Aspiration of single lead follicle Aspirin , , Assessment of ovarian reserve Atresia B Basal anti-Müllerian hormone body temperature , follicle stimulating hormone levels , , ovarian stromal blood flow Bilateral polycystic ovaries Bleeding per vaginum Blood gases tests Blunting of hepatic gluconeogenesis Body mass index Bone mineral density Breast cancer , discomfort and bloating Bright echogenic stroma Bromocriptine , , , , , Buserelin Butyrophenones C Cabergoline , Calcium channel blockers CAM therapy Causes of anovulation suitable for ovulation induction treatment hypogonadotropic hypogonadism poor ovarian response CC challenge test Central retinal vein occlusion Cerebral radiotherapy Cervical mucus Chemotherapy Chest wall stimulation X-ray Choice and dose of gonadotropin Chromohysteroscopy Chronic endometritis , , nonspecific endometritis renal failure Cimetidine Cirrhosis Clomiphene , , , , , and gonadotropin regime challenge test citrate , , , , challenge test failure in mild stimulation protocol resistance , therapy with gonadotropins and GnRH antagonist with single dose FSH Colony-stimulating factor Color and pulsed Doppler ultrasound Doppler imaging Combined oral contraceptive pills Complications of ovulation induction Congenital adrenal hyperplasia malformations uterine abnormalities Constipation Corpus luteal flow Correction of circulatory volume electrolyte imbalance Correlation of number of oocytes recovered with pregnancy rate Corticotropin releasing hormone stimulation test Craniopharyngioma , Creatinine Cryopreservation of embryo , , of oocyte , Cumulative conception rate pregnancy rate Cushing's syndrome , Cyproterone acetate Cystectomy Cytokines D Danazol D-chiroinositol , Decidual prolactin measurement Decreased renal function SHBG Dehydroepiandrosterone sulfate Dehydrogesterone Delayed puberty Dendritic cells Determination of ovarian tissue Development of GnRH antibodies Dexamethasone suppression test Diabetes insipidus mellitus Diminished ovarian reserve , , Dizziness , Dominant follicle ovary Donor oocytes Dopamine agonists , , receptor blockers synthesis inhibitors Dose of clomiphene FSH Drainage of hydrosalpinx Drug-induced hypersecretion Dynamic tests Dysgerminoma Dyslipidemia Dysparenia E Early follicular phase estradiol levels inhibin B levels Elevated circulating androgens Emotional stress Empty sella syndrome Endometrial aspiration biopsy , , , carcinoma growth hyperplasia lymphocytes morphology , polyps power thickness , , , vascularity , volume , Endometriosis , Endometritis Endometrium , Endothelial proliferation Epidermal growth factor Epileptic seizures Estrogen deficiency replacement therapy Evaluation of male partner Exogenous FSH ovarian reserve test , gonadotropin stimulation hCG Extended lithotomy position F Failure of medical therapy for ovulation induction Fasting blood sugar insulin levels serum insulin Fatigue Flexible antagonist protocol Fluid in cul-de-sac Flutamide Focal or diffuse hyperemia Follicle aspiration stimulating hormone , , , Free androgen index Functional hypothalamic chronic anovulation G Galactorrhea , , Glucocorticoids , , , , Glucose tolerance test GnRH agonist , , , , , , , , for ovulation trigger in luteal phase pulsatile therapy therapy GnRHa stimulation test Golan's classification of OHSS Goldmann's perimetry Gonadotropin , , , , releasing hormone analog challenge test stimulation therapy , Grades of severity of hypothalamic amenorrhea Granulomas Granulosa cells of primordial follicles and follicles Grasping of ovarian ligament Growth hormone replacement H Hair loss and dryness Hallucinations Hashimoto's thyroiditis Headache , , Hematoma Hepatic dysfunction High dose of gonadotropins , Highly purified human menopausal gonadotropin Hormonal control of endometrial preparation evaluation Hormones Hot flushes , Human chorionic gonadotropin , , , menopausal gonadotropin , , , pituitary gonadotropin Hydrothorax Hydroxyethyl starch administration Hypergonadotropic hypogonadism Hyperinsulinemia in polycystic ovarian syndrome Hyperplasia of lactotrophs Hyperprolactinemia , , , , , Hyperstimulation syndrome Hypertension and cardiovascular disease Hypoglycemia Hypogonadotropic hypogonadism , , , , , , , , , and ovulation induction Hypophysectomy Hypotension Hypothalamic pituitary dysfunction failure , stalk damage Hypothalmic disorders Hypothyroidism , , Hysteroscopy , I Idiopathic hyperprolactinemia Imipramines Induction of ovulation Infertility Injection of intrathoracic potassium chloride Insertion of needle Insulin resistance sensitizers , , sensitizing drugs Intolerable nausea and vomiting Intracranial injection of potassium chloride Intramuscular depot injections Intranasal route Intrathoracic injection of potassium chloride , Intravenous immunoglobulins Invasive hemodynamic monitoring Irregularity of follicle K Kallmann syndrome Ketoconazole Kidney function tests L Laparoscopic evaluation of pelvis ovarian drilling , , Laparotomy Large granular lymphocytes L-arginine Late follicular phase index values luteal phase index values Leaking per vaginum Letrozole , , , , , Leukemia inhibitory factor Leukocyte immunotherapy Leuprolide Liver disease dysfunction enzymes function tests , toxicity Loading of syringe Long-acting depot intramuscular injection Loss of remaining fetuses Low dose of gonadotropin oral contraceptive pills estradiol levels gonadotropins ovarian stimulation protocols progesterone levels in luteal phase resistance pattern Lubeck protocol , Lupus erythematosis Luteal estradiol , initiation of FSH phase defect , support , serum progesterone levels supplementation support , Lutein cyst formation Luteinized unruptured follicle , Luteinizing hormone , , Lymphocytic hypophysitis M Macro-adenoma Macroprolactin Maintaining normal endometrium Maintenance of intravascular volume and electrolyte imbalance renal function Management of ascitis clomiphene failure resistance hyperprolactinemia , luteal phase defect OHSS PCOS Mechanism of hyperinsulinemia and hyperandrogenemia Medical treatment of endometritis Meningioma Menstrual disturbance irregularities Metalloproteases Metformin , , , , , , Methods of endometrial evaluation multifetal pregnancy reduction Metoclopromide Micro-adenoma Micronized progesterone Micropolyps Mifepristone Mild ovarian stimulation , Mild stimulation protocol , Miscarriage Modes of ovulation induction in PCOS Monitoring of thyroid activity Mucosal edema Mullarian inhibiting substance Multiple endocrinopathies pregnancy , , , N N-acetyl cysteine Naltrexone , Nasal congestion spray Natural cycle killer cells Nausea , , Navot's classification Nitric oxide Nitroglycerin Nonapeptide agonists Nonspecific chronic endometritis Number of follicles , mature follicles O Obesity Oligomenorrhea , Oligo-ovulation Oocyte donation , pick-up Optic nerve Optimal endometrial score , and vascularity quantification perifollicular flow stromal artery flow uterine artery RI score Oral contraceptive , , , , pill , , Orthostatic hypotension Osteopontin Osteoporosis Ovarian artery cancer , and ovulation induction cyst drilling enlargement failure , , hyperstimulation syndrome , , , , reserve tests , sensitivity index size stimulation stroma stromal arteries peak systolic velocity suppressive agents transplant volume , , Ovaries , irrigated with normal saline Overnight dexamethasone suppression test Ovulation , documentation induction protocol treatment with clomiphene P Panhypopituitarism , Paracentesis of hydrothorax Pathophysiology of luteal phase defect ovarian hyperstimulation syndrome , Peak systolic velocity Perifollicular circumfrential vascularity Perivascular fibrosis Persistent hypersecretion of LH perifollicular reaction Phenothiazines Pioglitazone Piroxicam Pituitary capacity test dynamic testing failure function assessment gland hypersecretion stimulation test tumor Plenty of fluids Polycystic ovarian disease syndrome , ovaries , syndrome , Poor hormonal environment ovarian reserve and mild ovarian stimulation uterine artery blood flow Post-oocyte retrieval albumin Postpartum hemorrhage Postprandial insulin Pouch of Douglas Pregnancy , , and lactation complications monitoring rate , Premature luteinization menopause ovarian aging Prevention of endometrial hyperplasia thrombosis , Previous ovarian surgery Primary amenorrhea Principle of ELISA test Procedure of multifetal pregnancy reduction Progesterone supplementation , Prolactin and ovarian function secreting adenoma Prolactinoma Proliferation of spiral arteries Protocol of ovarian stimulation Pulmonary compromise Pulsatility index , , Pulse frequency and dose Purified urinary FSH Q Quantify blood flow Quinagolide , R Radioimaging of sella turcica Ranitidine Raynaud's phenomenon Recombinant FSH , , , hCG LH , Recurrent abortion Reducing uterine contractility Regularization of cycles Renal dysfunction failure , Resistance ovarian waveform pattern Resistive index , Results of gonadotropin therapy laparoscopic treatment Rheumatoid arthritis Ritodrine Role of androgens in ovulation induction chronic endometritis in endometrial receptivity GnRH agonists and antagonists in assisted reproductive technology GnRH antagonist in mild stimulation , LH supplementation metformin after conception natural killer cells oral contraceptive pill Rosiglitazone S Scanning electron microscopy Selective multifetal pregnancy reduction Sella turcica Serum estradiol levels prolactin estimation measurement Severe OHSS Sex hormone-binding globulin Sheehan's syndrome Sibutramine Side effect of bromocriptine Signs of ovulation on ultrasonography Sildenafil Simple needle puncture office procedure Slide test Small for gestational age Spindle damage Spiral arteries Spironolactone Standard flare protocol Steroid hormone receptor analysis Stimulation of folliculogenesis Storage of steroid hormones Strenuous exercise Stress , Strict intake output chart Study of pinopodes Subcutaneous injections Sub-endometrial blood flow , Suppressive therapy Supraseller pituitary mass extension Surgical ovulation induction , , , Systemic disorders T T regulatory cells Tamoxifen , , Termination of pregnancy Testosterone Tests for ovulation Thiazolidinediones Thioxanthenes Thyroid stimulating hormone , Transabdominal procedure of fetal reduction Transcervical aspiration of gestational sac Transdermal testosterone gel , , Transient hyperprolactinemia Transvaginal procedure of fetal reduction puncture and embryo aspiration , sonography ultrasound scan Treating anovulation Treatment of hyperprolactinemia luteal phase defect poor uterine receptivity prolactin secreting macroadenomas Triple layer endometrial thickness Triptorelin , Tube Tubercular endometritis , Tuberculosis and chronic nonspecific endometritis Tumor necrosis factor alpha Turner's syndrome Typical triple layer endometrium U Ultrasonographic scoring of uterine receptivity Ultrasonography of ovary Ultrasound machine Unexplained infertility , , Unilateral oophorectomy Use of general anesthesia LH recombinant FSH Uterine artery , dysfunction Uterus , V Vaginal dehydroepiandrosterone dryness Visual disturbances Visualization of needle tip Vomiting , , W Wall thickening and coiling WBC count Weight charts loss , , ,