SUMMARY
In Assisted Reproductive Technologies (ART), gametes and/or embryos are handled outside the body at shorter or longer periods, under the microscope, in incubators, and so on. This means that in order to achieve a pregnancy, the gametes and embryos must be replaced into the human body. The most common such procedure is transcervical embryo transfer (ET) into the uterine cavity. The historical background and alternatives to this technique are discussed in the present chapter. Embryo transfer is one of the crucial steps, where science and art meet, and where the optimum final solution for all patients is not yet found.
Introduction
Transfer of an embryo from one uterine cavity to another was first described in the rabbit by Walter Heape as early as in 1891.1 An in vivo fertilized embryo was flushed out from the uterus of one rabbit and placed into another hormonally synchronized animal, and this procedure resulted in a live offspring.
Embryo transfer (ET) following in vitro fertilization (IVF) was also first described in the rabbit in 1955,2 followed by successful experiments in the mouse in1968, and the rat in 1974.3 In the experimental design for these species, one animal's eggs were fertilized and following culture for various lengths of times, transferred to another animal. In the case of human beings, at least in the beginning, before the era of egg and embryo donation, the eggs were collected from the same woman to whom the resulting embryo was transferred after fertilization (autologous procedure). For that reason, the pioneers at Bourn Hall preferred the term ‘embryo replacement’ or initially, even ‘re-implantation’ in the human situation. Many other linguistic expressions, such as egg transfer (even if the oocyte is fertilized), pre-embryonic transfer to stress that the cleavage stage transfers (day 2 or 3) refer to pre-embryos and not embryos according to a strict definition have however, been used through the years ‘Blastocyst transfer’ is another 4expression utilized that refers to day 5 or 6 transfers. When referring to the human situation, where infertility problems are the main reason for applying the techniques, an umbrella term, which is nowadays frequently used, is ‘assisted reproductive technologies (ART)’ and one of the central techniques is embryo transfer.
Clinical Discussion
Methods for Transfer of Fertilized Eggs
Surgical transfer versus cervical ET has been compared in the past in various animal models, generally with a higher success rate with the use of surgical transfer. In the human, only a few clinical IVF groups have experience with surgical transfer on a large scale, and in the majority of the cases where surgical transfer has been used, vaginal or cervical malformations have been the reason for choosing this technique.4, 5 Although it was agreed that surgical transfer involved a decreased risk for infections, it probably caused more contractions and local release of substances (e.g. cytokines) in the uterine wall that could interfere negatively with the implantation process, especially in cases with active endometriosis for instance. In the human, surgical transfer is also regarded as a considerably more invasive and painful technique in the routine case.
Placement of the Embryos—The Fallopian Tube versus the Uterine Cavity
A major concern in relation to ET that has been discussed intensely, is the optimal location for placement of the embryo. Dependent on the indication for IVF, the embryo could be placed either in the fallopian tube-tubal embryo transfer (TET) in couple with male factor infertility and immunological infertility, or in the uterine cavity if the fallopian tubes are not patent. If at least one of the fallopian tubes is patent, it is a more physiological place to replace embryos in the 2-8 cell stages than the uterine cavity, and it was thus, not surprising that Asch in 1991,6 could demonstrate an almost doubled success rate applying TET for the right indications in comparison to IVF. The risk of ectopic pregnancy was not increased by the use of TET. Tubal embryo transfer is seldom used today, since the work-up of the patients prior to IVF does not always give detailed knowledge about tubal patency or peritubal conditions. Additionally, the use of gamete intrafallopian transfer (GIFT) has gradually disappeared for similar reasons. Apart from Asch's group, a number of publications describing similar techniques appeared in the late 1980's. One such technique, called zygote intrafallopian transfer (ZIFT), was reported from Brussels by Devroey et al7 in 1987. The advantage of ZIFT compared to GIFT was that fertilization of the oocyte could be verified under the microscope prior to transfer. Additionally, the IVF center in Bonn published at the same time, data on what they called transvaginal intratubal transfer as compared to laparoscopic intratubal transfer.8
While one was obviously more laborious, no significant differences were however found in their material between these two techniques (Fig. 1.1).
Embryo Placement Technique at Various Positions in the Uterine Cavity
Cervical versus fundal replacements have been discussed in the beginning of the IVF era. The two major concerns were uterine contractions and infection. Different kinds of transfer catheters have been introduced in order to minimize the risk for infections. In certain IVF programs, especially in Germany, prophylactic antibiotics were given to all patients prior to transfer. A few IVF groups preferred, at least in the beginning of the IVF era, to transfer embryos under general anesthesia and even now, some type of sedation is frequently utilized. Transfer is today, generally performed in the lithotomy position, even though the knee-chest position was initially sometimes used at centers such as Bourn Hall, with a vaginal speculum in position. The vagina and the outer parts of the cervical canal are carefully washed using embryotoxic physiological saline or especially prepared culture medium. Embryo toxic solutions containing iodine for instance, should naturally be avoided in the transfer procedure.
The so-called ‘mock’ or ‘dummy’ transfers prior to the real transfer, as described by Mansour and co-workers, have been recommended by certain IVF groups, but compiled data have not shown convincing advantages in routine cases. Since ultrasound is frequently used in connection with monitoring and oocyte pick-up, the position of the uterus is already relatively well known in most cases. Utilization of abdominal ultrasound in connection with ET has been recommended for a long time. It was first reported by Strickler et al.10 in 1985 using an abdominal transducer.
Fig. 1.2: Ultrasound scan showing the uterus and the endometrium. Arrow indicates the position of the transfer catheter tip at ET
Later, Hurley et al11 utilized a transvaginal transducer, which is not, however, used by all IVF-groups as a routine in all cases. This procedure, however, enables the doctor to place the tip of the transfer catheter in an optimal position, which is today, considered to be in the mid-region of the uterine cavity, not very close to the fundus (Fig. 1.2). At transfer, a small echo created by the injected transfer medium, is generally seen on the screen and this confirms the exact position of the tip of the catheter.
The amount of medium, which is used for transfer, has decreased from around 150 μl in the early days to approximately 15 μl today. Following transfer, the catheter is withdrawn gently, immediately or after a short delay. In the early days, when high transfer volumes were common, embryos were relatively frequently flushed up in the tubes and resulted in a high incidence of ectopic pregnancy. For such reasons, Steptoe recommended surgical occlusion or coagulation of the tubes close to the uterine wall prior to IVF.
Day of Transfer
Transfer of gametes, zygotes and embryos in different stages of development up to the blastocyst stage has been tried in animal models and in humans for many years without leading to a final conclusion or agreement about the optimal time for replacement in clinical IVF. The culture technique and culture conditions, as well as the IVF unit's cryopreservation programs, are of course, crucial for the choice of transfer day in various IVF programs. Transfer of zygotes and embryos in the 2-8 7cell stage into the uterine cavity does not offer an optimal uterine milieu, since physiologically, the embryo remains in the fallopian tube up to the blastocyst stage. A rapid passage of embryos through the fallopian tube by use of hormonal treatment (e.g. gestagen only pills or prostaglandins) has even been recommended for contraceptive purposes. The uterine wall also contracts rhythmically at this stage, which may increase the risk for expulsion of the embryo through the cervical canal.
Prolonged culture also gives the embryologist an increased possibility of evaluating the quality of embryo better at a later stage. A relatively new technique, also in support of prolonged culture, is a genetic evaluation of the embryo prior to transfer, the so-called preimplantation genetic screening (PGS). In order to make this evaluation, starting at the 8 cell stage, transfer cannot be performed until day 4-6 if freezing is not introduced in the procedure.12, 13
Initially, ET was mostly performed in the evening after the dark. This was based on studies, which had shown that a diurnal rhythm existed in the uterine activity with fewer contractions at night.14 Bed rest for 12-24 hours after transfer was recommended in the early days of IVF. Today, patients can generally leave the IVF clinic 30 minutes after the transfer and no adverse effects have been recorded by this simplification in procedure. Transfers are also performed during all times of the day, and if performed in a dark room, this is mainly done for protection of the embryos from bright light. Seasonal differences were also discussed in the early days, since such differences have been reported to be of importance in various animal models. Today, seasonal variations are not believed to be of importance in the human situation.
Transfer Media
In the past, relatively simple culture media were used and prolonged embryo culture was not practised, since this generally caused degeneration of the embryos. Today, more and more IVF-programs, utilizing sequential culture media, prolong their culture to the blastocyst stage, since a more optimal choice of the best embryo(s) can then be made. A majority of transfers are, however, still performed on day 2 or 3, very few on day 4, but gradually, an increasing percentage are being delayed to day 5 or 6.
The importance of serum for successful growth of human embryos was realized at an early stage and in the original setting at Bourn Hall, Hams F10 medium fortified with the patient's serum, was generally used for culture. It was later suggested that even higher concentrations of serum (75-100%) should be used in connection with ET. However, Menezo and co-workers15 could not confirm that increased viscosity in the transfer medium improved the pregnancy rates after ET. A decade later, Gardner and Lane16 demonstrated that replacement of protein with glycosamino glycan 8hyaluronate promoted implantation, at least in animal studies. Many culture and transfer media have been fortified with antibiotics for a considerable period of time, and for that reason, may cause allergic reactions in rare cases. Since the amount of medium introduced into the uterine cavity at transfer is so small, general reactions are very seldom noticed, however, a local intrauterine reaction, which may negatively interfere with the chances of implantation cannot be excluded.
Choice of Transfer Catheter
Ever since the start of clinical IVF, various types of catheters have been designed and compared. The original Bourn Hall catheter was soon replaced by a softer catheter (Wallace). Later came Craft, Tight difficult transfer (TDT) and Frydman catheters, all with variations in the material, flexibility, memory, softness, etc17, 18 Results were compared in terms of pregnancies and number of transfers that could be classified as easy or difficult. The conclusion from all these comparative studies is that the skill of the doctor performing the transfer is more important than the choice of catheter. In the small group of real difficult transfers, dilatation of the cervix in an earlier cycle-so called mock transfer, transfer under general anesthesia or transmyometrial surgical transfer, may be optimal methods of choice.17
Number of Embryos for Transfer
Initially, natural cycles of IVF were performed that resulted in a single embryo transfer (SET) in most cases. With the introduction of clomiphene citrate19 or gonadotropins,20 a higher number of fertilizable oocytes could be obtained and the temptation to transfer more than one embryo at a time to improve the results became so high that most IVF groups replaced up to five embryos in each cycle. Even though pregnancy rates increased, multiple pregnancy rates also increased dramatically. Today, an increasing number of IVF groups reduce the number of embryos for transfer to 2 or 1, and in some countries, strong recommendations from the central authorities for the replacement of fewer embryos must be followed.
In contrast to the situation 25 years ago, cryopreservation programs are now available for surplus embryos that can be replaced one at a time later on in natural cycles if the initial stimulated cycle failed to produce a pregnancy. With this procedure, the cumulative pregnancy rate could be as good as if more embryos are replaced in each transfer.21
CONCLUSION
The moment of ET in clinical IVF is a co-ordination between the embryologist's efforts in the laboratory on the one hand and the clinician with responsibility for the patient on the other. In the beginning, and 9occasionally even now, the laboratory and the operation theatre were often separated and only connected by a hole in the wall.
The laboratory delivers embryos in different stages of development, partly dependent upon the clinician's wishes. The embryos are enclosed in small volumes of medium, sometimes especially designed for transfer in catheters, which should be sterile and easy to handle in order to avoid blood or mucus on the catheter tip. At transfer, it is further important to avoid uterine contractions, or end up with retained or expelled embryos. The number of embryos to be replaced in each trial is nowadays, frequently discussed between the embryologist and the clinician, dependent upon alternatives such as prolonged culture, cryopreservation and other facilities in the laboratory outfit. Large differences in the management of the IVF laboratory, as well as in success rates, have been reported among various clinicians.
Joint efforts among personnel during the execution of the various steps in the IVF procedure seem to be of utmost importance since the importance of an optimal ET has been evaluated or analyzed separately.
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- Chang MC. La Fonction Tubaire et ses Troubles 1955:40–52. Mason et Cie, Paris.
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