Introduction:

In women with normal oocyte reserve undergoing in vitro fertilization embryo transfer (IVF-ET) with normal oocyte reserve (NOR) it is estimated that one could expect about 80-85% of the oocytes retrieved to be fully mature, i.e., they are in the metaphase II (MII) stage with 15-20% immature oocytes (germinal vesicle stage or MI stage) [1]. Culturing metaphase I oocytes or even germinal vesicle stage embryos another day, followed by fertilization by intracytoplasmic sperm injection (ICSI) (because the zona pellucida has been stripped) results in a much lower percentage of euploid embryos compared to those inseminated on the day of oocyte retrieval [2-4].

Conti and Franciosi, suggested that the low fertility potential of these rescued immature oocytes is related to lack of coordination of the normal events leading to euploidy of the zygote including coordination of fertilization events, reprogramming of the parental genomes, DNA replication, and embryonic genomic activation [5]. Indeed, most studies suspect that in vitro mature rescued MI oocyte lead to a much higher chance of an aneuploid embryo [6-9].

There have been live deliveries reported following culturing of MI to MII oocyte with subsequent embryo transfer dating back to the late 1990s [10,11]. These early studies simply transferred the embryos cultured for two days on the 3rd day after oocyte retrieval in contrast to M-II oocytes fertilized on the day of retrieval that would be allowed to develop for 3 days prior to embryo transfer. Though the appearance of morphologically normal appearing embryos reported in some studies was reasonable, and sometimes comparable to embryos derived from fertilization of retrieved MII oocytes, the live delivered pregnancy rates (LDPRs) were poor. For example, in the aforementioned study by Reichman et al, though they demonstrated similar fertilization rates of MI oocyte vs MII oocytes, embryo quality was only half as good. They had no live deliveries out of 17 embryos obtained by extended culture and delayed fertilization [4].

As mentioned, rescued oocytes seem to have lower fertilization rates [2-4, 12,13]. However, some claim that the embryo quality as determined by embryo morphology may be similar [12]. Nevertheless, these rescued oocytes seem to have defective cytoplasmic maturation, increased meiotic aberrations, and nuclear degeneration (14). Normal morphologic embryos lead to similar blastocyst formation, but yet higher rates of aneuploidy [15]. Furthermore, there is a difference in the morphokinetic characteristics of embryos derived from in-vitro matured vs in vivo matured oocytes after ovarian stimulation [16].

There may be one other explanation for lower pregnancy rates following the fresh transfer of embryos derived from rescued vs embryos from fertilization of MII oocytes and that is the possible adverse consequence of asynchrony between the stage of embryo development vs endometrial development, i.e., placing either day 3 embryos or blastocysts in the uterine cavity that is one day earlier than if the egg was fertilized on the day of retrieval. Thus, Ming et al found a higher pregnancy rate if instead of transferring fresh to a uterus with an asynchronous endometrium, to freeze the embryos derived from the fertilization of a rescued oocyte and delay transfer of the frozen thawed embryos in a subsequent cycle where proper synchronization could be achieved [17]. Aizer et al found that the LDPR following fresh embryo transfer derived from rescued oocytes was 1.6% vs 8.3% with frozen-thawed embryos derived from rescued oocyte embryos transferred to the uterus with a synchronous stage of development as the frozen-thawed embryo [18]. A live fetus in the 1st trimester was reported in a poor responder using a rescued oocyte. However, the patient was an unexpected non responder with no evidence of DOR before ovarian hyperstimulation [19].

Most infertility centers will cancel an IVF-ET cycle if they expect to retrieve only 1 or 2 oocytes. The majority of infertility centers will advocate the use of donor oocytes if there is marked diminished oocyte reserve (DOR). Nevertheless, at our institution we have had considerable experience in helping women with marked DOR to become pregnant with their own oocytes. For over 40 years, since we published a novel technique on restoring down-regulated FSH receptors in women in apparent premature ovarian failure (POF) that not only results in the induction of ovulation but also achieved live deliveries [20-22].

We encourage women with DOR to conceive without IVF-ET because without multiple oocytes to create more embryos it does not seem prudent to encounter the increased expense and invasiveness of IVF-ET. However, when there is a need for IVF, e.g. damaged fallopian tubes or a severe sperm defect, it is reasonable to consider IVF-ET even if there is a likelihood of only one egg being retrieved, or even with the combination of DOR and advanced age [23-25].

The question arises as to whether it is worthwhile to culture an MI oocyte to the MII stage in women with such severe DOR that they can only develop 1 dominant follicle, or in women that appear to be in overt menopause where development of a dominant follicle is attained by techniques aimed to up-regulate down-regulated FSH receptors [26, 27].

Material and Methods

A retrospective review was performed over a 25-year time period to identify women having such severe DOR that there was only one oocyte retrieved. The number of patients with a single metaphase II oocyte vs metaphase I was determined. Very immature oocytes, i.e., germinal vesicle stage, were not included. Following fertilization of the MI to MII oocytes. The embryos continued culture and were transferred fresh 4 days from the oocyte retrieval.

The oocyte stimulated protocol was a specific type of mild stimulation protocol known as an FSH receptor upregulation technique [27]. Details of the variations in this protocol according to the patients' serum hormonal levels (LH, FSH, estradiol (E2) and progesterone (P)) have been provided [27] This protocol may vary from patient to patient or cycle to cycle in the same patient [27]. There is evidence that this stimulation protocol maximizes the chance of a successful pregnancy based on evidence that raising the serum FSH too high can down-regulate a key FSH dependent cytokine or enzyme needed for the embryo to implant and the fetus to survive [28, 29].

Results

There were 42 women aged <39.9 who had a single MI oocyte cultured to MII. Fertilization occurred in 24 of 42 (53.9%) and cleavage to day 3 occurred in 18 of the 24 women (75%). Fourteen women had a fresh ET and 4 cryopreserved the embryo. None of the 4 women with cryopreserved embryos had a subsequent single embryo transfer so they were not included in the data. Two of the 14 women (14.3%) having a fresh ET conceived and had ultrasound evidence of pregnancy (clinical pregnancy) and 1 had a successful full-term delivery of a healthy child (7.1%)

There were 40 women > 40 where the sole oocyte retrieved was in the metaphase I stage. Fertilization occurred in 23 of 42 (54.8%) leading to 15 of 23 (65.2%) cleaved embryos to transfer; however, there were no pregnancies.

Discussion

In using the FSH receptor up regulation technique to mature oocytes for oocyte retrieval, live delivered pregnancy rates per (LDPRs) per transfer of a single embryo derived from an MII oocyte for women < age 39 is between 20-25% [25, 27-33]. This study shows that despite the appearance of normal morphologic embryos that were obtained from rescued oocytes, these embryos are much less likely to produce a live delivered baby. Nevertheless, a live delivery is possible (at least in women aged <39.9 and thus if they have already proceeded to oocyte retrieval) there does not seem to be any downside to culturing the rescued oocyte to the MII stage and do an embryo transfer. Whether higher pregnancy rates would be achieved by deferring fresh transfer and cryopreserving the embryo to allow a more synchronized transfer with luteal P effect on the endometrium in a subsequent cycle is at least possible [17, 18].

Women with extremely low oocyte reserve have a very reduced chance of conceiving a live baby with or without IVF-ET. Thus, IVF should be restricted to couples that require the procedure for damaged tubes or very poor sperm quality. This is the first case demonstrating that a live delivery is possible even when the only oocyte retrieved is MI oocyte and the woman has severe DOR.

Conclusion

In women with such severe DOR that there is only 1 oocyte to retrieve, one must consider that this group is much different than the previous publications of outcomes of rescued MI oocytes. Even those reporting outcome in poor responders, the finding of the poor response and the presence of only immature oocytes was unexpected, i.e., not predicted by serum AMH level or day 3 FSH. Furthermore, in the past fresh embryos were transferred 3 days after oocyte retrieval. We believe that this is the first evaluation of transfer of day 3 embryos, 4 days after oocyte retrieval, thus, allowing assisted embryo hatching.

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