Outcome of frozen embryo replacement cycles following elective cryopreservation of all embryos in women at risk of developing ovarian hyperstimulation syndrome

Aim: Our aim was to compare the outcome in subsequent frozen embryo replacement cycles in four groups of patients who had elective cryopreservation of all their embryos because they were considered to be at increased risk of developing severe ovarian hyperstimulation syndrome. Design: Sixty-two (91%) of 68 IVF cycles (68 patients) in which elective cryopreservation of all embryos was performed were analyzed. All patients continued on the GnRH agonist, buserelin, after oocyte recovery until the onset of vaginal bleeding. Frozen embryo replacement occurred in a hormone replacement cycle that started either on day 3 of the withdrawal bleed (group I;N=15) or after serum estradiol levels had fallen to <100 pmol/L (group II;N=16). The other patients commenced a frozen embryo replacement cycle several months later in either a hormone replacement (group III;N=15) or a natural (group IV;N=16) cycle. Results: Two patients developed severe ovarian hyperstimulation syndrome. There were no significant differences among the four groups regarding demographic variables, the dose of hMG used, and the clinical outcome. There was a higher but not significantly different clinical pregnancy rate in group I (26.7%), compared to group II (12.5%), group III (13.3%), and group IV (18.8%). Conclusions: Several options exist for the timing and protocol used for frozen embryo replacement in patients who had elective cryopreservation for the prevention of ovarian hyperstimulation syndrome, none of which was found to be clearly superior in this observational report.Presented at the 1994 Annual Conference of the American Fertility Society.     

Miscarriage rates following in-vitro fertilization are increased in women with polycystic ovaries and reduced by pituitary desensitization with buserelin

To assess the risk of miscarriage after in-vitro fertilization(IVF) with respect to age, cause of infertility, ovarian morphologyand treatment regimen, a retrospective analysis was performedof the first 1060 pregnancies conceived between June 1984 andJuly 1990 as a result of 7623 IVF cycles. Superovulation inductionwas achieved with human menopausal gonadotrophin (HMG) and/orpurified follicle stimulating hormone (FSH) together with eitherclomiphene citrate or the gonadotrophin hormone-releasing hormone(GnRH) agonist buserelin, the latter either as a short ‘flare’regimen or as a ‘long’ regimen to induce pituitarydesensitization. There were 282 spontaneous abortions (26.6%)and 54 ectopic pregnancies (5.1%). The mean age of women withongoing pregnancies was 32.2 (SD 3.9) years compared with 33.2(SD 4.1) years in those who miscarried, which were significantlydifferent (P = 0.008). There was no relation between the miscarriagerate and the indication for IVF. The miscarriage rate was 23.6%in women with normal ovaries compared with 35.8% in those withpolycystic ovaries [P = 0.0038, 95% confidence interval (CI)4.68–23.10%]. There was no difference in the miscarriagerate between treatment with HMG or FSH. Women whose ovarieswere normal on ultrasound were just as likely to miscarry ifthey were treated with clomiphene or with the long buserelinprotocol. Those with polycystic ovaries, however, had a significantreduction in the rate of miscarriage when treated with the longbuserelin protocol, 20.3% (15/74), compared with clomiphenecitrate, 47.2% (51/108) (P = 0.0003, 95% CI 13.82–40.09%).     

A controlled study comparing patients with and without polycystic ovaries undergoing in-vitro fertilization

The outcome of in-vitro fertilization and embryo transfer (IVF—ET)was compared in 76 patients with polycystic ovaries (PCO) diagnosedon pre-treatment ultrasound scan, and 76 control patients whohad normal ovaries and were matched for age, cause of infertilityand stimulation regimen. Despite receiving significantly lesshuman menopausal gonadotrophin (HMG), patients with PCO, ascompared with controls, had significantly higher serum oestradiollevels on the day of human chronic gonadotrophin administration(5940 ± 255 versus 4370 ± 240 pmol/1, P < 0.001),developed more follicles (14.9 ± 0.7 versus 9.8 ±0.6, P < 0.001) and produced more oocytes (9.3 ± 0.6versus 6.8 ± 0.5, P = 0.003). However, fertilizationrates were reduced in the PCO patients (52.8 ± 3.4% versus66.1 ± 3.4%, P = 0.007). There was no significant differencein cleavage rates. The pregnancy rate/embryo transfer was 25.4%in the PCO group and 23.0% in the group with normal ovaries.There were three high order multiple pregnancies in the PCOgroup compared with none in the group with normal ovaries. Ofthe PCO patients, 10.5% developed moderate/severe ovarian hyperstimulationsyndrome (OHSS) compared with none of the controls (P = 0.006).Patients with and without PCO undergoing IVF have comparablepregnancy and livebirth rates. However, it is important to diagnosePCO before ovarian stimulation is initiated as these patientsare more likely to develop moderate or severe OHSS following1VF—ET.     

In-vitro maturation of human oocytes

Immature human oocytes can be matured and fertilized in vitro. However, subsequent embryonic development is different when the immature oocytes are retrieved in different situations. Exposure to the LH surge in vivo may be important for the oocytes to acquire the competence for maturation and subsequent embryonic development. The size of the follicles may also be an important feature for subsequent embryonic development. However, the developmental competence of oocytes derived from small antral follicles does not seem to be adversely affected by the presence of a dominant follicle. Oocyte maturation in vitro is profoundly affected by culture conditions. Gonadotrophins are required for oocyte maturation in vivo, but any requirement in vitro is still unclear. Recent clinical results from in-vitro matured (IVM) human oocytes are promising, although further research remains to be done in order to address the mechanisms of oocyte maturation and to improve culture conditions and also the implantation rate of embryos generated from IVM oocytes.     

Current perspective on primordial follicle cryopreservation and culture for reproductive medicine

Mature oocytes are rare and precious cells. A technology which generates larger numbers would be very welcome in clinical practice, animal production technology and research. Since de-novo formation of female germ cells has ceased by the time of birth, the most attractive strategy, in theory, is to harvest and culture primordial follicles, the most abundant stage in the ovary at all ages. So far, there has been more success with cryopreservation of primordial follicles than with culture, and frozen-thawed ovarian tissue grafts have restored fertility to a number of species after oophorectomy. However, in-vitro development of isolated follicles is not sustained beyond the primary follicle stage. To meet their requirements for growth, metabolism and differentiation, a multistage protocol will probably be required for the prolonged period of development to maturity. The mouse is the only model, to date, in which a live offspring has ever been produced after growing follicles completely in vitro. A triple-stage process was required, involving culture of ovarian explants followed by isolation of granulosa-oocyte complexes and, finally, suitable conditions for completing meiotic maturation. Achievement of this goal for the larger and more slowly developing follicles from human and farm animal ovaries is still a remote possibility.     

Observation of pronuclei may not be an absolute indicator for fertilization in rescue intracytoplasmic sperm injection oocytes

This case report describes a successful full-term pregnancy and birth after the transfer of rescue intracytoplasmic sperm injection (ICSI) embryos derived from 1-day-old oocytes. A total of eight oocytes were retrieved and inseminated 3 h after collection. No oocytes were fertilized 16–18 h after insemination. A rescue ICSI was performed on the four metaphase II stage oocytes. At the regular time (16 h after ICSI) of examination for fertilization, there were no distinct two pronuclei (2PN) in the cytoplasm of any oocytes, but the culture of these oocytes led to the development of four two-cell stage embryos 26 h after ICSI. Further culture of these four embryos showed development to a four-cell stage the following day. The transfer of three embryos resulted in a full-term pregnancy with the delivery of a pair of healthy twins. This result suggests that the observation of 2PN at the normal time of fertilization assessment may not appear to be an absolute indicator of fertilization in the case of rescue ICSI. (Reprod Med Biol 2003; 2 : 83–85)     

Chromosome abnormality rates in human embryos obtained from in-vitro maturation and IVF treatment cycles

The aim of this retrospective study was to compare the incidence of chromosomal abnormality in embryos from in-vitro maturation (IVM) and IVF cycles. The copy numbers of chromosomes 13, 15, 16, 18, 21, 22, X and Y were assessed with fluorescence in-situ hybridization (FISH) in single blastomeres biopsied from cleavage stage embryos. Spare embryos that were not transferred or cryopreserved were also analysed in full. IVM and IVF groups comprised six and 30 couples, with mean ± SD embryos with FISH result of 8.0 ± 4.4 and 11.7 ± 3.8, respectively. The incidence of chromosomal abnormality per FISH result was similar in IVM and IVF embryos (58.7% versus 57.4%, respectively). When embryos were categorized based on maturation time of oocytes in IVM cycles, embryos derived from oocytes that matured 48 h after collection had a higher chromosomal abnormality rate compared with embryos derived from in-vivo matured oocytes and to embryos derived from oocytes that matured in the first 24 h after collection.     

Acrosome reactivity in spermatozoa of different morphology in response to stimulation with follicular fluid

The ability of the morphologically abnormal spermatozoon toundergo the processes necessary for fertilization is unknown;one of the essential processes is the acrosome reaction. Inorder to assess this, spermatozoa from 10 known fertile donorswere incubated with either follicular fluid or Earle's mediumcontaining 3 mg/ml bovine serum albumin at 37°C for 6 h.The spermatozoa were then stained with 2% Trypan Blue priorto being fixed in gluteraldehyde and stained for the presenceof the acrosomal cap using the triple stain. Fifty live spermatozoain each of four morphological categories (normal head, largehead, small head, abnormal neck or tail) were examined and thenumber of acrosome-reacted spermatozoa determined. There wasa significant difference between the morphological groups inthe baseline number of acrosome-reacted spermatozoa, determinedby examining sperm samples incubated in Earle's medium; however,the number of spermatozoa undergoing the acrosome reaction inresponse to stimulation with follicular fluid (i.e. the numberof spermatozoa acrosome-reacted in follicular fluid minus thenumber acrosome-reacted in Earle's medium) was similar for allmorphological groups. This suggests that abnormal sperm morphologydid not affect the response of spermatozoa to activation ofthe acrosome reaction by exogenous stimuli.     

Success of intrauterine insemination using cryopreserved donor sperm is related to the age of the woman and the number of preovulatory follicles

Objective: Our objective was to assess parameters associated with a successful outcome of intrauterine insemination (IUI) using cryopreserved donor sperm. Design: We analyzed 750 consecutive donor IUI cycles undertaken by 363 women in an assisted conception clinic. The main outcome measure was clinical pregnancy. Results: IUI was performed in 94.7% of the 750 IUI treatment cycles commenced and 180 clinical pregnancies occurred. The clinical pregnancy rate per cycle was 26.4%. The rate was significantly related to the patient's age (30.5% for age 35 years and 18.1% for age >35 years;P<0.006) and whether there was one or more than one preovulatory follicles [20.9, 34.4, and 31.5% for one, two, and three or four follicles with a mean diameter of 14 or more mm at the time of human chorionic gonadotropin (hCG) administration;P=0.006]. Two to four preovulatory follicles were present in 12.6% of the natural cycles, 43.6% of clomiphene citrate or tamoxifen, and 59.9% of gonadotropin stimulated cycles. The difference in the number of preovulatory follicles between stimulated and unstimulated cycles was highly significant (P<0.0001). Pregnancy rates were 29.9% in gonadotropin-stimulated cycles, 23.6% in clomiphene citrate- or tamoxifen-stimulated cycles, and 20.1% in unstimulated cycles. The difference in pregnancy rates between gonadotropin-stimulated and natural cycles was significant (P=0.038). Cycle fecundity rates were not significantly affected by the number of previous treatment cycles, duration of infertility, gravidity and parity of the patient, presence of a spontaneous luteinizing hormone (LH) surge before the administration of hCG, or number of motile sperm in the insemination specimen. Conclusions: Success of IUI using cryopreserved donor sperm is related to the age of the women and whether there is one or more than one preovulatory follicles.