John McBain

Preimplantation aneuploidy screening using comparative genomic hybridization or fluorescence in situ hybridization of embryos from patients with recurrent implantation failure

OBJECTIVE To select chromosomally euploid embryos for transfer by analyzing single biopsied blastomeres using either fluorescence in situ hybridization (FISH) for chromosomes 13, 16, 18, 21, and 22 or comparative genomic hybridization (CGH), which provides a full karyotype. DESIGN Prospective observational study. SETTING A large IVF unit and the research laboratory of a hospital clinical genetics unit. PATIENT(S) Twenty patients with recurrent implantation failure. INTERVENTION(S) Ovarian stimulation and IVF by intracytoplasmic sperm injection (ICSI), embryo biopsy, and embryo transfer. MAIN OUTCOME MEASURE(S) Chromosome normality of biopsied blastomeres and implantation and clinical pregnancy rates. RESULT(S) Comparative genomic hybridization was able to identify many chromosomal abnormalities that would have been missed if those cells had been analyzed by FISH. The clinical pregnancy rate per transfer and implantation rate was 11% and 7% for embryos analyzed by FISH and 21% and 15% for embryos analyzed by CGH. CONCLUSION(S) Comparative genomic hybridization is more effective than FISH for identifying chromosomally normal embryos, which may result in a higher clinical pregnancy rate and implantation rate after embryo transfer.

Factors affecting low birthweight after assisted reproduction technology: difference between transfer of fresh and cryopreserved embryos suggests an adverse effect of oocyte collection

BACKGROUND Data show that differences exist in the birthweight of singletons after frozen embryo transfer (FET) compared with fresh transfer or gamete intra-Fallopian transfer (GIFT). Factors associated with low birthweight (LBW) after assisted reproduction technology (ART) were studied. METHODS Birthweight, distribution of birthweight, z-score, LBW (<2500 g), gestation and percentage preterm (<37 weeks) for singleton births >19 weeks gestation, conceived by ART or non-ART treatments (ovulation induction and artificial insemination) between 1978 and 2005 were analysed for one large Australian clinic. RESULTS For first births, the mean birthweight was significantly (P < 0.005) lower, and LBW and preterm birth more frequent for GIFT (mean = 3133 g, SD = 549, n = 109, LBW = 10.9% and preterm = 10.0%), IVF (3166, 676, 1615, 11.7, 12.5) and ICSI (3206, 697, 1472, 11.5, 11.9) than for FET (3352, 615, 2383, 6.5, 9.2) and non-ART conceptions (3341, 634, 940, 7.1, 8.6). Regression modelling showed ART treatment before 1993 and fresh embryo transfer were negatively related to birthweight after including other covariates: gestation, male sex, parity, birth defects, Caesarean section, perinatal death and socio-economic status. CONCLUSIONS Birthweights were lower and LBW rates higher after GIFT or fresh embryo transfer than after FET. Results for FET were similar to those for non-ART conceptions. This suggests IVF and ICSI laboratory procedures affecting the embryos are not causal but other factors operating in the woman, perhaps associated with oocyte collection itself, which affect endometrial receptivity, implantation or early pregnancy, may be responsible for LBW with ART.

The Ultrastructure of the Preovulatory Human Egg Fertilized In Vitro

Preovulatory eggs in cumulus were inseminated in vitro with washed spermatozoa which had been preincubated for 1.5 hours. After 3 hours, three eggs were processed for electron microscopy and each was sectioned serially from pole to pole. In the two eggs which had been fertilized, the expanded chromatin of the fertilizing sperm head and the chromatin of the ovum were almost completely surrounded by a developing pronuclear envelope. In one of the penetrated eggs the developing male pronucleus and associated midpiece and sperm tail were located within an incorporation cone. The surface of the cone was free of microvilli but contained a zone of microfilaments immediately beneath the plasma membrane. A similar zone of microfilaments was present beneath the advancing surface of the extruding second polar body (PB2) which was connected to the ovum by an interbody and microtubules of the meiotic spindle. Cortical granules were completely absent from the fertilized eggs but were present in the unfertilized egg. PB2 contained a nucleus at a stage of development similar to that of the early pronuclei.

Serum supplement in human in vitro fertilization and embryo development

The effects of maternal preovulatory serum and human fetal cord serum supplement in culture medium in human in vitro fertilization and embryo development were compared in 208 cases over an 8-month period when there were no significant changes in other variables. A significantly higher pregnancy rate was observed in the fetal cord serum group despite no significant difference being found in fertilization and embryo cleavage rates. This suggests a difference in the health of the embryo cultured in different serum supplements.

AN UNEXPECTEDLY HIGH RATE OF ECTOPIC PREGNANCY FOLLOWING THE INDUCTION OF OVULATION WITH HUMAN PITUITARY AND CHORIONIC GONADOTROPHIN

Six tubal ectopic pregnancies occurred in a series of 193 pregnancies following ovulation induced with human pituitary gonadotrophin (hPG) and human chorionic gonadotrophin (hCG). The ectopic pregnancy rate of 3.1 per cent is higher than quoted incidences in the general population and occurred in the absence of predisposing factors. There was an association between ectopic pregnancy and elevated urinary oestrogen excretion in the peri‐ovulatory phases of the induced ovulatory cycles. A urinary oestrogen excretion of greater than 200 μg/24 hours on day 0 (the day after hCG was given) was associated with a 10 per cent chance of ectopic pregnancy (P <0.05).

A laparoscopic approach to a program of gamete intrafallopian transfer

Gamete intrafallopian transfer (GIFT) provides an effective method of achieving pregnancy in infertile women with normal fallopian tubes. Laparoscopic approach to ovum pickup and tubal catheterization provides a simple and rapid means of performing the operation. Equipment used to facilitate this process is described, and techniques of tubal catheterization are discussed. A clinical pregnancy rate of 27% is reported in a series of 71 treatment cycles. The application of GIFT in conjunction with in vitro fertilization is discussed, especially the use of excess gametes to provide embryos for freezing. The use of GIFT as a research and investigative tool may provide further insight into the causes for idiopathic infertility.

Detection of Antispermatozoal Antibodies of IgA Class in Cervical Mucus

ABSTRACT: A simple procedure for detection of antisperm antibodies of IgA class in human cervical mucus is described and the results of its application to samples from 102 patients are presented. The results suggest that the IgA immunobead test (IgA‐IBT) is a specific and clinically useful test for sperm antibodies. There was a strong correlation between the IgA‐IBT and the presence of complement‐dependent sperm immobilization in serum (Spearmans, r = 0.92, p < 0.001). Positive IgA‐IBT results occurred only in mucus samples that showed poor penetration by normal sperm. An added advantage of the IgA‐IBT is that both the immunoglobulin class and the site of binding to the sperm surface can be determined simultaneously.

The developmental potential of cryopreserved human embryos.

Using rigorously matched non-frozen controls we have shown that cryopreservation does not alter the implantation potential of early cleavage stage (day 2) human embryos if no blastomere loss occurs. Thawed intact 4-cell embryos have a significantly higher implantation (fetal heart) rate (16.9%) than similar 2-cell embryos (7.2%). This difference is not due to blastomere number per se since increasing the cell number in frozen embryos by allowing an extended period in culture prior to freezing does not alter their intrinsic developmental potential. Blastomere loss, which occurred in almost half of all thawed embryos, is directly related to a reduction in developmental potential. We estimate that approximately 30% of the expected fresh embryo implantations are lost as a consequence of cryopreservation. Both preimplantation and peri-implantation losses may contribute to this outcome.

Performance of patients with a “frozen pelvis” in an in vitro fertilization program

It is now possible to identify and study the performance of different subgroups of patients in in vitro fertilization (IVF) programs. Patients with severe pelvic adhesions due to pelvic inflammatory disease (PID) or endometriosis were classed as having a frozen pelvis if less than or equal to 20% of total ovarian surface was visible and if the rest of the ovary was bound down with significant adhesions. IVF offers the only hope of pregnancy for these patients. Fifty-one treatment cycles in 23 such patients were matched against 51 cycles in 48 patients with adhesion-free ovaries. The study group had a significantly higher number of cancelled oocyte retrievals because of poor estradiol (E2) response. They also had a significantly lower rate of E2 rise and a lower peak value of E2 before and after the administration of human chorionic gonadotropin. These patients took longer to respond to a hyperstimulation regime, and when a response occurred they formed fewer follicles, as measured with the use of ultrasound. Lower numbers of oocytes were obtained from this group, but the fertilization rate of oocytes was the same for both groups. One pregnancy occurred in the study group and 11 in the control group. It is possible that disruption of ovarian blood supply or mechanical factors due to the pressure of significant adhesions prevent a good follicular response in patients with a frozen pelvis.

Unexplained infertility: a review

Summary. When investigations fail to reveal a cause for infertility, treatment must then be based on possible, but unproven, causes, and since there is a high spontaneous pregnancy rate in unexplained infertility the effect of any treatment is difficult to assess. Such treatment has included correction of anatomical variants such as uterine retroversion and the use of hormonal manipulation during the follicular and luteal phases of the menstrual cycle. Ovum entrapment, occult spontaneous abortion and faults in sperm fertilizing capacity have all been implicated, and it is likely that immunological factors play a substantial role in unexplained infertility. Evidence does not support the use of bromocriptine in the absence of hyperprolactinaemia. Successful treatment by intrauterine insemination is unlikely if there are circulating anti‐sperm antibodies in the partner. Improving cervical mucus by treatment with oestrogens and clearing infections with antibiotics may have a modest place but it is very difficult to show that these treatments have more than a placebo effect. Endometriosis is often missed and the possibility of it having developed after initial investigation warrants repeat laparoscopy after 2 years. Three approaches are currently acceptable in the management of the couple with unexplained infertility: await spontaneous pregnancy, the empirical use of clomiphene and in‐vitro fertilization.