Cumulative pregnancy and live birth rates in women with antiphospholipid antibodies undergoing assisted reproduction.

The aims of this study were to investigate the influence of antiphospholipid antibodies (APA) on cumulative pregnancy and live-birth rates in patients undergoing assisted reproductive treatment. Serum samples from 173 patients were collected prior to initiation treatment cycle and tested by enzyme-linked immunosorbent assay (ELISA) for the presence of immunoglobulin (Ig)G, IgM and IgA against cardiolipin, phosphoserine, phosphoethanolamine, phosphoinositol, phosphatidic acid, and phosphoglycerol. Fifty-six samples from patients who had at least two failed cycles by assisted reproductive treatment were also tested by a bioassay for the presence of lupus anticoagulants.

Sugars exert a major influence on the vitrification properties of ethylene glycol-based solutions and have low toxicity to embryos and oocytes.

A systematic approach was taken to assess the vitrification properties of ethylene glycol-based solutions supplemented with carbohydrates. Solutions were prepared by weight (gravimetrically) using ethylene glycol as the cryoprotectant, 0.9% NaCl in water, and six different sugars: d-glucose, d(-)-fructose, d-sorbitol, sucrose, d(+)-trehalose, and raffinose.

The mechanism of action of epidermal growth factor and transforming growth factor alpha on aromatase activity in granulosa cells from polycystic ovaries.

We investigated aromatization and the mechanism of action of epidermal growth factor (EGF) and transforming growth factor alpha (TGFalpha) on oestradiol biosynthesis in freshly prepared granulosa cells from polycystic ovaries. Freshly prepared granulosa cells from polycystic ovaries incubated for only 3 h under basal conditions secreted significantly (P< 0.001) greater amounts of oestradiol-17beta than that of granulosa cells from normal ovaries.

Factors affecting the success of human blastocyst development and pregnancy following in vitro fertilization and embryo transfer.

Objective: To determine the factors affecting blastocyst development and pregnancy after IVF and ET.

Design: Retrospective analysis of data arising from a clinical trial.

Setting: Private in vitro fertilization clinic.

Oocyte maturation.

Primary oocytes recovered from small and growing follicles of > or = 3 mm in the ovaries of untreated women, can be matured in vitro, will fertilize and develop in vitro, and when transferred to the patient, develop to term. However, the implantation rate of cleaved embryos has been disappointingly low and when embryos are allowed to develop beyond the 4-cell in vitro, retardation of development and blockage is frequently observed, with relatively few embryos developing to blastocysts. We have devised new culture systems for human embryos to enable high rates of development of in-vivo matured oocytes to blastocysts within 5-6 days of culture, and high implantation rates of these blastocysts when they are transferred to the patients' uterus.

Potential benefits of cell cloning for human medicine.

The successful cloning of a mammal from an adult somatic cell nucleus opens new avenues for major advances in reproductive medicine, biotechnology and cellular-based transplantation therapies for degenerative diseases. At the same time, this breakthrough has generated much heated discussion concerning the ethics of cloning. Twinning is a form of cloning, and there are instances in clinical assisted reproduction in which the deliberate formation of twins by embryo dissection would seem ethically acceptable.

The effect of recipient oocyte volume on nuclear transfer in cattle.

This study compared the developmental potential of bovine nuclear transfer embryos with varying amounts of cytoplasm. Embryos formed from single cytoplasts fused to blastomeres by a single electrical pulse or from double cytoplasts using a double electrical pulse resulted in reconstituted embryos containing 75% and 150% of the original oocyte volume. No differences in fusion, cleavage, or development rates to blastocysts were observed between the groups.

Evolution of a culture protocol for successful blastocyst development and pregnancy.

A cell-free culture system was designed for human embryo development to the blastocyst stage by testing a range of culture conditions in a series of protocols. The culture system that was evolved has a brief 1 h exposure to spermatozoa and then culture of the pronucleate zygote for 2 days in IVF-50 medium. Two or three embryos were cultured together in 20 microl microdrops of medium under oil.

The stage-specific expression of TEC-1, -2, -3, and -4 antigens on bovine preimplantation embryos.

The preimplantation developmental period is associated with constant changes within the embryo, and some of these changes are apparent on the embryo cell surface. For example, during transition from maternal to embryonic genome control and the compaction and differentiation of embryonic cells, the cell surface undergoes morphologic alterations that reflect changes in gene control. In order to gain insight into the events occurring during embryonic development and cellular differentiation, monoclonal antibodies specific for cell surface antigens (TEC antigens) of embryonic cells have been generated previously and shown to recognise either the carbohydrate moiety of embryoglycan or a developmentally regulated protein epitope.

Constitutive and virus-induced interferon production by peripheral blood leukocytes.

The production of interferon-alpha (IFN-alpha) by normal human peripheral blood mononuclear cells (PBMNC) was studied using polyclonal antipeptide antibodies designed to react either with all IFN-alpha subtypes or with individual subtypes IFN-alpha 2 or IFN-alpha 4. In this study, we demonstrate the detection of intracellular IFN-alpha in PBMNC using immunofluorescence staining and flow-cytometric analysis. Virtually all cells of the PBMNC population were shown to produce IFN-alpha reactive with all three antisera after stimulation with Sendai virus.

Evaluation of propanediol, ethylene glycol, sucrose and antifreeze proteins on the survival of slow-cooled mouse pronuclear and 4-cell embryos.

Mouse pronuclear and 4-cell embryos were cryopreserved by slow cooling to -33 degrees C in 1.5 M 1,2-propanediol or 1.5 M ethylene glycol, with or without 0.

Expression of collagen alpha 1(IV), laminin and nidogen genes in the embryonic mouse lung: implications for branching morphogenesis.

The patterns of laminin A, B1, B2, nidogen and collagen alpha 1(IV) gene expression in the embryonic mouse lung were determined using in situ hybridization histochemistry at a stage when branching morphogenesis is taking place. Collagen alpha 1(IV), laminin B1 and B2 genes were expressed throughout the mesenchyme and epithelium. Nidogen gene expression was uniform throughout the mesenchyme but was not detected in epithelial cells.

Nicotinamide, a component of complex culture media, inhibits mouse embryo development in vitro and reduces subsequent developmental potential after transfer.

Objective: To determine the effects of B-group vitamins present in culture media on mouse embryo development in vitro and subsequent viability.

Design: Mouse zygotes were cultured in the presence of B-group vitamins. Embryo morphology and cell numbers were determined at 96 and 120 hours after hCG.

Enhanced rates of cleavage and development for sheep zygotes cultured to the blastocyst stage in vitro in the absence of serum and somatic cells: amino acids, vitamins, and culturing embryos in groups stimulate development.

The aim of this study was to develop a serum-free culture system that could support high levels of cleavage and blastocyst formation from sheep zygotes developed in vitro. To this end, we investigated the effects on sheep zygote development of amino acids, ammonium, vitamins, and culture of embryos in groups in Synthetic Oviduct Fluid (SOF) medium supplemented with BSA (32 mg/ml). The inclusion of amino acids in the culture medium had no effect on the percentage of embryos arrested at the 8-16-cell stage when embryos were cultured singly in the same drop of medium for 6 days (43% in SOF; 41% in SOF+amino acids).

The choice of the most appropriate microfertilization technique for human male factor infertility.

Comparisons were made among techniques used to treat male factor infertility. Patients with semen quality below that recognized by World Health Organization criteria as normal had a better success rate when treated by gamete intrafallopian transfer than by in vitro fertilization (25% v. 7% pregnancy rate per patient).

Embryo development capacity of oocytes fertilized by immature sperm and sperm treated with motility stimulants.

The fertilizing ability of mouse spermatozoa develops during maturation and coincides with the acquisition of motility. The lack of progressive motility of spermatozoa from the testis and precaudal segments of the epididymis interferes with their ability to fertilize oocytes after insemination in vitro. The removal of cumulus cells for insemination in vitro and the use of subzonal injection of a single spermatozoon resulted in a higher number of oocytes fertilized by immature caput and corpus spermatozoa.

Early sperm-egg interaction after sperm microinjection.

The early events of sperm-egg interaction occurring 1-3 h after multiple sperm injection into the perivitelline space (PVS) and after direct injection into the ooplasm of human oocytes are reported. The sperm acrosome reaction occurred in the PVS but was not detected within the ooplasm. Sperm in the PVS were incorporated into the ooplasm in the usual manner described in vitro, after completion of the acrosome reaction, and a block to polyspermy was evident at the oolemma.

Mouse testis Pdha-2 promoter upstream sequences confer tissue-and temporal-specific activity in transgenic mice.

Spermatogenesis is a complex process requiring the coordinate expression of a number of testis-specific genes. One of these, Pdha-2, codes for the murine testis-specific isoform of the E1 alpha subunit of the pyruvate dehydrogenase complex. To elucidate the mechanisms regulating its expression in vivo, we have begun to investigate the Pdha-2 promoter in transgenic mice.

Effects of stimulation or inhibition of lipid peroxidation on freezing-thawing of mouse embryos.

This study was conducted to determine whether the tolerance of embryos to the stress of freezing and thawing can be modified by including in the incubation medium stimulators or inhibitors of membrane lipid peroxidation. Mouse zygotes were cultured in medium M16, supplemented or not with FeSO4, apotransferrin, and/or ascorbate. In each culture supplement, 8-cell embryos were randomly allocated to an untreated (nonfrozen) control group, or treatment by freezing using slow or ultra-rapid cooling.

Expression of laminin and nidogen genes during the postimplantation development of the mouse placenta.

The expression patterns of laminin A, B1, B2, and nidogen genes were identified by in situ hybridization in postimplantation mouse extraembryonic tissues and maternal decidua during the period when the chorioallantoic placenta is established. Laminin and nidogen genes were not coordinately expressed either in the decidua or in trophoblast cells, indicating that these genes are regulated independently in these cell types during the establishment of the placenta. Laminin A mRNA was absent from the decidua except in the outer layer of cells adjacent to the myometrium and in the central decidual zone adjacent to the remnant of the uterine epithelium on Day 9.

Does the zona pellucida select spermatozoa from the medium with higher fertilizing potential?

The present study was carried out to test whether the zona pellucida selects spermatozoa with higher fertilization potential. Fertilization rates of mouse oocytes after sperm microinjection under the zona pellucida (SMUZ) of zona-bound spermatozoa and of spermatozoa incubated in the absence of oocytes and treated (acid-treated group) or not (control group) with acid Tyrode's solution were compared. SMUZ was performed at 15, 30, 60, and 90 min after the insemination of fresh oocytes required for selecting spermatozoa bound to the zona pellucida.

Uptake and metabolism of pyruvate and glucose by individual sheep preattachment embryos developed in vivo.

The uptake of pyruvate and glucose by individual sheep oocytes and preattachment sheep embryos at each state of development up to the hatching blastocyst was determined using a microfluorescence technique. After an initial increase at fertilization, pyruvate uptake was relatively constant (approximately 15 pmol/embryo/h) from the zygote through to the morula. Upon blastocyst formation and hatching, there were significant increases in uptake (39 pmol/embryo/h, P < 0.

Differential expression of laminin, nidogen and collagen IV genes in the midgestation mouse placenta.

The distribution of laminin A, B1, B2, nidogen and collagen alpha 1(IV) mRNA was studied in the 12.5-day mouse placenta and uterus. This was compared to the pattern of laminin, nidogen and collagen IV immunoreactivity in the placenta at this time.