F. M. Fusi

Premature ovarian failure: steroid synthesis and autoimmunity.

The androgen biosynthesis and autoimmunity of 25 patients with premature ovarian failure (POF) and 18 control subjects with normal cycles were examined. Serum levels of dehydroepiandrosterone sulfate (DHEAS), 17-hydroxyprogesterone (17-OHP), androstenedione, and testosterone were analyzed in POF patients with or without organ-specific autoimmunity, and the results compared with those of women with normal ovarian function. The comparative analysis of DHEAS, 17-OHP, androstenedione and testosterone showed that POF patients had significantly lower values than normal women (DHEAS, androstenedione and testosterone p < 0.01, 17-OHP p < 0.05). Furthermore, we found one or more organ-specific autoantibodies in 11 patients with POF (44%), while only one woman in the control group showed autoimmunity (antithyroid microsome) (5.5%). Only one patient had both anti-ovarian and anti-adrenal antibodies (4%). The comparison of androgen levels in POF patients with or without autoimmunity revealed a statistically significant reduction of DHEAS levels in POF patients with organ-specific autoimmunity (p < 0.01). These data reveal the reduction in androgen synthesis in POF patients, particularly in those with organ-specific autoimmunity.

In controlled ovarian hyperstimulation, steroid production, oocyte retrieval, and pregnancy rate correlate with gene expression of vascular endothelial growth factor.

Purpose: Whether the gene expression of vascular endothelial growth factor (VEGF) in human granulosa cells is a predictor of fertilization was evaluated in patients participating in an in vitro fertilization program.Methods: Fifty patients with normal ovaries who were participating in an in vitro fertilization program at the University of Milan, San Raffaele Scientific Institute, were included in the study. We correlated E2and P serum levels on the day of oocyte collection, the number of follicles, oocytes collected, and fertilized, and pregnancies with mRNA for VEGF of luteinizing granulosa cells obtained at the time of oocyte retrieval.Results: Comparing E2and P serum levels, the number of follicles, oocytes collected and fertilized, and pregnancies with gene expression for VEGF, we found a positive correlation. E2and P serum levels were higher in patients with increased VEGF (P < 0.01). Furthermore, there were more follicles, oocytes collected and fertilized, and pregnancies in patients with maximum expression of VEGF, and the difference was statistically significant (P < 0.05).Conclusions: Our results suggest that VEGF may be important for vascular development during follicular growth and luteal differentiation, oocyte maturation, and fertilization.

Soluble Intercellular Adhesion Molecule-1 in Ovarian Follicles: Production by Granulosa Luteal Cells and Levels in Follicular Fluid

OBJECTIVEnTo determine the concentration of the soluble form of intercellular adhesion molecule (ICAM)-1 in granulosa luteal cell-conditioned media and in follicular fluid (FF).nnnDESIGNnGranulosa cells and FF samples were obtained at the time of oocyte retrieval for IVF. In 10 women, a total of 33 fluids were obtained from individual follicles, whereas in 70 women, the follicular aspirates were pooled.nnnSETTINGnClinica L. Mangiagalli and Reproductive Center, San Raffaele Hospital, Milan, Italy.nnnPATIENT(S)nEighty women referred for IVF for tubal factor or male factor infertility.nnnINTERVENTION(S)nWomen underwent ovarian hyperstimulation.nnnMAIN OUTCOME MEASURE(S)nSoluble ICAM-1 was measured by an ELISA, and its levels were correlated with follicular size, the number of retrieved oocytes, and the number of follicles with a diameter of >15 mm.nnnRESULT(S)nThe concentration of soluble ICAM-1 in granulosa luteal cell-conditioned media was 17.8 +/- 1.8 ng/5 x 10(5) cells. Interleukin-1beta can stimulate soluble ICAM-1 release in a dose-dependent manner. A significant positive correlation was demonstrated between levels of soluble ICAM-1 in pooled FF and the number of retrieved oocytes or the number of follicles with a diameter of >15 mm.nnnCONCLUSION(S)nSoluble ICAM-1 can be released by granulosa luteal cells and can be detected in FF after ovarian hyperstimulation. Levels of soluble ICAM-1 in FF correlate directly with some indices of ovarian function.

Success in inducing ovulation in a case of premature ovarian failure using growth hormone-releasing hormone

Ovulation was obtained in a 29-year-old woman affected by premature ovarian failure who had previously failed to respond to two attempts performed administering human menopausal gonadotropin or follicle-stimulating hormone after the spontaneous gonadotropin production was suppressed using a gonadotropin-releasing hormone analog (buserelin). Induction of ovulation succeeded when 1000 mg/day growth hormone-releasing hormone was added to the induction scheme. Five mature follicles were obtained after 27 days therapy and the serum level of 17 beta-estradiol was 975 pg/ml (195 pg/ml per follicle) at the time of human chorionic gonadotropin administration.

Effects of the coculture with human endometrial cells on the function of spermatozoa from subfertile men

Objective To evaluate the effects of a coculture with human endometrial cells on the function of spermatozoa from samples obtained from infertile couples. Design In a prospective study, human spermatozoa selected by swim-up from fresh samples were cultured on human endometrial feeder layers. Thereafter, their viability, motility, acrosome integrity, and ability to penetrate zona-free hamster oocytes were evaluated. Spermatozoa from the same samples incubated under the same conditions but in the absence of endometrial cells, as well as in the medium previously spent for cell culture, were used as controls. Setting Andrology Laboratory of the Infertility Center of San Raffaele Hospital. Patients Spermatozoa were obtained from 17 infertile men attending the Infertility Center at our hospital. Results Spermatozoa incubated in the presence of endometrial cell feeder layers did not differ from controls with regard to their viability or motility. Conversely, the percent spontaneous acrosome reactions after 18hours of incubation was significantly higher for spermatozoa cocultured (19.7±2.2 versus 11.2±1.9; mean±SE). The mean number of spermatozoa penetrating hamster oocytes was also significantly improved (1.24±0.3 versus 0.68±0.24). This effect did not seem to be solely due to the secretion of soluble factors by endometrial cells in the medium, in that spermatozoa incubated in the medium spent for endometrial cell culture had a similar acrosome reaction percentage but a lower rate of hamster egg penetration. Conclusions The coculture with human endometrial cells appeared to be beneficial for improving the sperm function. This effect partially may be due to the secretion of steroids in the medium, which increases the quota of spontaneous acrosome reaction and in part due to the direct contact of cells with spermatozoa, maybe for the detoxification of the medium or the release of trophic factors. Coculture might be a promising approach to preparing spermatozoa for assisted fertilization in cases of subfertile males.

The use of Matrigel∗ at low concentration enhances in vitro blastocyst formation and hatching in a mouse embryo model ∗

OBJECTIVEnTo determine the effect of Matrigel at a low concentration on the growth of mouse embryos in culture.nnnDESIGNnRandomized case-control study of mouse embryos.nnnSETTINGnAn academic research environment.nnnANIMALSnMouse embryos.nnnINTERVENTION(S)nEmbryos were cultured in Quinns or Celbios human tubal fluid (HTF) enriched with 1.5% bovine serum albumin and 0.8% liquid Matrigel. Each HTF was compared with the same medium devoid of Matrigel. Afterward, Quinns and Celbios HTF, both containing Matrigel, were compared directly. Embryos were cultured in four-well dishes, and their morphology and viability were assessed at 96 hours.nnnMAIN OUTCOME MEASURE(S)nLevel of interleukin-1alpha in media collected at the end of culture.nnnRESULT(S)nIn both types of HTF, the presence of Matrigel allowed a larger number of embryos to reach the blastocyst stage and to hatch; blastocyst morphology also was improved. These positive effects were enhanced in Quinns HTF: embryos cultured in its Matrigel-enriched version secreted a higher level of interleukin-1alpha than those in Celbios HTF plus Matrigel and also showed a better morphology.nnnCONCLUSION(S)nIn the mouse embryo model, Matrigel improves culture conditions in terms of both embryo viability and morphology, and these effects are enhanced in Quinns HTF.

Immunomagnetic separation of antibody-labelled from antibody-free human spermatozoa as a treatment for immunologic infertility. A preliminary report.

Summary. A method is described where superparamagnetic polymer microspheres coated with monoclonal antibodies are used to isolate antibody—labelled from antibody‐free spermatozoa in male autoimmune infertility. Autoimmune sperm samples or antibody‐free spermatozoa adsorbed with antisperm‐antibodies from sera were incubated with microspheres coated with a specific monoclonal antibody to murine immunoglobulins, after their preincubation with mouse anti‐human IgG and IgA. Using a magnet, the microsphere‐labelled spermatozoa were separated from the samples. Immunobead binding was performed before and after the treatment in order to detect changes in the percentage of antibody‐bound spermatozoa. After the immunomagnetic separation, approximately 50% of the IgA‐labelled spermatozoa was isolated while no difference was demonstrated when antisperm antibodies of IgG class were involved. The evaluation of sperm motility and membrane integrity after treatment seemed to indicate that the technique did not have any relevant effect on sperm characteristics. The fact that only a partial success in separation of IgA‐bound spermatozoa and no success for IgG‐labelled sperm was obtained indicates that the method needs to be improved before its clinical utilization might be postulated.