Jan S. Kruessel

The Role of Gonadotropin-Releasing Hormone in Murine Preimplantation Embryonic Development

Previous studies have established the presence of an extrahypothalamic GnRH in a variety of tissues. GnRH receptor is known to be present in the placenta, which produces and secretes the decapeptide from the very early stages of placentation. We hypothesized that GnRH may play a role in the preimplantation development of embryos. To examine this hypothesis, we assessed GnRH and GnRH receptor messenger RNA (mRNA; RT-PCR) and protein expression (Immunohistochemistry) in preimplantation murine embryos at various developmental stages. Furthermore, preimplantation murine embryos were cultured with GnRH agonist and antagonist in vitro to assess the influence of GnRH analogs on embryo development. GnRH is expressed in the developing mouse embryo from morula to hatching blastocyst stages at the mRNA and protein levels. GnRH receptor mRNA is also present in the developing embryos studied. Preimplantation embryonic development was significantly enhanced by incubation with increasing concentrations of GnRH agonist a...

Different pattern of interleukin-1β-(IL-1β), interleukin-1 receptor antagonist- (IL-1ra) and interleukin-1 receptor type I- (IL-1R tI) mRNA-expression in single preimplantation mouse embryos at various developmental stages1

The interleukin-1 (IL-1) system has been shown to play an important role in human and murine embryo implantation. Recent studies have documented immunohistochemical evidence of interleukin-1 beta (IL--1 beta), interleukin-1 receptor antagonist (IL-1ra) and interleukin-1 receptor type I (IL-1R tI) in human preimplantation embryos and protein levels of interleukin-1 alpha (IL-1 alpha), IL-1 beta and IL1ra in human preimplantation embryo culture fluid have been correlated with successful implantation and pregnancy. Our aim in this study was to detect IL-1 beta, Il-1ra and Il-1R tI mRNA in single preimplantation mouse embryos and to describe the frequency of positive mRNA-expression at different developmental stages. B6C3F1-mice, 12 weeks old were pregnant mare serum gonadotropin (PMSG)/human chorionic gonadotropin (hCG)-stimulated and mated. Animals were sacrificed at day 0.5, and zygotes were flushed from the tubes and cultured in HAMs-F10 medium. 2-cell- (2C-), 8-cell- (8C-), morula- (M-), early blastocyst- (EB-) and hatching blastocyst- (HB-) stage embryos were examined by one round of reverse transcriptase (RT) followed by two rounds of polymerase chain reaction (PCR) carried out on individual mouse embryos for beta-actin (internal standard), IL-1 beta, IL-1ra and IL-1R tI-mRNAs. The frequencies of positive mRNA-expressions were as follows (2C/8C/M/EB/HB); beta-actin: 91/96/100/100/98%; IL-1b: 0/0/2.5/6.25/19; IL-1ra; 0/5/30/41/74% and IL-1R tI: 0/0/10/20/25%. The incidence of IL-1ra mRNA expression increased with developmental stage. IL-1ra mRNA seems to be expressed in a very high percentage (74%) of embryos near the time of implantation, whereas the percentage of IL-1 beta-mRNA positive embryos is surprisingly low (19%).

Immunoreactive gonadotropin-releasing hormone expression in cycling human endometrium of fertile patients.

OBJECTIVE To investigate the protein expression of GnRH in the endometrium of fertile patients throughout the menstrual cycle. DESIGN Prospective longitudinal study. SETTING Department of Gynecology and Obstetrics, Reproductive Immunology Laboratory, Stanford University Medical Center. PATIENT(S) Twenty-two fertile premenopausal women submitted to laparoscopic surgery for benign gynecologic indications. None of the 22 women had endometriosis or pelvic inflammatory disease. INTERVENTION(S) An endometrial biopsy specimen using the Novak curette was obtained at the time of surgery. MAIN OUTCOME MEASURE(S) Protein expression and localization from unfractioned endometrial tissue was analyzed by immunohistochemistry. RESULT(S) Gonadotropin-releasing hormone is expressed at the protein level in both the endometrial stroma and epithelium throughout the entire menstrual cycle of fertile women. Immunostaining in the human epithelium reached maximal levels in the midluteal phase and was elevated in the stroma throughout the entire luteal phase. CONCLUSION(S) Our results demonstrate the presence of GnRH in the human endometrium at the protein level throughout the entire menstrual cycle of fertile women, with an increase in the luteal phase compared with the preovulatory endometrium.

Genital schistosomiasis as a cause of female sterility and acute abdomen.

OBJECTIVE To report the case of a Nigerian patient who suffered from sterility and underwent abdominal laparotomy to remove an adnexal tumor. Histologic analysis revealed Schistosoma haematobium ova. DESIGN Case report. SETTING Multidisciplinary group practice and teaching hospital. PATIENTS One patient who underwent an abdominal laparotomy to remove an adnexal tumor. INTERVENTION(S) In vitro fertilization and adnexal tumor resection via laparotomy. MAIN OUTCOME MEASURE(S) Treatment of the adnexal tumor affecting the reproductive health. RESULT(S) Histologic analysis performed on the tissue removed at surgery revealed fibrous patches around aggregates of calcified Schistosoma haematobium eggs in the fallopian tube wall. CONCLUSION(S) Schistosomiasis needs to be considered as a differential diagnosis of female infertility and sterility.

Interleukin-1β regulation of gonadotropin-releasing hormone messenger ribonucleic acid in cultured human endometrial stromal cells

Abstract Objective To investigate the role of interleukin-1β (IL-1β) in regulating GnRH mRNA expression in cultured human endometrial stromal cells using a modified semiquantitative competitive reverse transcription and polymerase chain reaction (PCR). Design A controlled study. Setting Clinical and academic research setting in a university medical center. Patient(s) Women undergoing hysterectomy for nonmalignant indications. Intervention(s) Confluent stromal cell cultures treated with steroid hormones were stimulated with IL-1β and attenuated by anti-IL-1β antibody or IL-1 receptor antagonist. Main outcome measure(s) The human endometrial stromal cell expression of GnRH and its receptor were determined by PCR. Interleukin-1β-mediated regulation of stromal cell GnRH mRNA expression was determined by quantitative competitive PCR. Result(s) The GnRH and GnRH receptor mRNA expression were amplified in cultured stromal cells by PCR and two rounds of nested PCR, respectively. Treatment with IL-1β stimulated stromal cell GnRH mRNA expression at concentrations of IL-1β above 10 IU/mL. Recombinant IL-1 receptor antagonist and anti-IL-1β antibody attenuated the increase of gene expression of GnRH initiated by IL-1β. Conclusion(s) These results provide indirect evidence that IL-1β may play a crucial role at the level of embryo–maternal interaction by regulating stromal cell expression of GnRH and its receptor, both known to be important in mediating trophoblast invasion and placental hormone regulation.

Gonadotropin-releasing hormone messenger ribonucleic acid and protein expression in Vero cells.

Purpose: Interleukin-1 (IL-1) is a major regulator of local cellular interactions during embryonic implantation. We hypothesized that gonadotropin-releasing hormone (GnRH) may also play a role in the embryonic/epithelial dialogue during early implantation. To examine this hypothesis, we examined the ability of IL-1 to regulate GnRH mRNA and protein expression in Vero cells.Methods: Viable Vero cells (1 × 105/well) were cultured in multiple-well tissue culture plates for in vitro studies and in 4-well chamber slides for immunohistochemical study. Confluent Vero cells were cultured with increasing concentrations of recombinant human IL-1β for an additional 24 hr. Vero cell expression of GnRH and GnRH receptor mRNAs was measured with polymerase chain reaction (PCR) and nested PCR, respectively. GnRH protein expression was validated by immunohistochemistry study. The quantitative level of GnRH mRNA expression regulated by IL-1β in Vero cells was determined by quantitative competitive PCR (QC PCR) with standard curve methodology.Results: RT-PCR revealed β-actin, GnRH, and GnRH receptor mRNA expression in Vero cell cultures. Immunostaining confirmed the presence of GnRH protein in Vero cells. Quantitative PCR demonstrated IL-1β up-regulation of Vero cell GnRH mRNA expression (p < 0.05).Conclusions: These results suggest that Vero cell mRNA and protein expression of GnRH may play a substantial role in early embryo/epithelial dialogue during embryo coculture, with an embryotrophic effect due to expression of GnRH by Vero cells.