Clinical Assisted Reproduction: Gonadotropin-Releasing Hormone Messenger Ribonucleic Acid and Protein Expression in Vero Cells

摘要

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.