GnRH regulates trophoblast invasion via RUNX2-mediated MMP2/9 expression

Abstract

Study hypothesis: We hypothesized that Runt-related transcription factor 2 (RUNX2), matrix metalloproteinase (MMP)2 and MMP9 are involved in basal and gonadotrophin-releasing hormone (GnRH)-induced human extravillous trophoblast (EVT) cell invasion. studyfinding: Our finding indicates that GnRH-induced RUNX2 expression enhances the invasive capacity of EVT cells by modulating the expression of MMP2 and MMP9. what is known already: GnRH is expressed in first-trimester placenta and exerts pro-invasive effects on EVT cells in vitro. RUNX2 regulates MMP2 and MMP9 expression and is often associated with invasive phenotypes. study design, samples/materials, methods: First-trimesterhuman placenta (n 1/4 9) was obtained fromwomenundergoing elective termination of pregnancy. The localization of RUNX2, MMP2 and MMP9 in first-trimester human placenta was examined by immunohistochemistry. Primary or immortalized (HTR-8/SVneo) EVT cells were treated alone or in combination with GnRH, GnRH antagonist Antide, MAPK kinase inhibitor PD98095, phosphatidylinositol 3-kinase inhibitor LY294002, MMP2/9 inhibitor or small interfering RNAs (siRNAs) targeting RUNX2, MMP2 and/or MMP9. Protein and mRNA levels were measured by western blot and RT-PCR, respectively. Cell invasiveness was evaluated by transwell Matrigel or collagen I invasion assays. main results and the role of chance: RUNX2, MMP2 and MMP9 were detected in the cell column regions of human firsttrimester placental villi. GnRH treatment increased RUNX2 mRNA and protein levels in HTR-8/SVneo cells and primary EVTs, and these effects were attenuated by co-treatment with Antide, PD98095 or LY294002. Down-regulation of RUNX2 by siRNA reduced basal and GnRH-induced MMP2/9 expression and cell invasion. Moreover, pharmacological inhibition or siRNA-mediated knockdown of MMP2/9 reduced basal and GnRH-induced cell invasion. limitations, reasons for caution: The lack of an in vivo model is the major limitation of our in vitro study. wider implications of the findings: Our findings provide important insight into the functions of the GnRH - GnRH receptor system in early implantation and placentation. large scale data: Not applicable. study funding and competing interest(s): This research was supported by Canadian Institutes of Health Research (Grant #143317) to P.C.K.L. The authors have nothing to disclose.