201.Regulation of mouse mural granulosa cell progesterone synthesis by oocyte paracrine factors

Abstract

Oocytes secrete soluble paracrine factors that play an important role in the growth and development of surrounding follicular cells. It is known that oocytes suppress FSH-induced progesterone production by mural granulosa cells (MGC), however it is unclear which growth factor(s) are involved. Some candidate molecules include growth differentiation factor 9 (GDF9), bone morphogenetic protein 15 (BMP15) and BMP6. The aim of this study was to examine the role of these factors in the regulation of FSH-induced MGC progesterone production. Prepubertal 129/SV mice were primed with eCG and ovaries were collected ~46�h later. MGC from large antral follicles were cultured with either denuded oocytes (DO; 0.25/μl), GDF9 or BMP15 (0.25-4% v/v) or BMP6 (10-200ng/ml), in the absence or presence of FSH. Cells were cultured for 18�h followed by a further 6�h pulse of 3H-thymidine. After 24�h cells and media were harvested for assessment of MGC DNA and progesterone synthesis, respectively. Treatment with FSH increased MGC progesterone production 9-fold, which, as expected, was antagonised by coculture with DO (by 73%). GDF9 and BMP15 both decreased FSH-induced MGC progesterone in a dose dependent manner, significantly reducing control levels (100%) to 17% and 30%, respectively, at doses of 2%v/v. All doses of BMP6 abolished FSH-stimulated progesterone. Even though all treatments inhibited progesterone production, only two of these, GDF9 (0.25%v/v) and DO (0.25DO/μl), stimulated MGC DNA synthesis (2.1 and 3.3 fold above controls, respectively). The BMP receptor type-II (BMPR-II) is a known receptor of several oocyte factors. Treatment with a BMPR-II ectodomain completely antagonised DO- and GDF9-stimualted MGC DNA synthesis, but progesterone levels were only partially restored (by 50%). These data indicate that BMP15 and BMP6 mimic the progesterone-regulating, but not the growth-promoting, activities of oocytes, whereas GDF9 does both. Although the BMPR-II ECD antagonises these oocyte factors, this receptor-signalling system may not necessarily be the means by which oocytes regulate MGC progesterone synthesis.