Differential Regulation and Sterol-Responsiveness of the HDL Receptor SR-BI and the LDL Receptor in Follicle-Stimulating Hormone (FSH) and Transforming Growth Factor Beta1 (TGFBeta1)-Promoted Steroidogenesis in Ovarian Granulosa Cells.

Ovarian steroidogenesis is fundamental to female reproductive competence and systemic health. Orchestration of pituitary gonadotropins and intra-ovarian factors including TGFbeta1 tightly regulate such ovarian function. We and others have demonstrated TGFbeta1 potently enhances FSH-promoted steroidogenesis in ovarian granulosa cells, and this is partly attributed to upregulation of cellular steroidogenic machinery through activation of cAMP/PKA and PI3K/Akt pathways. Steroidogenic cells, like other cells, could uptake cholesterol through low-density lipoprotein receptor (LDLR)-mediated endocytosis. Additionally, they selectively uptake cholesteryl ester through a high-density lipoprotein (HDL) receptor, the scavenger receptor class B member 1 (SR-BI/SCARB1). During rodent ovarian luteinization, SR-BI is upregulated to supply cholesterol for progesterone synthesis. Also, studies showed FSH stimulates SR-BI and LDLR expression in rat granulosa cells. This study was to investigate whether FSH and TGFbeta1 upregulation of steroidogenesis in rat granulosa cells involves differential regulation of SR-BI and LDLR. There were two specific aims. The first was to address whether and how FSH and TGFbeta1 regulate SR-BI and LDLR expression. The second aim was to ask whether there is a functional segregation between SR-BI and LDLR. Primary culture of granulosa cells isolated from eCG-primed immature rats was used. FSH treatment within 6 hours maximally increased SR-BI protein level that gradually decreased after 12 hours of treatment, while FSH alone did not significantly affect LDLR level. On the other hand, combined treatment with FSH and TGFbeta1 increased both proteins up to 48 hours-post treatment. Interestingly, FSH and TGFbeta1 induction of Scarb1 may involve PI3K and PKA signaling, while the induction of Ldlr involves PI3K but not PKA. Also, FSH and TGFbeta1 increased SR-BI localization at plasma membrane microvillar extensions. In the presence of HDL₃, SR-BI actively supplied lipoprotein-derived cholesterol for progesterone synthesis as suggested by SR-BI inhibitor BLT-1 blockade of steroidogenesis. In non-steroidogenic cells, uptake and synthesis of cholesterol are inversely regulated depending on intracellular cholesterol level. To understand whether this mode of regulation also applies to granulosa cells, we examined responses of Scarb1, Ldlr, and Hmgcr (HMG-CoA reductase, the rate-limiting enzyme in cholesterol de novo synthesis) to HDL₃ supplementation. HDL₃ dose-dependently reduced the FSH and TGFbeta1-upregulated Ldlr and Hmgcr, and interestingly, this had no obvious effect on Scarb1. This suggests that Scarb1 expression, unlike that of Ldlr and Hmgcr, is relatively insensitive to cholesterol feedback control. This study implicates that granulosa cells like non-steroidogenic cells possess the fundamental cholesterol feedback control system, and are additionally equipped with a feedback control-insensitive system (like SR-BI) to meet the demand of cholesterol for steroidogenesis under stimulation by FSH and TGFbeta1. This research was supported by a grant from Taiwan National Science Council to JJH (NSC96-2320-B-010-022-MY3). WAL was supported by an award (DD9801N) from National Health Research Institutes, Taiwan.

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