Regulation of progesterone production in granulosa cells is important for normal reproductive functions. Steroidogenic acute regulatory protein (StAR) is recognized as the key regulatory protein involved in the rate-limiting step of steroidogenesis. TGF-β1 protein is detected in human follicular fluid, and TGF-β1 and its receptors are expressed in human granulosa cells. However, the functional role of TGF-β1 in the regulation of StAR expression and progesterone production in human granulosa cells remains unknown. Our objective was to investigate the effects of TGF-β1 on StAR expression and progesterone production in human granulosa cells. SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization and immortalized with SV40 large T antigen. SVOG cells were used to investigate the effects of TGF-β1 on StAR expression and progesterone production at an academic research center. Levels of mRNA and protein were examined by RT-qPCR and western blotting, respectively. The accumulation levels of progesterone were measured by enzyme-linked immunosorbent assay (ELISA). TGF-β1 treatment downregulated StAR expression and decreased progesterone production. The suppressive effects of TGF-β1 on StAR expression and progesterone production were abolished by the inhibition of TGF-β type I receptor. In addition, treatment with TGF-β1 activated the Smad2/3 and ERK1/2 signaling pathways. The inhibition of the Smad3 and ERK1/2 signaling pathways attenuated the TGF-β1-induced downregulation of StAR expression and progesterone production. TGF-β1 downregulated StAR expression and decreased progesterone production by activating the Smad3 and ERK1/2 signaling pathways in human granulosa cells.

Steroidogenic acute regulatory protein (StAR) mediates the rate-limiting step in ovarian steroidogenesis and progesterone (P4) synthesis. Melatonin and its receptors are expressed in human granulosa cells, and have been shown to influence basal P4 production. However, previous studies addressing the regulation of StAR expression by melatonin and its impact on P4 secretion yielded contradictory results. Here, we demonstrate that melatonin upregulates StAR expression in primary cultures of human granulosa-lutein (hGL) cells obtained from women undergoing in vitro fertilization (IVF). Using pharmacological inhibitors, we show that the stimulatory effect of melatonin on StAR expression is mediated via both MT1 and MT2 melatonin receptors. Melatonin exposure activates the PI3K/AKT signaling pathway and its inhibition attenuates the stimulatory effect of melatonin on StAR expression. Moreover, siRNA-mediated knockdown of StAR abolishes melatonin-induced P4 production. Importantly, clinical analyses demonstrate that melatonin levels in human follicular fluid are positively correlated with P4 levels in serum. By illustrating the potential physiological role of melatonin in the regulation of StAR expression and P4 production in hGL cells, our results may serve to improve current strategies used to treat clinical infertility.

Aberrant regulation of ovulation is one of the major causes of infertility. In animal models, 3 epidermal growth factor (EGF)-like growth factors, amphiregulin (AREG), betacellulin (BTC), and epiregulin (EREG), have been shown to be involved in ovulation by regulating cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production. However, whether the same is true in humans remains largely unknown. Our objective was to investigate the effects of AREG, BTC, and EREG on COX-2 expression and PGE2 production in human granulosa cells. SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization and immortalized with SV40 large T antigen. SVOG cells were used to investigate the effect of AREG, BTC, and EREG on ovulation-related functions at an academic research center. Levels of mRNA and protein were examined by quantitative RT-PCR and Western blotting, respectively. The protein levels of PGE2 were measured by ELISA. LH treatment upregulated AREG, BTC, EREG, and COX-2. Knockdown of EGF receptor (EGFR) attenuated LH-induced COX-2 expression and PGE2 production. Treatment with AREG, BTC, and EREG upregulated COX-2 expression and PGE2 production. The stimulatory effects of AREG, BTC, and EREG on COX-2 expression and PGE2 production were blocked by inhibition of EGFR activity and expression. AREG-, BTC-, and EREG-activated ERK1/2 signaling, but not Akt signaling, was required for AREG-, BTC-, and EREG-induced COX-2 expression and PGE2 production. AREG, BTC, and EREG induced PGE2 production by upregulating COX-2 expression through ERK1/2 signaling in human granulosa cells.

Cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production have been shown to play key roles in the regulation of ovulation. The TGF-β superfamily members are important molecules that regulate many ovarian functions under normal physiological and pathological conditions. TGF-β1 and its receptors are expressed in human granulosa cells. However, to date, whether TGF-β1 can regulate COX-2 expression and PGE2 production, which in turn contribute to the process of ovulation, remains unknown. The objective of the study was to investigate the effects of TGF-β1 on COX-2 expression and PGE2 production in human granulosa cells. SVOG cells are human granulosa cells that were obtained from women undergoing in vitro fertilization and immortalized with Simian virus 40 large T antigen. SVOG cells were used to investigate the effect of TGF-β1 on COX-2 expression and PGE2 production. The study was conducted at an academic research center. mRNA and protein levels were examined by RT-quantitative real-time PCR and Western blotting, respectively. The concentrations of PGE2 in the culture medium were measured by an ELISA. TGF-β1 treatment induced COX-2 expression and PGE2 production. The inductive effects of TGF-β1 on COX-2 and PGE2 were abolished by the inhibition of TGF-β type I receptor (TβRI). In addition, treatment with TGF-β1 activated phosphorylated mothers against decapentaplegic (Smad)-2 and Smad3 signaling pathways. Inhibition of the Smad signaling pathways by small interfering RNA-mediated approaches attenuated the TGF-β1-induced COX-2 expression and PGE2 production. TGF-β1 induced PGE2 production by inducing the COX-2 expression through a Smad-dependent signaling pathway in human granulosa cells.