Activation of the p38 MAPK pathway by follicle-stimulating hormone regulates steroidogenesis in granulosa cells differentially.
In the present study, we started out to test whether the follicle-stimulating hormone (FSH)-activated p38 MAPK signaling cascade was involved in the regulation of steroidogenesis in granulosa cells (GCs). GCs were prepared from the ovaries of DES-treated immature rats and cultured in serum-free medium. Treatment of GCs with FSH (50 ng/ml) induced the phosphorylation of p38 MAPK rapidly with the phosphorylation being observed within 5 min and reaching the highest level at 30 min. Such activation was protein kinase A-dependent as indicated by the results using specific inhibitors. FSH stimulated the production of progesterone and estradiol as well as the expression of the steroidogenic acute regulatory protein (StAR) in a time-dependent manner, with a maximum level being observed in the production of progesterone and StAR at 48 h. Moreover, the potent p38 MAPK inhibitor SB203580 (20 microM) augmented FSH-induced progesterone and StAR production, while reduced FSH-induced estradiol production at the same time (P<0.01). RT-PCR data showed that inclusion of SB203580 in the media enhanced FSH-stimulated StAR mRNA production, while decreased the FSH-stimulated P450arom mRNA expression (P<0.05). Immunocytochemical studies showed that FSH treatment together with the inhibition of p38 MAPK activity resulted in a higher expression of StAR in mitochondria than FSH treatment alone. FSH also significantly up-regulated the protein level of LRH-1, a member of the orphan receptor family that activates the expression of P450arom in ovaries and testes. p38 MAPK inactivation down-regulated the basal and FSH-induced LRH-1 expression significantly. The intra-cellular level of DAX-1, another orphan receptor that inhibits StAR expression, also decreased upon p38 MAPK being inactivated. For the first time, the present study suggests that FSH-activated p38 MAPK signal pathway regulates progesterone and estrogen production in GCs differentially, and that the transcription factors LRH-1 and DAX-1 might play important roles in the process.
State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100080, China.
Fu-Qing
FQ
Yu
Chun-Sheng
CS
Han
Wei
W
Yang
Xuan
X
Jin
Zhao-Yuan
ZY
Hu
Yi-Xun
YX
Liu
D000818:Animals; D001141:Aromatase / Q000032:analysis / Q000235:genetics; D002478:Cells, Cultured; D004789:Enzyme Activation; D004958:Estradiol / Q000032:analysis / Q000096:biosynthesis; D004967:Estrogens / Q000096:biosynthesis* / Q000235:genetics; D005260:Female; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D006107:Granulosa Cells / Q000187:drug effects / Q000378:metabolism*; D007150:Immunohistochemistry / Q000379:methods; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D010750:Phosphoproteins / Q000032:analysis / Q000235:genetics; D011374:Progesterone / Q000032:analysis / Q000096:biosynthesis; D012313:RNA / Q000032:analysis; D011863:Radioimmunoassay / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*; D000098682:Steroidogenic Acute Regulatory Protein
D004967:Estrogens; D010750:Phosphoproteins; D000098682:Steroidogenic Acute Regulatory Protein; D011374:Progesterone; D004958:Estradiol; D012313:RNA; D005640:Follicle Stimulating Hormone; D001141:Aromatase; D048051:p38 Mitogen-Activated Protein Kinases