title: AMPK activation by dihydrotestosterone reduces FSH-stimulated cell proliferation in rat granulosa cells by inhibiting ERK signaling pathway.
abstract: We have previously reported that 5α-dihydrotestosterone (DHT) inhibits FSH-mediated granulosa cell proliferation by reducing cyclin D2 mRNA expression and blocking cell cycle progression at G1/S phase. The present study investigated the role of AMP activated protein kinase (AMPK) in DHT-mediated inhibition of granulosa cell proliferation. Granulosa cells harvested from 3-d estradiol primed immature rats were exposed to different concentrations of DHT (0, 45, and 90 ng/ml) for 24 h. Western blot analysis of immunoprecipitated AMPK showed a dose-dependent activation (P < 0.05) as evidenced by the increased phosphorylation at thr 172. In addition, time-courses studies (0, 6, 12, and 24 h) using DHT (90 ng/ml) showed a time-dependent increase in AMPK activation with maximum effect at 24 h. FSH inhibited AMPK phosphorylation and promoted granulosa cell proliferation, but pretreatment with DHT (90 ng/ml) for 24 h prior to FSH treatment reduced this effect. Pharmacological activation of AMPK with 5-aminoimidazole-4-carboxamide-1-β4-ribofuranoside abolished FSH-mediated ERK phosphorylation, indicating that AMPK is a negative upstream regulator of ERK. Furthermore, inhibition of AMPK activation by compound C reversed the DHT-mediated reduction in positive cell cycle regulator, cyclin D2, and 5-bromo-2'-deoxyuridine incorporation. These results suggest that elevated levels of DHT activate AMPK, which in turn inhibits ERK phosphorylation. Thus, inhibition of ERK phosphorylation by activated AMPK in response to DHT might contribute to decreased granulosa cell mitogenesis and ovulatory dysfunction seen in hyperandrogenic states.
authors.affiliation: Department of Obstetrics and Gynecology, University of Michigan Medical School, Ann Arbor, Michigan 48109, USA.
authors.forename: Pradeep P
authors.initials: PP
authors.lastname: Kayampilly
authors.forename: K M J
authors.initials: KM
authors.lastname: Menon
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D000728:Androgens / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D056742:Cyclin D2 / Q000235:genetics / Q000378:metabolism; D013196:Dihydrotestosterone / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005260:Female; D005640:Follicle Stimulating Hormone / Q000494:pharmacology*; D015870:Gene Expression / Q000187:drug effects; D006107:Granulosa Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D006728:Hormones / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
chemical_list: D000728:Androgens; D056742:Cyclin D2; D006728:Hormones; D013196:Dihydrotestosterone; D005640:Follicle Stimulating Hormone; D048049:Extracellular Signal-Regulated MAP Kinases; D055372:AMP-Activated Protein Kinases