title: Androgen receptor-dependent activation of endothelial nitric oxide synthase in vascular endothelial cells: role of phosphatidylinositol 3-kinase/akt pathway.
abstract: The mechanisms of testosterone-induced vasodilatation are not fully understood. This study investigated the effect of testosterone on nitric oxide (NO) synthesis and its molecular mechanism using human aortic endothelial cells (HAEC). Testosterone at physiological concentrations (1-100 nm) induced a rapid (15-30 min) increase in NO production, which was associated with phosphorylation and activation of endothelial NO synthase (eNOS). Then, the involvement of the androgen receptor (AR), which is abundantly expressed in HAEC, was examined. The effect of testosterone on eNOS activation and NO production were abolished by pretreatment with an AR antagonist nilutamide and by transfection with AR small interference RNA. In contrast, testosterone-induced eNOS phosphorylation was unchanged by pretreatment with an aromatase inhibitor or by transfection with ERalpha small interference RNA. 5alpha-Dihydrotestosterone, a nonaromatizable androgen, also stimulated eNOS phosphorylation. Next, the signaling cascade that leads to eNOS phosphorylation was explored. Testosterone stimulated rapid phosphorylation of Akt in a time- and dose-dependent manner, with maximal response at 15-60 min. The rapid phosphorylation of eNOS or NO production induced by testosterone was inhibited by Akt inhibitor SH-5 or by phosphatidylinositol (PI) 3-kinase inhibitor wortmannin. Co-immunoprecipitation assays revealed a testosterone-dependent interaction between AR and the p85alpha subunit of PI3-kinase. In conclusion, testosterone rapidly induces NO production via AR-dependent activation of eNOS in HAEC. Activation of PI3-kinase/Akt signaling and the direct interaction of AR with p85alpha are involved, at least in part, in eNOS phosphorylation.
authors.affiliation: Department of Geriatric Medicine, Graduate School of Medicine, The University of Tokyo; 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan.
authors.forename: Jing
authors.initials: J
authors.lastname: Yu
authors.forename: Masahiro
authors.initials: M
authors.lastname: Akishita
authors.forename: Masato
authors.initials: M
authors.lastname: Eto
authors.forename: Sumito
authors.initials: S
authors.lastname: Ogawa
authors.forename: Bo-Kyung
authors.initials: BK
authors.lastname: Son
authors.forename: Shigeaki
authors.initials: S
authors.lastname: Kato
authors.forename: Yasuyoshi
authors.initials: Y
authors.lastname: Ouchi
authors.forename: Tetsuro
authors.initials: T
authors.lastname: Okabe
mesh_terms: D000704:Analysis of Variance; D000726:Androgen Antagonists / Q000494:pharmacology; D000728:Androgens / Q000494:pharmacology; D015153:Blotting, Western; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000378:metabolism*; D005456:Fluorescent Dyes; D006801:Humans; D048289:Imidazolidines / Q000494:pharmacology; D047468:Immunoprecipitation; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D034741:RNA, Small Interfering; D011944:Receptors, Androgen / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013739:Testosterone / Q000378:metabolism / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection
chemical_list: D000726:Androgen Antagonists; D000728:Androgens; D005456:Fluorescent Dyes; D048289:Imidazolidines; D034741:RNA, Small Interfering; D011944:Receptors, Androgen; D013739:Testosterone; C021277:nilutamide; D052250:Nitric Oxide Synthase Type III; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt