Androgen Receptor-Dependent Activation of Endothelial Nitric Oxide Synthase in Vascular Endothelial Cells: Role of Phosphatidylinositol 3-Kinase/Akt Pathway

• Published in: Endocrinology (Philadelphia) 2010-04, Vol.151 (4), p.1822-1828
• Main Authors: Yu, Jing, Akishita, Masahiro, Eto, Masato, Ogawa, Sumito, Son, Bo-Kyung, Kato, Shigeaki, Ouchi, Yasuyoshi, Okabe, Tetsuro
• Format: Article
• Language: English
• Subjects: 1-Phosphatidylinositol 3-kinase; AKT protein; Analysis of Variance; Androgen Antagonists - pharmacology; Androgen receptors; Androgens; Androgens - pharmacology; Aorta; Aromatase; Biological and medical sciences; Blotting, Western; Cell activation; Cells, Cultured; Chemical synthesis; Dihydrotestosterone; Dose-Response Relationship, Drug; Endothelial cells; Endothelial Cells - metabolism; Fluorescent Dyes; Fundamental and applied biological sciences. Psychology; Humans; Imidazolidines - pharmacology; Immunoprecipitation; Kinases; Molecular modelling; Nitric oxide; Nitric Oxide Synthase Type III - metabolism; Nitric-oxide synthase; Phosphatidylinositol 3-Kinases - metabolism; Phosphorylation; Phosphorylation - drug effects; Physiological effects; Pretreatment; Proto-Oncogene Proteins c-akt - metabolism; Receptors; Receptors, Androgen - genetics; Receptors, Androgen - metabolism; RNA, Small Interfering; RNA-mediated interference; Signal Transduction - drug effects; Signal Transduction - physiology; siRNA; Testosterone; Testosterone - metabolism; Testosterone - pharmacology; Time Factors; Transfection; Vasodilation; Vertebrates: endocrinology; Wortmannin
• ISSN: 0013-7227; 1945-7170
• DOI: 10.1210/en.2009-1048

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 ERα small interference RNA. 5α-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 p85α 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 p85α are involved, at least in part, in eNOS phosphorylation. Testosterone rapidly induces nitrogen oxide (NO) production via androgen receptor (AR)-dependent activation of endothelial NO synthase in human aortic endothelial cells, partly through direct interaction of AR with p85α.