title: Cyclosporin A inhibits flow-mediated activation of endothelial nitric-oxide synthase by altering cholesterol content in caveolae.
abstract: Fluid shear stress generated by blood flowing over the endothelium is a major determinant of arterial tone, vascular remodeling, and atherogenesis. Nitric oxide (NO) produced by endothelial NO synthase (eNOS) plays an essential role in regulation of vascular function and structure by blood flow. Although cyclosporin A (CsA), an inhibitory ligand of cyclophilin A, is a widely used immunosuppressive drug, it causes arterial hypertension in part by impairing eNOS-dependent vasodilation. Here we show that CsA inhibits fluid shear stress-mediated eNOS activation in endothelial cells via decreasing cholesterol content in caveolae. Exposure of cultured bovine aortic endothelial cells to 1 mum CsA for 1 h significantly inhibited NO production and eNOS phosphorylation at Ser-1179 induced by flow (shear stress=dynes/cm2). The effect of CsA was not related to inhibition of two known eNOS kinases, protein kinase B (Akt) and protein kinase A, because CsA did not affect Akt or protein kinase A activation. In rabbit aorta perfused ex vivo, CsA also significantly inhibited flow-induced eNOS phosphorylation at Ser-1179 but had no effect on Akt measured by phosphorylation at Ser-473. However, CsA treatment decreased cholesterol content in caveolae and displaced eNOS from caveolae, which may be caused by CsA disrupting the association of caveolin-1 and cyclophilin A. The magnitude of the cholesterol depleting effect was similar to that of beta-cyclodextrin, a cholesterol-binding molecule, and beta-cyclodextrin had a similar inhibitory effect on flow-mediated eNOS activation. Treating bovine aortic endothelial cells for 24 h with 30 mug/ml cholesterol blocked the CsA effect and restored eNOS phosphorylation in response to flow. These data suggest that decreasing cholesterol content in caveolae by CsA is a potentially important pathogenic mechanism for CsA-induced endothelial dysfunction and hypertension.
authors.affiliation: Department of Medicine and Center for Cardiovascular Research, University of Rochester School of Medicine and Dentistry, Aab Institute of Biomedical Sciences, Rochester, New York 14642, USA.
authors.forename: Andreea O
authors.initials: AO
authors.lastname: Lungu
authors.forename: Zheng-Gen
authors.initials: ZG
authors.lastname: Jin
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Yamawaki
authors.forename: Tatsuo
authors.initials: T
authors.lastname: Tanimoto
authors.forename: Chelsea
authors.initials: C
authors.lastname: Wong
authors.forename: Bradford C
authors.initials: BC
authors.lastname: Berk
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001011:Aorta / Q000378:metabolism; D015153:Blotting, Western; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002784:Cholesterol / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry; D016572:Cyclosporine / Q000494:pharmacology*; D003902:Detergents / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000201:enzymology / Q000378:metabolism; D004789:Enzyme Activation; D015151:Immunoblotting; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D007700:Kinetics; D009569:Nitric Oxide / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry; D011518:Proto-Oncogene Proteins / Q000737:chemistry; D051057:Proto-Oncogene Proteins c-akt; D011817:Rabbits; D012694:Serine / Q000737:chemistry; D013314:Stress, Mechanical; D013997:Time Factors; D047392:beta-Cyclodextrins / Q000494:pharmacology
chemical_list: D003902:Detergents; D007166:Immunosuppressive Agents; D011518:Proto-Oncogene Proteins; D047392:beta-Cyclodextrins; D009569:Nitric Oxide; D012694:Serine; D016572:Cyclosporine; D002784:Cholesterol; D019001:Nitric Oxide Synthase; D052250:Nitric Oxide Synthase Type III; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D017868:Cyclic AMP-Dependent Protein Kinases; C031215:betadex