Adiponectin stimulates production of nitric oxide in vascular endothelial cells.
Adiponectin is secreted by adipose cells and mimics many metabolic actions of insulin. However, mechanisms by which adiponectin acts are poorly understood. The vascular action of insulin to stimulate endothelial production of nitric oxide (NO), leading to vasodilation and increased blood flow is an important component of insulin-stimulated whole body glucose utilization. Therefore, we hypothesized that adiponectin may also stimulate production of NO in endothelium. Bovine aortic endothelial cells in primary culture loaded with the NO-specific fluorescent dye 4,5-diaminofluorescein diacetate (DAF-2 DA) were treated with lysophosphatidic acid (LPA) (a calcium-releasing agonist) or adiponectin (10 microg/ml bacterially produced full-length adiponectin). LPA treatment increased production of NO by approximately 4-fold. Interestingly, adiponectin treatment significantly increased production of NO by approximately 3-fold. Preincubation of cells with wortmannin (phosphatidylinositol 3-kinase inhibitor) blocked only adiponectin- but not LPA-mediated production of NO. Using phospho-specific antibodies, we observed that either adiponectin or insulin treatment (but not LPA treatment) caused phosphorylation of both Akt at Ser473 and endothelial nitric-oxide synthase (eNOS) at Ser1179 that was inhibitable by wortmannin. We next transfected bovine aortic endothelial cells with dominant-inhibitory mutants of Akt (Akt-AAA) or AMP-activated protein kinase (AMPK) (AMPKK45R). Neither mutant affected production of NO in response to LPA treatment. Importantly, only AMPKK45R, but not Akt-AAA, caused a significant partial inhibition of NO production in response to adiponectin. Moreover, AMPK-K45R inhibited phosphorylation of eNOS at Ser1179 in response to adiponectin but not in response to insulin. We conclude that adiponectin has novel vascular actions to directly stimulate production of NO in endothelial cells using phosphatidylinositol 3-kinase-dependent pathways involving phosphorylation of eNOS at Ser1179 by AMPK. Thus, the effects of adiponectin to augment metabolic actions of insulin in vivo may be due, in part, to vasodilator actions of adiponectin.
Chen
Hui
H
Diabetes Unit, Laboratory of Clinical Investigation, NCCAM, National Institutes of Health, Bethesda, Maryland 20892, USA.
Montagnani
Monica
M
Funahashi
Tohru
T
Shimomura
Iichiro
I
Quon
Michael J
MJ
D055372:AMP-Activated Protein Kinases; D052242:Adiponectin; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001011:Aorta; D002417:Cattle; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019793:Fluorescein; D005456:Fluorescent Dyes; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins; D008164:Luminescent Proteins / Q000235:genetics; D008246:Lysophospholipids / Q000494:pharmacology; D051379:Mice; D009097:Multienzyme Complexes / Q000235:genetics; D009154:Mutation; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics; D011506:Proteins / Q000494:pharmacology*; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D011994:Recombinant Proteins; D014162:Transfection; D014664:Vasodilation; D000077191:Wortmannin; D000097573:Red Fluorescent Protein
D055372:AMP-Activated Protein Kinases; D052242:Adiponectin; D000730:Androstadienes; D004791:Enzyme Inhibitors; D019793:Fluorescein; D005456:Fluorescent Dyes; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D008164:Luminescent Proteins; D008246:Lysophospholipids; D009097:Multienzyme Complexes; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017346:Protein Serine-Threonine Kinases; D011506:Proteins; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D011994:Recombinant Proteins; D000077191:Wortmannin; D000097573:Red Fluorescent Protein; C414689:4,5-diaminofluorescein diacetate; C496314:NOS3 protein, human; C496315:Nos3 protein, mouse; C496316:Nos3 protein, rat; C494918:AKT1 protein, human; C494920:Akt1 protein, rat