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Caveolin-1 phosphorylation regulates vascular endothelial insulin uptake and is impaired by insulin resistance in rats.

ABSTRACT: As insulin entry into muscle interstitium is rate-limiting for its overall peripheral action, defining the route and regulation of its entry is critical. Caveolin-1 is required for caveola formation in vascular endothelial cells (ECs) and for EC insulin uptake. Whether this requirement reflects simply the need for caveola availability or involves a more active role for caveolae/caveolin-1 is not known. Here, we examined the role of insulin-stimulated tyrosine 14 (Tyr(14))-caveolin-1 phosphorylation in mediating EC insulin uptake and the role of cellular Src-kinase (cSrc), TNF-?/IL-6 and high fat diet (HFD) in regulating this process.Freshly isolated ECs from normal or HFD-fed rats and/or cultured ECs were treated with FITC-labelled or regular insulin with or without a Src or phosphotidylinositol-3-kinase inhibitor, TNF-? or IL-6, or transfecting FLAG-tagged wild-type (WT) or mutant (Y14F) caveolin-1. Tyr(14)-caveolin-1/Tyr(416) cSrc phosphorylation and FITC-insulin uptake were quantified by immunostaining and/or western blots.Insulin stimulated Tyr(14)-caveolin-1 phosphorylation during EC insulin uptake. Inhibiting cSrc, but not phosphotidylinositol-3-kinase, reduced insulin-stimulated caveolin-1 phosphorylation. Furthermore, inhibiting cSrc reduced FITC-insulin uptake by ?50%. Overexpression of caveolin-1Y14F inhibited, while overexpression of WT caveolin-1 increased, FITC-insulin uptake. Exposure of ECs to TNF-? or IL-6, or to 1-week HFD feeding eliminated insulin-stimulated caveolin-1 phosphorylation and inhibited FITC-insulin uptake to a similar extent.Insulin stimulation of its own uptake requires caveolin-1 phosphorylation and Src-kinase activity. HFD in vivo and proinflammatory cytokines in vitro both inhibit this process.

SUBMITTER: Wang H

PROVIDER: S-EPMC4417063 | biostudies-literature | 2015 Jun

REPOSITORIES: biostudies-literature

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Caveolin-1 phosphorylation regulates vascular endothelial insulin uptake and is impaired by insulin resistance in rats.

Wang Hong H Wang Aileen X AX Aylor Kevin K Barrett Eugene J EJ

Diabetologia 20150310 6

<h4>Aims/hypothesis</h4>As insulin entry into muscle interstitium is rate-limiting for its overall peripheral action, defining the route and regulation of its entry is critical. Caveolin-1 is required for caveola formation in vascular endothelial cells (ECs) and for EC insulin uptake. Whether this requirement reflects simply the need for caveola availability or involves a more active role for caveolae/caveolin-1 is not known. Here, we examined the role of insulin-stimulated tyrosine 14 (Tyr(14)) ...[more]

PMID: 25748795

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Project description:BackgroundAtherosclerosis is driven by synergistic interactions between pathological, biomechanical, inflammatory, and lipid metabolic factors. Our previous studies demonstrated that absence of caveolin-1 (Cav1)/caveolae in hyperlipidemic mice strongly inhibits atherosclerosis, which was attributed to activation of endothelial nitric oxide (NO) synthase (eNOS) and increased production of NO and reduced inflammation and low-density lipoprotein trafficking. However, the contribution of eNOS activation and NO production in the athero-protection of Cav1 and the exact mechanisms by which Cav1/caveolae control the pathogenesis of diet-induced atherosclerosis are still not clear.MethodsTriple-knockout mouse lacking expression of eNOS, Cav1, and Ldlr were generated to explore the role of NO production in Cav1-dependent athero-protective function. The effects of Cav1 on lipid trafficking, extracellular matrix remodeling, and vascular inflammation were studied both in vitro and in vivo with a mouse model of diet-induced atherosclerosis. The expression of Cav1 and distribution of caveolae regulated by flow were analyzed by immunofluorescence staining and transmission electron microscopy.ResultsWe found that absence of Cav1 significantly suppressed atherogenesis in Ldlr-/-eNOS-/- mice, demonstrating that athero-suppression is independent of increased NO production. Instead, we find that the absence of Cav1/caveolae inhibited low-density lipoprotein transport across the endothelium and proatherogenic fibronectin deposition and disturbed flow-mediated endothelial cell inflammation. Consistent with the idea that Cav1/caveolae may play a role in early flow-dependent inflammatory priming, distinct patterns of Cav1 expression and caveolae distribution were observed in athero-prone and athero-resistant areas of the aortic arch even in wild-type mice.ConclusionsThese findings support a role for Cav1/caveolae as a central regulator of atherosclerosis that links biomechanical, metabolic, and inflammatory pathways independently of endothelial eNOS activation and NO production.

Dataset Information

Caveolin-1 phosphorylation regulates vascular endothelial insulin uptake and is impaired by insulin resistance in rats.

Publications

Caveolin-1 phosphorylation regulates vascular endothelial insulin uptake and is impaired by insulin resistance in rats.

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