Interleukin-6 inhibits endothelial nitric oxide synthase activation and increases endothelial nitric oxide synthase binding to stabilized caveolin-1 in human vascular endothelial cells

Abstract

We hypothesized a possible mechanism for atherosclerosis in which interleukin-6 (IL-6) might affect the endothelial nitric oxide synthase (eNOS)-caveolin-1 interaction and result in decreased nitric oxide bioavailability in the setting of low-grade inflammation. Because eNOS and caveolin-1 are crucial for vascular tone control, we studied the effects of IL-6 on the expression and activation of eNOS and caveolin-1 in human vascular endothelial cells. IL-6 inhibited the phosphorylation of eNOS at Ser1177 and the bradykinin-stimulated nitric oxide production; however, eNOS protein expression was not changed. In addition, IL-6 inhibited bradykinin-stimulated Akt phosphorylation at Ser473 and Thr 308 without affecting the Akt protein expression. IL-6 did not alter the mRNA level of caveolin-1; however, the caveolin-1 protein level was significantly increased dose-dependently. The binding of eNOS and caveolin-1 in endothelial cells, as demonstrated by coimmunoprecipitation assay, was increased by IL-6 treatment. IL-6 treatment was found to stabilize caveolin-1 protein and its half-life was estimated to prolong from 7.5 h to longer than 12 h. Furthermore, treatment with PD98059 and short interference RNA of extracellular signal-regulated kinase gene reversed the effects of IL-6 on eNOS and caveolin-1. In addition to decreasing Akt phosphorylation, the results of this study demonstrate, for the first time, the molecular mechanism underlying the effect of IL-6 to decrease the nitric oxide bioavailability by increasing the half-life and, therefore, the protein levels of caveolin-1. The increased caveolin-1 proteins bind more eNOS and consequently decrease eNOS activation by reducing the Ser1177 phosphorylation.