Abstract 13748: Caveolin-1/Adiponectin Receptor 1 Interaction Positively Regulates Adiponectin Initiated Intracellular Signaling in Endothelial Cells
The potent anti-inflammatory/vascular protective effects of adiponectin (APN) are now well recognized. Reduced APN levels have been shown to contribute to vascular injury in diabetic patients. However, molecular mechanisms responsible for APN transmembrane signaling remain undefined. Recent studies have demonstrated that caveolin (Cav) is a critical regulator of transmembrane signaling. However, the role of Cav in APN's vasculoprotective function has never been investigated. Human umbilical vascular endothelial cells (HUVEC) were treated with APN in the presence and absence of TNFα, and the effect of siRNA-mediated caveolin-1 knockdown (Cav1-KD) on APN's physiologic and pharmacologic effects were determined. In control HUVEC (Con), APN significantly increased eNOS phosphorylation, elevated NO concentration, and inhibited TNFα-initiated inflammatory response as determined by ICAM-1 expression and nitrative stress. However, in Cav1-KD HUVEC, basal NO level was elevated, but APN failed to induce significant eNOS phosphorylation (+1.2±0.19 fold vs. +1.92±0.24 fold in Con, p0.05 vs. vehicle). Moreover, Cav1-KD abrogated APN's anti-TNFα effects as evidenced by diminished ICAM-1 downregulation (−10.6±2.5% vs. −51±6.3% in Con, P<0.01) and blunted anti-nitrative effect (−17.3±3.1% in nitrotyrosine content vs. −68.4±4.3% in Con, P<0.01). The expression of key APN signaling molecules, including APN receptors 1 and 2 (AdipoR1 and AdipoR2), APPL1, and AMPK were all normal in Cav1-KD cells. Con-focal microscopic observation demonstrated that AdipoR1 co-localizes with Cav-1 in HUVEC. Co-immunoprecipitation demonstrated that AdipoR1 and Cav-1 directly interact and form a complex in the caveolea. Moreover, two Cav binding motifs in AdipoR1 were identified to be sufficient and necessary for AdipoR1/Cav-1 interaction. In conclusion, we demonstrate for the first time that, similar to insulin signaling but differing from the signaling of many other proteins binding Cav, Cav-1/APNR1 interaction positively regulates APN transmembrane signaling, and is required for APN's physiologic NO stimulatory effects and pharmacologic anti-inflammatory effects in HUVEC.
- © 2010 by American Heart Association, Inc.