Statins Inhibit NADPH Oxidase Activity by Interference with Membrane Raft Clustering Independent of Rac1 Inactivation in Endothelial Cells

Abstract

Statins, the inhibitor of 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) reductase, inhibit mevalonate pathway to block cholesterol biosynthesis, thereby lowering plasma cholesterol and being used for prevention and treatment of atherosclerotic diseases. Our previous studies have shown that statins including pravastatin and simvastatin interfere with the formation of OxLDL‐induced membrane raft (MR)‐redox signaling platforms in human coronary arterial endothelial cells (HCAECs) and in mouse coronary arterial endothelium. However, it remains unknown how statins produce this action that is independent of plasma cholesterol lowering. Here, we tested a hypothesis that inhibition of enhanced MR clustering is essential for the inhibitory effect of statins on Rac1 and NADPH oxidase (NOX) activity. We first demonstrated that statins had no effect on protein expression of NOX subunits, gp91phox or p47phox and Rac1 in HCAECs and mRNA levels of gp91phox or p47phox in coronary arteries of mice, confirming that statins primarily inhibit NADPH oxidase activation rather than alter their gene expression. Overexpression of oncogenic Rac1 in HCAECs was unable to reverse the inhibitory effects of statins on OxLDL‐induced MR clustering suggesting that the effect of statins on MR clustering does not require Rac1 activation. Electron spin resonance spectrometry demonstrated that incubation of HCAECs with sphingomyelinase, a strong stimulator of MR clustering significantly increased O2•− production, which was abolished by pravastatin and simvastatin or by a MR disruptor methyl‐β‐cyclodextrin. Taken together, these results indicate that statins inhibit NOX activity by interference with MR clustering independent of Rac1 activation. This action of statins on MR clustering may be associated with reduction of cholesterol in cell membrane of ECs (supported by NIH grants HL057244, HL091464 and HL075316).