Enhancement of Autophagy by Simvastatin through Inhibition of Rac1‐mTOR Signaling Pathway

Abstract

In addition to their action lowering lower blood cholesterol levels, the statins have been found to modulate biological characteristics and function nof vascular smooth muscle cells (VSMCs) such as viability, proliferation, apoptosis, survival and contraction. The present study was designed to test whether simvastatin as a prototype enhances autophagy in VSMCs and thereby exerts their beneficial effects in atherosclerosis. Using Laser flow cytometry with a Cyto‐ID Green dye that detects autophagosomes, we found that simvastatin significantly increased autophagosome formation in VSMCs isolated from mouse coronary arteries. Western blot analysis also demonstrated that simvastatin significantly increased the protein expression of typical autophagy markers including LC3B and beclin‐1 in these mouse VSMCs. By activated Rac1‐GTPase pull‐down assay, simvastatin was shown to decrease basal Rac1‐GTPase activity and blocked angiotensin II‐induced Rac1 activation in VSMCs. Analysis of mTOR activity by assay of phosphorlylation of its substrate S6K demonstrated that simvastatin inhibited both basal and angiotensin II‐induced mTOR activity in VSMCs. Stimulation of mTOR activity by phosphatidic acid, an mTOR agonist, inhibited simvastatin‐induced autophagy in VSMCs. Moreover, simvastatin increased protein expression of a contractile phenotype marker calponin in VSMCs, while such increase in calponin expression was blocked by the autophagy inhibitor 3‐MA. Collectively, our results indicate that simvastatin inhibits Rac1‐mTOR pathway and thereby increases the autophagy in coronary VSMCs that may stabilize VSMCs in the contractile phenotype and prevent these cells from proliferation and growth (Supported by NIH grants HL‐091464, HL57244 and HL‐75316).