Regulation of autophagy and apoptosis in response to ox-LDL in vascular smooth muscle cells, and the modulatory effects of the microRNA hsa-let-7g

Abstract

Regulation of autophagy and apoptosis during treatment of vascular smooth muscle cells (VSMCs) with pro-atherogenic stimuli, such as oxidized low density lipoprotein (ox-LDL), remains unclear. We examined the expression of autophagy and apoptosis upon treatment of VSMCs with ox-LDL. Exposure to ox-LDL in modest amounts (10-40 μg/ml) enhanced autophagy (expression of beclin-1, LC3-II/LC3-1 ratio and Atg5) and apoptosis (expression of caspase-3, Bax, Bcl-2 and Bcl-xL); however, exposure to higher concentrations (≥ 60 μg/ml) induced high levels of apoptosis but autophagy declined. Pretreatment of VSMCs with the miRNA hsa-let-7 g inhibited autophagy, as LOX-1 expression and apoptosis declined. Hsa-let-7 g treatment also resulted in a decrease in intracellular ROS generation. Treatment with LOX-1 antibody had similar effects as hsa-let-7 g. Next, we studied autophagy and apoptosis in aortic segments from wild-type and LOX-1 knockout mice fed a high cholesterol diet, and observed increased autophagy as well as apoptosis in lipid-rich sections of aortas from wild-type mice and LOX-1 knockout mice (vs. corresponding controls); however, both autophagy and apoptosis in lipid-rich areas in aortic sections of LOX-1 knockout mice were less than in WT mice. These in vivo data are in keeping with in vitro data showing enhanced autophagy and apoptosis of VSMCs exposed to modest amount of ox-LDL. This study provides first set of data on the regulation of autophagy and apoptosis in ox-LDL-treated VSMCs. Our observations also suggest that hsa-let-7 g acts as a critical regulator of autophagy and apoptosis by modulating LOX-1.