The Impact of 3-MA on Autophagy and Atherosclerosis via Wnt/β-catenin and AMPK/mTOR Pathways

Abstract

Abstract Objectives To study the mechanism of 3-methyladenine (3-MA) regulating autophagy and atherosclerosis (AS).Methods Ox-LDL-treated vascular smooth muscle cells (VSMCs) were used to construct an in vitro model of AS. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium assay was selected to detect the absorbance (OD) value of VSMCs. WB(Western Blotting) was utilized to analyze the levels of Beclin-1, p62, ULK1, α-SMA, SM22-α, OPN, Wnt, β-catenin, AMPK and mTOR proteins. Real-time fluorescence quantitative PCR (RTqPCR) was used to detect the expression of α-SMA, SM22-α, OPN, Wnt, β-catenin, AMPK, p62 mTOR, Beclin-1 and ULK1. Transwell was used to detect the migration ability of VSMCs. Lipid droplets in VSMC were stained by oil red O staining method.Results The protein expression levels of p62 in 3-MA + ox-LDL group were higher than those in ox-LDL group, while the protein expression levels of Wnt, β-catenin, p-AMPK/AMPK, p-mTOR/mTOR, Beclin-1 and ULK1 were lower than those in ox-LDL group. The gene expressions of p62 in 3-MA + ox-LDL group were higher than those in ox-LDL group, while the gene expressions of Wnt, β-catenin, AMPK, mTOR, Beclin-1 and ULK1 were lower than those in ox-LDL group. Reversing the regulation of the corresponding genes was achieved by IWP-4 intervention.Conclusions This study demonstrated that 3-MA can promote autophagy inhibition of AS via the Wnt/β-catenin and AMPK/mTOR pathway. It provides theoretical basis for improving clinical diagnosis and treatment of AS.