Autophagy mediates tumor necrosis factor-α-induced phenotype switching in vascular smooth muscle A7r5 cell line

PloS One
Marina García-MiguelMario Chiong

Abstract

Vascular smooth muscle cells (VSMC) dedifferentiation from a contractile to a synthetic phenotype contributes to atherosclerosis. Atherosclerotic tissue has a chronic inflammatory component with high levels of tumor necrosis factor-α (TNF-α). VSMC of atheromatous plaques have increased autophagy, a mechanism responsible for protein and intracellular organelle degradation. The aim of this study was to evaluate whether TNF-α induces phenotype switching of VSMCs and whether this effect depends on autophagy. Rat aortic Vascular smooth A7r5 cell line was used as a model to examine the phenotype switching and autophagy. These cells were stimulated with TNF-α 100 ng/mL. Autophagy was determined by measuring LC3-II and p62 protein levels. Autophagy was inhibited using chloroquine and siRNA Beclin1. Cell dedifferentiation was evaluated by measuring the expression of contractile proteins α-SMA and SM22, extracellular matrix protein osteopontin and type I collagen levels. Cell proliferation was measured by [3H]-thymidine incorporation and MTT assay, and migration was evaluated by wound healing and transwell assays. Expression of IL-1β, IL-6 and IL-10 was assessed by ELISA. TNF-α induced autophagy as determined by increased LC3-II (1.91±0....Continue Reading

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Citations

Jan 28, 2020·Expert Opinion on Therapeutic Targets·Dorien G De MunckWim Martinet
Dec 15, 2019·Journal of Drug Targeting·Yin-Yu ZhangLi Qin
Jun 5, 2020·American Journal of Hypertension·Fred S LambRyan J Stark
Jan 7, 2021·Toxins·Nikolaos C KyriakidisPeter Stenvinkel

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Methods Mentioned

BETA
protein assay
ELISA
PCR
confocal microscopy

Software Mentioned

UN
IT gel
SCAN

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