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Author Notes:

E-mail: mchiong@uchile.cl

The authors finally thanks Fidel Albornoz and Gindra Latorre for their excellent technical assistance

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Research Funding:

This research was funded in part by Comision Nacional de Ciencia y Tecnologia (CONICYT), Chile (FONDECYT 1140329 and 1180157 to M.C.; FONDECYT 3160298 to J.A.R.; FONDAP 15130011 to M.C.). HL113167 and HL095070 from the National Institute of Health (to ASM). P.M. and DM-R hold CONICYT PhD fellowships.

Keywords:

  • Science & Technology
  • Multidisciplinary Sciences
  • Science & Technology - Other Topics
  • I-KAPPA-B
  • PROLIFERATION
  • ATHEROSCLEROSIS
  • METABOLISM
  • MECHANISMS
  • INHIBITORS
  • SECRETION
  • PATHWAY
  • GROWTH

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

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Journal Title:

PLoS ONE

Volume:

Volume 13, Number 5

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Type of Work:

Article | Final Publisher PDF

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±0.21, p<0.001) and decreased p62 (0.86±0.02, p<0.05) when compared to control. Additionally, TNF-α decreased α-SMA (0.74±0.12, p<0.05) and SM22 (0.54±0.01, p<0.01) protein levels. Consequently, TNF-α induced migration (1.25±0.05, p<0.05), proliferation (2.33±0.24, p<0.05), and the secretion of IL-6 (258±53, p<0.01), type I collagen (3.09±0.85, p<0.01) and osteopontin (2.32±0.46, p<0.01). Inhibition of autophagy prevented all the TNF-α-induced phenotypic changes. TNF-α induces phenotype switching in A7r5 cell line by a mechanism that required autophagy. Therefore, autophagy may be a potential therapeutic target for the treatment of atherosclerosis.

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This is an Open Access work distributed under the terms of the Creative Commons Universal : Public Domain Dedication License (http://creativecommons.org/publicdomain/zero/1.0/).

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