5-Aza-2′-deoxycytidine inhibited PDGF-induced rat airway smooth muscle cell phenotypic switching

Abstract

Airway smooth muscle (ASM) cell phenotypic switching played an important role in airway remodeling in asthma. In vitro platelet-derived growth factor (PDGF) induced ASM cell phenotypic switching from a mature to pro-remodeling phenotype, but the mechanism remained incompletely understood. This study was to explore the effect of DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (Aza-CdR) on PDGF-induced rat ASM cell phenotypic switching and biological behaviors. Rat airway smooth muscle (RASM) cells were obtained by primary explant techniques. Western blot, 3-dimensional gel contraction, transwell and wound healing assay, and MTT were applied to detect cell phenotypic switching, contractility, migration and proliferation, respectively. Cytoskeleton rearrangement was observed by immunofluorescence. Results showed Aza-CdR inhibited PDGF-induced down-regulation of contractile markers in RASM cells and increased cell contractility. Aza-CdR inhibited PDGF-induced RASM cell migration by abrogating cell morphology change and cytoskeletal reorganization and attenuated the effect of PDGF on proliferating cell nuclear antigen expression and cell cycle progression, ultimately cell proliferation. PDGF-induced DNA methyltransferase 1 (DNMT1) expression was mediated by activation of PI3K/Akt and ERK signaling in RASM cells. Selective depletion of DNMT1 protein by Aza-CdR inhibited PDGF-induced RASM cell phenotypic switching, revealing DNMT1-mediated DNA methylation was implicated in asthmatic ASM remodeling. We proposed for the first time that DNMT1 played a key role in PDGF-induced RASM cell phenotypic switching and Aza-CdR is promising in intervening ASM remodeling in asthma. Although study of abnormal DNA methylation in PDGF-stimulated ASM cells is in its infancy, this work contributes to providing new insights into the mechanism of ASM remodeling and may be helpful for developing effective treatments for airway remodeling in asthma.