Role of Cation Channel TRPV4 in Mechanosensing, Myofibroblast Differentiation, and Pulmonary Fibrosis

Myofibroblast differentiation is a key process in the pathogenesis of fibrotic diseases. Transforming growth factor-β1 (TGF-β1) is a powerful inducer of myofibroblast differentiation and is implicated in pathogenesis of tissue fibrosis. This study was undertaken to determine the role of mitochondrial deacetylase SIRT3 in TGF-β1-induced myofibroblast differentiation in vitro and lung fibrosis in vivo. Treatment of human lung fibroblasts with TGF-β1 resulted in increased expression of fibrosis markers, smooth muscle α-actin (α-SMA), collagen-1, and fibronectin. TGF-β1 treatment also caused depletion of endogenous SIRT3, which paralleled with increased production of reactive oxygen species (ROS), DNA damage, and subsequent reduction in levels of 8-oxoguanine DNA glycosylase (OGG1), an enzyme that hydrolyzes oxidized guanine (8-oxo-dG) and thus protects DNA from oxidative damage. Overexpression of SIRT3 by adenovirus-mediated transduction reversed the effects of TGF-β1 on ROS production and mitochondrial DNA damage and inhibited TGF-β1-induced myofibroblast differentiation. To determine the antifibrotic role of SIRT3 in vivo, we used the bleomycin-induced mouse model of pulmonary fibrosis. Compared with wild-type controls, Sirt3-knockout mice showed exacerbated fibrosis after intratracheal instillation of bleomycin. Increased lung fibrosis was associated with decreased levels of OGG1 and concomitant accumulation of 8-oxo-dG and increased mitochondrial DNA damage. In contrast, the transgenic mice with whole body Sirt3 overexpression were protected from bleomycin-induced mtDNA damage and development of lung fibrosis. These data demonstrate a critical role of SIRT3 in the control of myofibroblast differentiation and lung fibrosis.

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Bromodomain and Extraterminal (BET) Protein Inhibition Restores Redox Balance and Inhibits Myofibroblast Activation

BioMed Research International ◽

10.1155/2019/1484736 ◽

2019 ◽

Vol 2019 ◽

pp. 1-11 ◽

Cited By ~ 5

Author(s):

Carmel J. W. Stock ◽

Charalambos Michaeloudes ◽

Patricia Leoni ◽

Andrew L. Durham ◽

Sharon Mumby ◽

...

Keyword(s):

Pulmonary Fibrosis ◽

Lung Disease ◽

Critical Role ◽

Redox Balance ◽

Protein Isoforms ◽

Related Factor ◽

Myofibroblast Differentiation ◽

Ros Production ◽

Bet Protein

Background and Objective. Progressive pulmonary fibrosis is the main cause of death in patients with systemic sclerosis (SSc) with interstitial lung disease (ILD) and in those with idiopathic pulmonary fibrosis (IPF). Transforming growth factor-β (TGF-β) and NADPH oxidase- (NOX-) derived reactive oxygen species (ROS) are drivers of lung fibrosis. We aimed to determine the role of the epigenetic readers, bromodomain and extraterminal (BET) proteins in the regulation of redox balance in activated myofibroblasts. Methods. In TGF-β-stimulated fibroblasts, we investigated the effect of the BET inhibitor JQ1 on the mRNA expression of the prooxidant gene NOX4 and the antioxidant gene superoxide dismutase (SOD2) by quantitative RT-PCR, the antioxidant transcription factor NF-E2-related factor 2 (Nrf2) activity by a reporter assay, and intracellular ROS levels by dichlorofluorescein staining. Myofibroblast activation was determined by α-smooth muscle actin immunocytochemistry. The role of specific BET protein isoforms in NOX4 gene regulation was studied by siRNA silencing and chromatin-immunoprecipitation. Results and Conclusions. Affymetrix gene array analysis revealed increased NOX4 and reduced SOD2 expression in SSc and IPF fibroblasts. SOD2 silencing in non-ILD control fibroblasts induced a profibrotic phenotype. TGF-β increased NOX4 and inhibited SOD2 expression, while increasing ROS production and myofibroblast differentiation. JQ1 reversed the TGF-β-mediated NOX4/SOD2 imbalance and Nrf2 inactivation and attenuated ROS production and myofibroblast differentiation. The BET proteins Brd3 and Brd4 were shown to bind to the NOX4 promoter and drive TGF-β-induced NOX4 expression. Our data indicate a critical role of BET proteins in promoting redox imbalance and pulmonary myofibroblast activation and support BET bromodomain inhibitors as a potential therapy for fibrotic lung disease.

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