Inhibition of Myocardin-Related Transcription Factor/Serum Response Factor Signaling Decreases Lung Fibrosis and Promotes Mesenchymal Cell Apoptosis

Abstract

Myofibroblasts are crucial to the pathogenesis of tissue fibrosis. Their formation of stress fibers results in the release of myocardin-related transcription factor (MRTF), a transcriptional coactivator of serum response factor (SRF). MRTF-A (Mkl1)-deficient mice are protected from lung fibrosis. We hypothesized that the SRF/MRTF pathway inhibitor CCG-203971 would modulate myofibroblast function in vitro and limit lung fibrosis in vivo. Normal and idiopathic pulmonary fibrosis lung fibroblasts were treated with/without CCG-203971 (N-[4-chlorophenyl]-1-[3-(2-furanyl)benzoyl]-3-piperidine carboxamide) and/or Fas-activating antibody in the presence/absence of transforming growth factor (TGF)-β1, and apoptosis was assessed. In vivo studies examined the effect of therapeutically administered CCG-203971 on lung fibrosis in two distinct murine models of fibrosis induced by bleomycin or targeted type II alveolar epithelial injury. In vitro, CCG-203971 prevented nuclear localization of MRTF-A; increased the apoptotic susceptibility of normal and idiopathic pulmonary fibrosis fibroblasts; blocked TGF-β1-induced myofibroblast differentiation; and inhibited TGF-β1-induced expression of fibronectin, X-linked inhibitor of apoptosis, and plasminogen activator inhibitor-1. TGF-β1 did not protect fibroblasts or myofibroblasts from apoptosis in the presence of CCG-203971. In vivo, CCG-203971 significantly reduced lung collagen content in both murine models while decreasing alveolar plasminogen activator inhibitor-1 and promoting myofibroblast apoptosis. These data support a central role of the SRF/MRTF pathway in the pathobiology of lung fibrosis and suggest that its inhibition can help resolve lung fibrosis by promoting fibroblast apoptosis.

Mechanisms of Lung Fibrosis Resolution

“…69 Apoptosis has also been identified as a primary mechanism of myofibroblast removal in the bleomycin lung fibrosis model. 55,70,71 An accumulating body of literature demonstrates that lung fibroblasts and myofibroblasts in both mouse fibrosis models and IPF acquire an apoptosis-resistant phenotype. 41,48,55,59,70,71 Several mechanisms by which myofibroblasts evade apoptosis have been identified, 72 and several common mechanistic themes have emerged.…”

“…62,76 Several recent murine studies demonstrate that pharmacological strategies to induce myofibroblast apoptosis after fibrosis were already established and can accelerate resolution. 41,55,70,71 The histone deacetylase inhibitor…”

“…71 Targeting myofibroblast stress fiber formation through, either the Rho-associated kinase pathway or myocardin-related transcription factor/serum response factor signaling, induced preexisting lung myofibroblasts of IPF patients to undergo apoptosis. 41,55 Specific pharmacological agents targeting these pathways also reduced postinflammatory lung fibrosis in bleomycin-treated mice. 41,55 Pharmacological targeting inhibitor of apoptosis family proteins X-linked inhibitor of apoptosis and cellular IAP reduced postinflammatory lung fibrosis in bleomycin-treated mice while promoting fibroblast/ myofibroblast apoptosis.…”

“…41,55 Specific pharmacological agents targeting these pathways also reduced postinflammatory lung fibrosis in bleomycin-treated mice. 41,55 Pharmacological targeting inhibitor of apoptosis family proteins X-linked inhibitor of apoptosis and cellular IAP reduced postinflammatory lung fibrosis in bleomycin-treated mice while promoting fibroblast/ myofibroblast apoptosis. 70 In summary, experimental evidence demonstrates that strategies that augment apoptosis in myofibroblasts hasten lung fibrosis resolution.…”

“…Recent preclinical investigations support the concept that targeting intrinsic lung myofibroblast dedifferentiation, apoptosis, or senescence pathways effectively promotes resolution and may prove to be practical approaches for treatment of lung fibrosis. 41,50,55,70,71 There are clearly significant gaps in understanding lung fibrosis resolution likely reflecting the biological complexity and limitations in the animal models and basic scientific methods previously available to investigators. However, accelerated improvements in biotechnology and bioinformatics are revolutionizing biomedical research, and it is likely these new techniques and discoveries will continue to improve the understanding of the mechanisms of lung fibrosis resolution.…”