Redistribution of the chromatin remodeler Brg1 directs smooth muscle–derived adventitial progenitor–to–myofibroblast differentiation and vascular fibrosis
Austin J. Jolly, Sizhao Lu, Allison M. Dubner, Keith A. Strand, Marie F. Mutryn, Aaron Pilotti-Riley, Etienne P. Danis, Raphael A. Nemenoff, Karen S. Moulton, Mark W. Majesky, Mary C.M. Weiser-Evans
Austin J. Jolly, Sizhao Lu, Allison M. Dubner, Keith A. Strand, Marie F. Mutryn, Aaron Pilotti-Riley, Etienne P. Danis, Raphael A. Nemenoff, Karen S. Moulton, Mark W. Majesky, Mary C.M. Weiser-Evans
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Research Article Vascular biology

Redistribution of the chromatin remodeler Brg1 directs smooth muscle–derived adventitial progenitor–to–myofibroblast differentiation and vascular fibrosis

Abstract

Vascular smooth muscle–derived Sca1+ adventitial progenitor (AdvSca1-SM) cells are tissue-resident, multipotent stem cells that contribute to progression of vascular remodeling and fibrosis. Upon acute vascular injury, AdvSca1-SM cells differentiate into myofibroblasts and are embedded in perivascular collagen and the extracellular matrix. While the phenotypic properties of AdvSca1-SM–derived myofibroblasts have been defined, the underlying epigenetic regulators driving the AdvSca1-SM–to–myofibroblast transition are unclear. We show that the chromatin remodeler Smarca4/Brg1 facilitates AdvSca1-SM myofibroblast differentiation. Brg1 mRNA and protein were upregulated in AdvSca1-SM cells after acute vascular injury, and pharmacological inhibition of Brg1 by the small molecule PFI-3 attenuated perivascular fibrosis and adventitial expansion. TGF-β1 stimulation of AdvSca1-SM cells in vitro reduced expression of stemness genes while inducing expression of myofibroblast genes that was associated with enhanced contractility; PFI blocked TGF-β1–induced phenotypic transition. Similarly, genetic knockdown of Brg1 in vivo reduced adventitial remodeling and fibrosis and reversed AdvSca1-SM–to–myofibroblast transition in vitro. Mechanistically, TGF-β1 promoted redistribution of Brg1 from distal intergenic sites of stemness genes and recruitment to promoter regions of myofibroblast-related genes, which was blocked by PFI-3. These data provide insight into epigenetic regulation of resident vascular progenitor cell differentiation and support that manipulating the AdvSca1-SM phenotype will provide antifibrotic clinical benefits.

Authors

Austin J. Jolly, Sizhao Lu, Allison M. Dubner, Keith A. Strand, Marie F. Mutryn, Aaron Pilotti-Riley, Etienne P. Danis, Raphael A. Nemenoff, Karen S. Moulton, Mark W. Majesky, Mary C.M. Weiser-Evans

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Figure 1

Carotid artery ligation increases Brg1 expression in AdvSca1-SM cells.

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Carotid artery ligation increases Brg1 expression in AdvSca1-SM cells. (...
(A) AdvSca1-SM cells were isolated from injured and uninjured carotid artery tissues from SM22α-Cre-YFP mice by FACS 3 days after ligation, and RNA extraction was performed. (B) Smarca4 (Brg1) mRNA expression was assessed by qPCR. Data represent 3 independent biological replicates. A 2-tailed unpaired t test was performed to compare the means of the 2 experimental groups; ***P < 0.001. (C) Uninjured and injured carotid arteries were harvested from tamoxifen-treated Gli1-CreERT-YFP AdvSca1-SM reporter mice 3 days after ligation, embedded in OCT, and immunofluorescently stained for YFP (green) to identify AdvSca1-SM cells and for Brg1 (red). A, Adventitia; M, Media. Elastin autofluorescence is observed on the green and red channel. Scale bar: 50 μm. (D) Total YFP+ and YFP+/Brg1+ cells were counted from stained slides from each group. Data are shown as total number per high-magnification field. n = 6 (uninjured), n = 6 (injured). Blue data points represent male samples, and red data points represent female samples. A 2-tailed unpaired t test was performed to compare the means of the 2 experimental groups; ***P < 0.001, ****P < 0.0001. (E) Smarca4 (Brg1) mRNA expression from human left ventricular tissue was obtained from the publicly available Genome Tissue Expression Database. The data were stratified by NPPB expression, nonhypertrophic tissue was defined as the lowest 30% of NPPB expression, and hypertrophic tissue was defined as the highest 30% of NPPB expression.