Smooth muscle–derived progenitor cell myofibroblast differentiation through KLF4 downregulation promotes arterial remodeling and fibrosis
Sizhao Lu, Austin J. Jolly, Keith A. Strand, Allison M. Dubner, Marie F. Mutryn, Karen S. Moulton, Raphael A. Nemenoff, Mark W. Majesky, Mary C.M. Weiser-Evans
Sizhao Lu, Austin J. Jolly, Keith A. Strand, Allison M. Dubner, Marie F. Mutryn, Karen S. Moulton, Raphael A. Nemenoff, Mark W. Majesky, Mary C.M. Weiser-Evans
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Research Article Stem cells Vascular biology

Smooth muscle–derived progenitor cell myofibroblast differentiation through KLF4 downregulation promotes arterial remodeling and fibrosis

Abstract

Resident vascular adventitial SCA1+ progenitor (AdvSca1) cells are essential in vascular development and injury. However, the heterogeneity of AdvSca1 cells presents a unique challenge in understanding signaling pathways orchestrating their behavior in homeostasis and injury responses. Using smooth muscle cell (SMC) lineage-tracing models, we identified a subpopulation of AdvSca1 cells (AdvSca1-SM) originating from mature SMCs that undergo reprogramming in situ and exhibit a multipotent phenotype. Here we employed lineage tracing and RNA-sequencing to define the signaling pathways regulating SMC-to-AdvSca1-SM cell reprogramming and AdvSca1-SM progenitor cell phenotype. Unbiased hierarchical clustering revealed that genes related to hedgehog/WNT/beta-catenin signaling were significantly enriched in AdvSca1-SM cells, emphasizing the importance of this signaling axis in the reprogramming event. Leveraging AdvSca1-SM–specific expression of GLI-Kruppel family member GLI1 (Gli1), we generated Gli1-CreERT2-ROSA26-YFP reporter mice to selectively track AdvSca1-SM cells. We demonstrated that physiologically relevant vascular injury or AdvSca1-SM cell–specific Kruppel-like factor 4 (Klf4) depletion facilitated the proliferation and differentiation of AdvSca1-SM cells to a profibrotic myofibroblast phenotype rather than macrophages. Surprisingly, AdvSca1-SM cells selectively contributed to adventitial remodeling and fibrosis but little to neointima formation. Together, these findings strongly support therapeutics aimed at preserving the AdvSca1-SM cell phenotype as a viable antifibrotic approach.

Authors

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

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Figure 1

Global analysis of genes differentially expressed between mature SMCs, SMC-derived AdvSca1-SM cells, and non–SMC-derived AdvSca1-MA cells.

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Global analysis of genes differentially expressed between mature SMCs, S...
Mature SMCs, AdvSca1-SM cells, and AdvSca1-MA cells were recovered from the carotid artery + aortic arch (CA+arch) and descending aortae (dAo) of SMC reporter mice as described in Methods. Total RNA was isolated from cell populations from pooled, digested arteries and analyzed by RNA-Seq. N = 3 independent experiments using arteries from 10–12 pooled mice per experiment were used for analysis. Differentially expressed genes were identified in all pairwise comparisons between the recovered populations (5265 genes). Hierarchical clustering was performed on the set of 5265 genes and clusters with high expression in specific populations were identified: cluster 1 — genes highly and selectively expressed in mature SMCs; cluster 2 — genes highly and selectively expressed in SMC-derived AdvSca1-SM cells; cluster 3 — genes highly expressed in non–SMC-derived AdvSca1-MA cells. Red, upregulated genes; green, downregulated genes.