Investigating the Mechanisms Underlying the Heterogeneous VSMC Contribution to Vascular Disease
In healthy blood vessels, vascular smooth muscle cells (VSMCs) exist in a contractile, quiescent state but upon vascular insult can switch phenotype to activate proliferation, migration and remodelling of the extracellular matrix. Phenotypically switched VSMCs contribute most cells within neointimal lesions, characteristic of atherosclerosis and in-stent restenosis, diseases that underlie heart attack and stroke. Using multicolour “Confetti” VSMC-specific lineage tracing in animal models of vascular disease, our lab and others have shown that the extensive VSMC contribution to these lesions results from the clonal expansion of few cells. To understand how oligoclonal VSMC lesion contribution arises and to identify the signals activating VSMC proliferation in vivo, I used quantified VSMC clonal development over time in two models of vascular disease. Following acute vascular injury, the number and sizes of patches of clonally expanded VSMCs steadily increased before reaching a plateau, suggesting rare activation of VSMC proliferation in few cells, rather than clonal competition following widespread VSMC activation. Interestingly, only a subset of medial patches gave rise to neointimal patches, suggesting that VSMC lesion invasion represents a second selective event underlying mature lesion oligoclonality. Infrequent activation of VSMC proliferation in atherosclerosis was evidenced by the absence of plaques with high numbers of colours at any stage of plaque development. Tamoxifen-inducible, VSMC-specific deletion of contractile master regulator Myocd in adult mice had modest phenotypic effects on baseline VSMC contractile marker expression, and on injury-induced VSMC transcriptional response and clonality. In both vascular disease models, VSMC activation was greatly enriched in vascular regions displaying elastic lamina alterations, medial acellularity and immune cell recruitment, implicating these as potential proliferation-inducing cues. However, not all VSMCs in these regions formed patches, suggesting that VSMCs must be primed to respond proliferatively. Consistent with the hypothesis that VSMCs marked by stem cell antigen-1 (SCA1) may represent such a primed population, profiling of chromatin accessibility in SCA1+ VSMCs revealed substantial opening of chromatin at genes showing increased expression in injury-activated compared to healthy VSMCs. Manipulation of RUNX1 and CEBP, transcription factors whose motifs were enriched at activation-specific open chromatin regions, could allow for control of VSMC priming and proliferation.
Cardiovascular Disease, Atherosclerosis, Vascular Smooth Muscle Cell, Clonal Expansion, Myocardin, Lineage-Tracing, Confocal Microscopy, ATAC-seq
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