Primed smooth muscle cells acting as first responder cells in disease

Abstract

Rationale; Vascular smooth muscle cell (VSMC) dysregulation is a hallmark of vascular disease, including atherosclerosis. In particular, the majority of cells within atherosclerotic lesions are generated from pre-existing VSMCs and a clonal nature has been documented for VSMC-derived cells in multiple disease models. However, the mechanisms underlying the generation of oligoclonal lesions and the phenotype of proliferating VSMCs are unknown. Objective; To understand the cellular mechanisms underlying clonal VSMC expansion in disease. Methods and Results; Here we analyse clonal dynamics in multi-color lineage-traced animals over time after vessel injury. We demonstrate that VSMC proliferation is initiated in a small fraction of VSMCs that initially expand clonally in the medial layer and then migrate to form the oligoclonal neointima. Selective activation of VSMC proliferation also occurs in vitro, suggesting that this is a cell-autonomous feature. Mapping of VSMC trajectories using single-cell RNA-sequencing reveals a continuum of cellular states after injury and suggests that VSMC proliferation initiates in cells that have downregulated the contractile phenotype and show evidence of pronounced phenotypic switching. We show that proliferation is associated with induced expression of stem cell antigen 1 (SCA1) and the expression signature previously identified in SCA1+ VSMCs in healthy arteries. A remarkably increased proliferation of SCA1+ VSMCs, directly validated in functional assays, indicates that SCA1+ VSMCs act as "first responders" in vascular injury. Early atherosclerotic lesions also had clonal VSMC contribution and we show that the proliferation-associated injury response is conserved in plaque VSMCs, extending these findings to atherosclerosis. Finally, we identify VSMCs in healthy human arteries that correspond to the SCA1+ state in mouse VSMCs and show that genes identified as differentially expressed in this human VSMC subpopulation are enriched for genes showing genetic association with cardiovascular disease. Conclusions; We show that cell-intrinsic, selective VSMC activation drives clonal proliferation after injury and in atherosclerosis. Our study suggests that healthy mouse and human arteries contain VSMCs characterised by expression of disease-associated genes that are predisposed for proliferation. Targeting such "first responder" cells in patients undergoing vascular surgery could effectively prevent injury-associated VSMC activation and neoatherosclerosis.

RBPMS promotes contractile smooth muscle splicing and alters phenotypic behaviour of human embryonic stem cell derived vascular smooth muscle cells

“…This is because artefactual molecular differences arising from culture conditions, matrix stiffness and other non-physiological factors will obscure differences reflective of the true VSMC phenotypic state or of the effects of specific genetic perturbations. Second, to properly map the mature/differentiated adult SM-AS network, we would need to compare its splicing profile with a cognate phenotypically switched network, ideally in vivo during, for example, injury response [3, 59]. This is achievable in rodent systems but very challenging to model with human VSMCs.…”

“…This switch is accompanied by major changes in the cellular transcriptome including loss of expression of the contractile marker network. More recently, with the advent of single cell RNA sequencing, we have been able to dissect these molecular changes in far greater detail and better appreciate the phenotype switching process and the heterogeneity underlying contractile VSMCs of different embryonic lineages [1][2][3][4][5]. However, most of our knowledge of the molecular networks characterising these VSMC states is centred around the mRNA abundance profiles of marker genes, leaving us, currently, with a view only of the transcriptional component of the VSMC transcriptome.…”

Emerging Concepts of Vascular Cell Clonal Expansion in Atherosclerosis

“…Existence of distinct subpopulations of SMCs in the vascular wall that have greater propensity for proliferation when exposed to pathological stimuli may also explain the varying nature of atherosclerotic progression in different types of arteries. 51,53,56 Benditt et al 57 first identified clonal populations in human atherosclerotic lesions, presumed to be mostly SMCs, and described intimal masses as benign tumors. These results were based on X chromosome-linked (X linked) inactivation (using glucose-6-phosphate dehydrogenase) as a rudimentary marker for clonal analysis, which lacks cell-type specificity.…”

“…Existence of distinct subpopulations of SMCs in the vascular wall that have greater propensity for proliferation when exposed to pathological stimuli may also explain the varying nature of atherosclerotic progression in different types of arteries. 51,53,56…”

“…61,70 These plaque SCA1 + SMCs induce macrophage inflammation and are more proliferative than their SCA1 − counterparts. 56,60,61 Antibody staining and SCA1-GFP mice have been used to show that only ≈0.5% to 1% of medial SMCs express SCA1. 61 Interestingly, SCA1 + SM-derived cells from the atherosclerotic lesion had similar transcriptomic profiles to SCA1 + SMCs in the medial wall.…”