Epigenetic and Phenotypic Modulation of Adventitial Fibroblasts in Atherosclerosis by Coronary Artery Risk Gene TCF21

The vascular adventitia has long been postulated to play a key role in the development of atherosclerosis, but the relevant adventitial cell populations and the mechanisms by which they influence atherosclerosis have remained elusive. To characterize relevant adventitial cells and their regulators, we have combined single cell multi-omic murine model and human multi-omic data with human genetics. Single cell transcriptomic and epigenetic analysis of both murine and human arterial tissue revealed a unique population of adventitial fibroblasts (AdvFib) that reside exclusively in the adventitia. Murine lineage tracing using a novel AdvFib-specific model revealed specific expansion of AdvFib in atherosclerosis. Cell-cell communication analysis demonstrated an increase in AdvFib-driven crosstalk with other cell types during early phases of atherosclerosis development. Furthermore, akin to phenotypic modulation in SMC, AdvFibs also undergo dramatic phenotypic alterations, activating pro-inflammatory (Ccl2/7/8, Lcn2, Cxcl12) and cell proliferation programs early in atherosclerosis, suggesting a key coordinating role in disease. Transcriptome and epigenome analysis revealed disproportionate enrichment for coronary artery disease (CAD) GWAS risk loci and genes in AdvFib, suggesting that AdvFib actively contributes to disease. Single cell data show that one CAD risk gene, TCF21 , is expressed primarily in AdvFib. Alteration of TCF21 appears to be central to AdvFib activation in atherosclerosis, as its expression also changed during atherosclerosis, with corresponding epigenetic remodeling in AdvFib enriched for Tcf21 binding motifs. AdvFib-specific TCF21 loss changed AdvFib proliferation and inflammatory cell recruitment, consistent with the higher TCF21 expression in human with lower risk of myocardial infarction. Mechanistically, through histone modification cut-and-tag, we demonstrated that TCF21 regulates AdvFib activation through epigenetic reprograming of key cytokine and cell fate regulators. Collectively, our results begin to establish a role for AdvFib in atherosclerosis development and highlight regulation of TCF21 in this process through epigenetic mechanisms.