Abstract 1044: Single Cell RNA-sequencing Of Endothelial Cells In Atherosclerosis Reveals A Single Phenotypic Transition To A Mesenchymal Cell State

Abstract

Introduction: Coronary artery disease (CAD) is the leading cause of mortality in the United States. Multiple cell types contribute to the atherosclerotic plaques causing CAD, influencing disease burden. Endothelial-to-mesenchymal transition (EndMT), the process by which endothelial cells (ECs) acquire characteristics of mesenchymal cells and loose canonical endothelial features, is correlated with an unstable plaque phenotype, although this transition remains poorly characterized. Hypothesis: A specific population of ECs may undergo EndMT, potentially contributing to the progression of atherosclerosis. Methods: Endothelial-specific lineage-tracing was induced in Tg Cdh5-CreERT2 , ApoE -/- mice, followed by 16 weeks of high-fat diet (HFD) to produce atherosclerosis. Aortic roots were collected from male and female disease-condition animals (n=13) and a baseline cohort that was not fed HFD (n=6). Single cell RNA sequencing was performed using 10X Genomics and analyzed with the Seurat package. RNAscope staining of sections (n=10 per genotype) was performed to characterize the lesion. Results: Multiple endothelial subtypes in the aortic root were identified. One population of arterial ECs, characterized by Klk10 expression, appears to give rise to a disease-specific population of ECs expressing Edn1 . These ECs appear to acquire a mesenchymal phenotype, consistent with the occurrence of EndMT. RNAscope staining revealed Edn1 + cells at the luminal surface and deep within the plaque. Upstream regulator analysis comparing Edn1+ and Klk10+ populations predict EndMT-associated transcription regulators including SNAI2, TWIST2, STAT3, SNAI1 and YAP1 as drivers of this transition. This analysis predicts upregulation of both fibrotic and cytokine signaling. In contrast to a previous study of carotid ECs under disturbed flow that identified multiple disparate EC phenotypes, we observed a single disease-associated EC transition. Conclusion: We identified ECs undergoing EndMT in a HFD-induced mouse model of atherosclerosis and determined that this transition is the primary EC cell state transition during disease. Thus, the process of EndMT may account for both mesenchymal and inflammatory changes observed in ECs during atherosclerosis.