The exploration to identify key molecules in the interaction between endothelial cells and mono-macrophages in atherosclerosis based on the single cell transcriptome

Coronary artery disease (CAD) is the leading cause of global cardiovascular deaths, and atherosclerosis (AS) is the characteristic pathological change in CAD. The interactions between monocytes/macrophages and endothelial cells play an essential role in the formation and progression of atherosclerotic plaques. In this study, we used published single-cell RNA sequencing datasets of calcified atherosclerotic core (AC) plaques and patient-matched proximal adjacent (PA) carotid artery tissue to characterize the single-cell profiles of carotid artery cells. We specifically analyzed endothelial cells (ECs) and monocyte-macrophage (MM) populations to identify cell subpopulations and signaling pathways associated with AS. By comparing cell-cell communication (CCC) between AC and PA, we identified signaling pathways and corresponding ligand-receptor pairs significantly altered in AS. For preliminary validation, Transwell co-culture experiments were conducted for 24 h with human umbilical vein endothelial cells (HUVECs) and human aortic endothelial cells (HAECs) co-cultured with M0, M1, or M2 macrophages. Through integrative analyses, we identified 15 cell clusters and 6 cell subpopulations closely related to AS. Among the altered pathways, MK (Midkine), GALECTIN (Galactose lectin), and SPP1 (Secreted phosphoprotein 1) showed significant changes in AC, with the MK pathway being the most prominent. Notably, the MDK-NCL pair was upregulated in the AS-associated EC and MM subpopulations. In vitro assays confirmed that MDK and NCL expression levels were significantly higher in the ECs and M1 macrophage co-culture group than in the M0 or M2 control groups. In conclusion, this work characterizes AS-related cell heterogeneity and identifies potential key signaling pathways and interactions, providing candidate targets for the prevention and treatment of CAD.