Abstract Or100: Targeting Endothelial CD45 Ameliorates Endothelial-to-Mesenchymal Transition in Atherosclerosis

Background: Protein-tyrosine-phosphatase CD45 is exclusively expressed in all nucleated cells of the hematopoietic system but is rarely expressed in endothelial cells. Interestingly, our recent study indicated that activation of the endogenous CD45 promoter in human endothelial colony forming cells induced expression of multiple endothelial-to-mesenchymal transition (EndMT) markers. However, the detailed molecular mechanisms underlying CD45-driven EndMT and the therapeutic potential of manipulating CD45 expression in atherosclerosis are entirely unknown. Here, we aim to investigate whether endothelial CD45 genetic deletion reduced atheroma burden in ApoE-deficient mice fed with the western diet. Methods and Results: We generated a tamoxifen-inducible EC-specific CD45 deficient mouse strain (EC-iCD45KO) in an ApoE-/- background and fed with a Western diet (C57BL/6) for atherosclerosis and molecular analyses. We isolated and enriched mouse aortic endothelial cells (MAECs) with CD31 beads to perform single-cell RNA sequencing. Firstly, the absence of endothelial CD45 leads to reduced lesion development, foam cells, plaque macrophages, and expression of ICAM-1, which is negatively associated with atheroma development. Importantly, our single-cell data indicated that EC-specific knock-out CD45 a novel sub-population of EndoMT cells- EndMT2. It also showed an upregulation of EC marker genes (Cdh5, Flt1, Mecom) while a downregulation of SMC (Mef2c, Pde4d, Pdcd4) and EndMT marker genes (Fbln2, Fbln5, Isg15) in EndMT2 cells in EC-iCD45KO/ApoE-/- WD mice. Next, we found that CD45 deficiency led to decreased TGFβR 2-3 expression on MAECs which treated with oxidized low-density lipoprotein. In addition, we observed that the absence of endothelial CD45 led to the reduction of alpha-smooth muscle actin on MAECs which pretreated with TGFβ 1 or TGFβ 2 . Finally, we found that endothelial CD45 deficiency positively correlated with KLF2 expression on the mouse aortic root, which counters EndMT. Conclusion: These findings demonstrate that the loss of endothelial CD45 protects against EndMT-driven atherosclerosis, promoting KLF2 expression while inhibiting TGFβ signaling and EndMT markers. Thus, targeting endothelial CD45 may be a novel therapeutic strategy for EndMT and atherosclerosis.