CD45 Is Sufficient to Initiate Endothelial-to-Mesenchymal Transition in Human Endothelial Cells—Brief Report

Endothelial-to-mesenchymal transition (EndMT) is a dynamic process in which endothelial cells acquire mesenchymal properties and in turn contribute to tissue remodeling and growth. Previously, we found EndMT associated with mitral valve adaptation after myocardial infarction. Furthermore, mitral valve endothelial cells collected at 6 months post-myocardial infarction expressed the pan-leukocyte marker CD45 and EndMT markers. Additionally, mitral valve endothelial cells induced to undergo EndMT with TGF (transforming growth factor)-β1 strongly coexpressed CD45 but not CD11b or CD14. Pharmacologic inhibition of the CD45 PTPase (protein tyrosine phosphatase) domain in mitral valve endothelial cells blocked TGFβ-induced EndMT. This prompted us to speculate that, downstream of TGFβ, CD45 induces EndMT. We activated the endogenous CD45 promoter in human endothelial colony forming cells (ECFCs) using CRISPR (cluster regularly interspaced short palindromic repeats)/inactive Cas9 (CRISPR-associated protein 9) transcriptional activation. Bulk RNA sequencing was performed on control ECFCs and CD45-positive ECFCs to identify transcriptomic changes. Three functional assays-cellular migration, collagen gel contraction, and transendothelial electrical resistance-were conducted to assess mesenchymal properties in CD45-positive ECFCs. Activation of the endogenous CD45 promoter in ECFC and 3 additional sources of endothelial cells induced expression of several genes implicated in EndMT. In addition, CD45-positive ECFCs showed increased migration, a hallmark of EndMT, increased collagen gel contraction, a hallmark of mesenchymal cells, and decreased cell-cell barrier integrity, indicating reduced endothelial function. CD45 is sufficient to incite an EndMT phenotype and acquisition of mesenchymal cell properties in normal human ECFCs. We speculate that CD45, through its C-terminal PTPase domain, initiates signaling events that drive EndMT.