Single-cell analysis reveals a novel regulatory mechanism of endothelial-to-mesenchymal transition
The transdifferentiation of endothelial cells (ECs) towards a mesenchymal-like phenotype, referred to as endothelial-to-mesenchymal transition (EndMT), is critical for embryonic development, and in adults it is one of the major contributors to the onset of diseases, including cancer, fibrosis and a number of cardiovascular disorders. Here we identified mitochondrial calcium signaling as a  key regulator of EndMT in three in vitro EndMT models, as well as in physiopathological conditions in vivo. Pharmacological inhibition of the mitochondrial calcium uniporter (MCU) prevents EndMT. Deletion of MCU in ECs confirms loss of EndMT during cardiac embryo development as well as in a hind limb ischemia mouse model. Together, our data provide evidence of a novel regulatory mechanism of endothelial transdifferentation, thus potentially allowing for the development of new therapeutic interventions for EndMT-related diseases. Overall design: To identify novel regulators of EndMT, we single cell profiled three in vitro EndMT models. EndMT was induced in freshly isolated human umbilical vein endothelial cells (HUVECs) by: 1) long-term TGF-b treatment, 2) ERG silencing (shERG) and 3) SNAI1 overexpression (SNAIOE). Each experimental condition was compared to control HUVECs (untreated, pLKO-empty and pRRL-empty, respectively). For each model, a pooled sample of two HUVEC donors was subjected to scRNA-seq. We then perfomed gene expression profiling analysis of each model individually and combined.