Accelerated Endothelial to Mesenchymal Transition increased fibrosis via deleting Notch signalling in wound vasculature

Skin wound healing in adults is characterised by a peak of angiogenesis followed by regression of the excessive vasculature in parallel with collagen deposition and fibrosis in the wound. We hypothesized that regressing vessels in healing wounds were in fact entering an endothelial to mesenchymal transition (EndMT) contributing to scarring. Using vascular specific fate tracking (Cdh5-creERt2/ROSA-YFP mice), full-thickness excisional wounds were analysed to reveal a time dependent transition from endothelial phenotype characterised by VE-Cadherin, CD31 and CD34 towards a mesenchymal phenotype characterised by alpha-smooth muscle actin and FSP1 expression. We next conditionally ablated RBPJ in the vasculature (Rbpjfl/fl/Cdh5-creERt2ROSA-YFP) to evaluate the role of canonical Notch signalling in this process. EndMT was clearly accelerated following loss of Notch signalling within the vasculature. The acceleration of EndMT resulted in delayed wound healing, increased fibrosis and extensive scar tissue formation, with the rapid loss of key endothelial genes and proteins and upregulation of mesenchymal protein expression (α-smooth muscle actin & fibroblast specific protein-1) in vessels. Our findings here uncover a cellular contributor to skin wound scarring through the process of EndMT in skin wounds and demonstrate the importance of Notch signalling in regulating this critical process during healing