High Content Analysis uncovers functionally relevant metastable phenotypic states in human Endothelial Cells monolayers
SummaryEndothelial cells (EC) present distinct cell properties in different tissues. Heterogeneity within the same vascular bed has been proposed to underpin emergent behaviours in EC monolayers. Quantification and functional relevance of EC phenotypic variance are challenging to assess. Here we developed an EC profiling tool (ECPT) to uniquely enable single EC profiling within a monolayer providing spatial and relational information regarding cell proliferation, inter-endothelial Junctions and NOTCH activation. We used ECPT to characterise differential phenotypes in arterial, venous and microvascular EC populations. Our analysis highlighted extensive heterogeneity within individual monolayers and revealed VEGF-modulable metastability of NOTCH signalling which in turn regulates inter-endothelial junction’s stability. We suggest that accounting for adaptive emerging endothelial behaviours is necessary to develop revascularisation strategies for regenerative medicine and to design more effective EC-targeting drugs for cardiovascular diseases and cancer.