Endothelial cell flow-mediated quiescence is temporally regulated and utilizes the cell cycle inhibitor p27

Abstract

ABSTRACTBackgroundEndothelial cells regulate their cell cycle as blood vessels remodel and transition to quiescence downstream of blood flow-induced mechanotransduction. Laminar blood flow leads to quiescence, but how flow-mediated quiescence is established and maintained is poorly understood.MethodsPrimary human endothelial cells were exposed to laminar flow regimens and gene expression manipulations, and quiescence depth was analyzed via time to cell cycle re-entry after flow cessation. Mouse and zebrafish endothelial expression patterns were examined via scRNA seq analysis, and mutant or morphant fish lacking p27 were analyzed for endothelial cell cycle regulation andin vivocellular behaviors.ResultsArterial flow-exposed endothelial cells had a distinct transcriptome, and they first entered a deep quiescence, then transitioned to shallow quiescence under homeostatic maintenance conditions. In contrast, venous-flow exposed endothelial cells entered deep quiescence early that did not change with homeostasis. The cell cycle inhibitor p27 (CDKN1B)was required to establish endothelial flow-mediated quiescence, and expression levels positively correlated with quiescence depth. p27 lossin vivoled to endothelial cell cycle upregulation and ectopic sprouting, consistent with loss of quiescence.HES1andID3, transcriptional repressors of p27 upregulated by arterial flow, were required for quiescence depth changes and the reduced p27 levels associated with shallow quiescence.ConclusionsEndothelial cell flow-mediated quiescence has unique properties and temporal regulation of quiescence depth that depends on the flow stimulus. These findings are consistent with a model whereby flow-mediated endothelial cell quiescence depth is temporally regulated downstream of p27 transcriptional regulation by HES1 and ID3. The findings are important in understanding endothelial cell quiescence mis-regulation that leads to vascular dysfunction and disease.GRAPHICAL ABSTRACTHIGHLIGHTSDifferent quiescence stimuli lead to distinct transcriptional and functional quiescence profiles in endothelial cellsp27 is required for endothelial cell quiescence and depth is temporally regulated in a flow stimulus-dependent manner that correlates with p27 levels and flow-regulated repressorsHES1andID3p27 is expressed in endothelial cells according to flow magnitudein vivoand is functionally required for cell cycle regulation and sproutingin vivo