Understanding and accelerating the mechanisms of endothelial wound healing is of fundamental interest for biotechnology and of significant medical utility in repairing pathologic changes to the vasculature induced by invasive medical interventions. We report the fundamental mechanisms that determine the influence of substrate topography and flow on the efficiency of endothelial regeneration. We exposed endothelial monolayers, grown on topographically engineered substrates (gratings), to controlled levels of flow-induced shear stress. The wound healing dynamics were recorded and analyzed in various configurations, defined by the relative orientation of an inflicted wound, the topography and the flow direction. Under flow perpendicular to the wound, the speed of endothelial regeneration was significantly increased on substrates with gratings oriented in the direction of the flow when compared to flat substrates. This behavior is linked to the dynamic state of cell-to-cell adhesions in the monolayer. In particular, interactions with the substrate topography counteract Vascular Endothelial Cadherin phosphorylation induced by the flow and the wounding. This effect contributes to modulating the mechanical connection between migrating cells to an optimal level, increasing their coordination and resulting in coherent cell motility and preservation of the monolayer integrity, thus accelerating wound healing. We further demonstrate that the reduction of vascular endothelial cadherin phosphorylation, through specific inhibition of Src activity, enhances endothelial wound healing in flows over flat substrates.
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