Dissecting VEGF-Induced Acute Versus Chronic Vascular Hyperpermeability: Essential Roles of Dimethylarginine Dimethylaminohydrolase-1

Abstract

Abnormal vascular leakiness plays crucial roles in several pathogenesis including cancer, macular degeneration, cardiovascular diseases and even in COVID-19-related pulmonary edema. Vascular endothelial cell growth factor (VEGF), also well-known as vascular permeability factor (VPF), activates VEGF receptor 2 (VEGFR2) and its downstream signaling leading to both acute and chronic vascular permeability. While many key downstream components are identified in cultured cells and murine models upon acute VEGF administration, the key molecular mediators of VEGF-induced chronic permeability remain unclear due to lack of appropriate in experimental model. Here we demonstrated that chronic VEGF exposure induces unique pattern of signaling as compared to that of acute exposure, including phosphorylation of VEGFR2, c-Src, and steady increases of nitric oxide (NO). Utilizing heat-inducible VEGF transgenic zebrafish (Danio rerio) and establishing an algorithm incorporating segmentation techniques to precisely quantify the extravasated dye, we monitored acute and chronic VEGF-induced vascular hyperpermeability in real time. Importantly, knockdown of dimethylarginine dimethylaminohydrolase-1 (DDAH1), an enzyme essential for NO generation, significantly altered downstream signaling pathways due to both acute and chronic VEGF exposure in human endothelial cells and the VEGF-inducible zebrafish permeability model. Finally, the effect of DDAH1 on VEGF-induced hyperpermeability was confirmed using DDAH1 transgenic mice as well as DDAH1 knockout mice by Miles assays, which exhibited increased and decreased leakage of Evan’s blue dye, respectively. Taken together, our data reveal novel divergent signaling of VEGF-induced acute versus chronic hyperpermeability and identify DDAH1 as a critical player and potentially a therapeutic target of vascular hyperpermeability-mediated pathogenesis.