Regular Article VASCULAR BIOLOGY NRP1 acts cell autonomously in endothelium to promote tip cell function during sprouting angiogenesis

In vertebrates, organ development, homeostasis, and repair rely on properly perfused blood vessel networks. The first blood vessels in the embryo are assembled from single-cell precursors, which coalesce into a honeycomb-shaped vascular plexus in a process known as vasculogenesis. This primitive plexus then expands through sprouting and remodeling to supply all tissues and organs in a mechanism termed angiogenesis. Angiogenesis is normally quiescent in adults but can be reactivated, for example during tissue repair in wound healing, in ischemic eye disease, or during solid tumor growth. Angiogenesis depends on the vascular endothelial growth factor (VEGF)-A, which binds several different receptors on endothelial cells. The tyrosine kinase receptor VEGFR2, also known as FLK1 in mouse and KDR in humans, is the main signal transducing VEGF-A receptor in endothelial cells; it stimulates endothelial cell proliferation and migration through a plethora of downstream signaling events, and loss of VEGFR2 therefore halts blood vessel formation by vasculogenesis early during embryogenesis. After vasculogenesis, VEGFR2 promotes angiogenesis by acting in the specialized endothelial tip cells that head vascular sprouts and extend filopodia to sense growth factor gradients composed of VEGF-A isoforms with different matrix affinities. Consistent with an important role for VEGFR2 in endothelial guidance, high levels of VEGFR2 confer a competitive advantage to endothelial cells as they negotiate the tip cell position with their neighboring cells. In cultured human endothelial cells, VEGFR2 forms complexes with NRP1, a nontyrosine kinase receptor for the VEGF165 isoform of VEGF-A. Complex formation is thought to promote VEGF165 signaling through VEGFR2 and activate signaling pathways involved in cell migration and angiogenic sprout formation. The VEGF165mediated interaction between NRP1 and VEGFR2 may occur when both receptors are coexpressed on the same endothelial cell or on neighboring endothelial cells (homotypic interaction). Alternatively, NRP1 may be expressed on nonendothelial cells that interact with VEGFR2-expressing endothelial cells (heterotypic interaction). For example, it has been proposed that endothelial VEGFR2 interacts with tumor cell NRP1 in trans to stimulate VEGFR2 phosphorylation. Consistent with the idea of heterotypic roles for NRP1 in angiogenesis, treatment with a soluble dimerized form of NRP1 can rescue defective vascular outgrowth from paraaortic splanchnopleural explants of NRP1-deficient mouse embryos in vitro. However, the relevance of nonendothelial NRP1 for angiogenesis in vivo was not established in these studies. In the mouse, NRP1 overexpression leads to the formation of excess blood vessels that are leaky and hemorrhagic, causing