Neuropilin-1 regulates attachment in human endothelial cells independently of vascular endothelial growth factor receptor-2

Abstract

IntroductionVascular endothelial growth factor (VEGF, also named vascular permeability factor, VPF) is a key regulator of physiologic and pathologic angiogenesis, and a molecular target for effective therapeutic intervention. [1][2][3][4][5][6] The biologic activities of VEGF are mediated, for the most part, by 2 receptor tyrosine kinases, Flt-1 (VEGFR-1) and KDR (VEGFR-2). 7,8 VEGFR-1 and VEGFR-2 are essential for fetal angiogenesis, and mouse embryos null for either receptor die in utero between days 8.5 and 9.5. 9,10 Compelling evidence indicates that VEGFR-2, and not VEGFR-1, is the major mediator of VEGF-induced proliferation and migration of endothelial cells, despite both receptors having high affinity binding to VEGF. [11][12][13][14] Chimeric fusion receptors containing the intracellular domain of VEGFR-1 or VEGFR-2 and the extracellular domain of other receptors provided evidence that VEGFR-1 can serve as a competitive inhibitor of VEGFR-2 mitogenic signals. 13,14 Gene targeting studies have shown that whereas hematopoietic and vascular development are severely impaired in VEGFR-2 null mice, 10 overgrowth of endothelial cells and disorganized blood vessels are the cause of death in VEGFR-1 null embryos, 9 providing additional evidence for VEGFR-1's role as a negative regulator of VEGF function. Besides these canonical VEGF receptors, the angiogenic roles of VEGF are modulated by the co-receptor molecules neuropilin NRP-1 and NRP-2. 15,16 NRP-1 and NRP-2, originally identified as receptors for class-3 semaphorins mediating neuronal guidance, 17,18 were subsequently found to bind several isoforms of VEGF and to form complexes with VEGFR-1 and VEGFR-2. [19][20][21][22] When coexpressed with VEGFR-2, NRP-1 enhanced the binding of VEGF to VEGFR-2 and enhanced VEGF-mediated chemotaxis. 16 When complexed with VEGFR-1, NRP-1 prevents the binding of VEGF to NRP-1, providing evidence for a common surface for interaction of NRP-1 with either VEGFR-1 or VEGF. 23 Consistent with NRP-1 playing an important role in regulating VEGF availability and function, NRP-1-deficient mice die at the embryo stage with defects in the heart, vasculature, and nervous system. 24,25 Transgenic mice overexpressing NRP-1 also exhibited abnormalities in the cardiovascular and nervous systems, contributing to embryonic lethality. 26,27 NRP-1 knockin mice expressing NRP-1 mutants that bind either VEGF or class 3 semaphorins demonstrated that VEGF/NRP-1 signaling is required for vascular development, whereas semaphorin-3A/NRP-1 signaling is required for neuronal, but not vascular, development. 28 In addition, endothelial cell-specific NRP-1 null mice displayed marked vascular abnormalities, confirming that NRP-1 expression in endothelial cells is essential for normal vascular development. 28 NRP-1 is a single spanning transmembrane protein with only a short intracytoplasmic tail of 40 amino acids. Deletion studies indicated that semaphorin function in neuronal cells is not dependent upon the intracellular domain of NRP-1 but requires plexi...

Galectin-1, a novel ligand of neuropilin-1, activates VEGFR-2 signaling and modulates the migration of vascular endothelial cells

“…As shown in Figure 4a, knockdown of NRP1 completely abolished Gal-1-induced HUVEC migration, but had only a marginal effect on Gal-1-induced HUVEC proliferation. Silencing of NRP1 expression also led to a marked decrease in the adhesion of HUVECs to Gal-1 (Figure 4b), and significantly decreased EC attachment to laminin and fibronectin, a result that is consistent with the reported decrease in the adhesive ability to the extracellular matrix of HUVECs transfected with NRP1 siRNA (Murga et al, 2005). As described earlier, Gal-1 binds both a 7 b 1 and a 5 b 1 integrins, and regulates smooth muscle cell differentiation and tumor cell growth (Gu et al, 1994;Fischer et al, 2005).…”

“…Silencing of VEGFR-2 expression and inhibition of JNK activation have only minimal effects, however, on the adhesion capacity of endothelial cells to Gal-1 (Supplementary Figures S4 and S5). Previous studies have reported that NRP1 alone can mediate HUVEC adhesion in a VEGFR-2-independent manner (Murga et al, 2005). Additional studies will be required to clarify the mechanisms of action of NRP1 in Gal-1-enhanced endothelial cell adhesion.…”

Biomarkers of angiogenesis and their role in the development of VEGF inhibitors

“…Additional co-receptors include neuropilins that have traditionally been implicated in cell guidance (Kawasaki et al, 1999) and increased binding of VEGF to its signalling receptor (Soker et al, 1998). However, recent data have suggested that NRP-1 may regulate EC function independently of VEGFR-2 (Murga et al, 2005), and that VEGF 121 can directly interact with NRP-1 without forming an NRP-1 -VEGFR-2 complex (Pan et al, 2007). Vascular endothelial growth factor-A interacts with both VEGFR-1 and VEGFR-2 to mediate angiogenesis, whereas VEGF-B and PlGF have high affinity for only VEGFR-1.…”

Cilengitide downmodulates invasiveness and vasculogenic mimicry of neuropilin 1 expressing melanoma cells through the inhibition of αvβ5 integrin

“…Since we and others have suggested a role for NRP-1 in endothelial or tumour cells independent of other VEGFRs, 16,32,33 we investigated whether cilengitide might modulate the response of M14-N cells to VEGF-A, using an in vitro chemotactic assay. Interestingly, cilengitide was found to inhibit the chemotactic response to VEGF-A, whereas it did not affect cell migration in response to human fibroblast conditioned medium (Fig.…”