Soluble perlecan domain i enhances vascular endothelial growth factor-165 activity and receptor phosphorylation in human bone marrow endothelial cells

BACKGROUND: Immobilized recombinant perlecan domain I (PlnDI) binds and modulates the activity of heparin-binding growth factors, in vitro. However, activities for PlnDI, in solution, have not been reported. In this study, we assessed the ability of soluble forms to modulate vascular endothelial growth factor-165 (VEGF(165)) enhanced capillary tube-like formation, and VEGF receptor-2 phosphorylation of human bone marrow endothelial cells, in vitro. RESULTS: In solution, PlnDI binds VEGF(165 )in a heparan sulfate and pH dependent manner. Capillary tube-like formation is enhanced by exogenous PlnDI; however, PlnDI/VEGF(165 )mixtures combine to enhance formation beyond that stimulated by either PlnDI or VEGF(165 )alone. PlnDI also stimulates VEGF receptor-2 phosphorylation, and mixtures of PlnDI/VEGF(165 )reduce the time required for peak VEGF receptor-2 phosphorylation (Tyr-951), and increase Akt phosphorylation. PlnDI binds both immobilized neuropilin-1 and VEGF receptor-2, but has a greater affinity for neuropilin-1. PlnDI binding to neuropilin-1, but not to VEGF receptor-2 is dependent upon the heparan sulfate chains adorning PlnDI. Interestingly, the presence of VEGF(165 )but not VEGF(121 )significantly enhances PlnDI binding to Neuropilin-1 and VEGF receptor-2. CONCLUSIONS: Our observations suggest soluble forms of PlnDI are biologically active. Moreover, PlnDI heparan sulfate chains alone or together with VEGF(165 )can enhance VEGFR-2 signaling and angiogenic events, in vitro. We propose PlnDI liberated during basement membrane or extracellular matrix turnover may have similar activities, in vivo.