(259a) Covalently Bound VEGF Upregulates VEGFR-2 Migration Pathway Signaling in HUVECs

The binding of vascular endothelial growth factor (VEGF) to VEGF receptor-2 (VEGFR-2) begins a signaling cascade that ultimately leads to the sprouting and formation of blood vessels. Studies conducted in vivo have found that the structure of the blood vessel network depends on the affinity of VEGF for the extracellular matrix (ECM). Soluble VEGF signaling has been linked to the formation of large, tumor-like blood vessels, while matrix bound VEGF signaling leads to smaller, capillary-like networks. However, the molecular signaling involved that result in these differences have not been thoroughly investigated. Utilizing self assembled monolayers (SAMs) on gold, surfaces were engineered that are capable of both covalent and electrostatic immobilization of VEGF. Soluble and bound VEGF activated tyrosine residue 1175 of VEGFR-2. This phosphorylation site is typically associated with the proliferation pathway. Both covalently and electrostatically bound VEGF increased activation of tyrosine residue 1214 of VEGFR-2, a residue responsible for initiating the migration pathway. Covalently bound VEGF sustained the activation up to 60 minutes. Meanwhile, soluble VEGF did not activate Y1214 as strongly, and for not as long. Covalently bound VEGF sustained activation of p38 and cdc42 over the soluble control. Electrostatically bound VEGF also activated p38, but not for the extended time period of covalently bound VEGF. Taken together, these results indicate that covalently binding growth factors to a biomaterial surface alters their signaling activities. Sustained phosphorylation of the receptor is a result of a ligand that remains bound to the surface, and is not internalized with the receptor.