Micron-scale spatially patterned, covalently immobilized vascular endothelial growth factor on hydrogels accelerates endothelial tubulogenesis and increases cellular angiogenic responses.

Spontaneous formation of endothelial tubules was restricted to patterned micron-scale regions presenting cell adhesion ligands and angiogenic signaling protein on poly(ethylene glycol) hydrogels. Arginine-glycine-aspartic acid-serine (RGDS), an integrin ligand, and vascular endothelial growth factor (VEGF), a rate-limiting signaling protein involved in angiogenesis, were covalently bound through photopolymerization via laser scanning lithography to the surface of poly(ethylene glycol) hydrogels in patterned micron-scale regions. Endothelial cells cultured in this restricted environment underwent accelerated tubulogenesis, forming endothelial tubes within 2 days, whereas cells cultured on larger patterned areas remained spread and did not form tubules by day 2. Tubules formed in 2 days on RGDS and VEGF patterns were observed to possess lumens; however, tubule-like structures formed in 2 days on RGDS-only control patterns did not have observable lumens. Additionally, tubules that formed on restricted areas of RGDS and VEGF expressed more VEGF receptor 1, VEGF receptor 2, and ephA7 surface markers, in addition to higher expression of laminin, than cells remaining spread on wide patterned lines. This work reports spatial control and acceleration of endothelial tubule formation using biocompatible hydrogel materials to allow the formation of highly organized vascularized tissues.