Bio-based heparin-functionalized hydrogels for controlled VEGF-release II: Radical-initiated cross-linking

Purpose Presently insufficient oxygen and/or nutrient supply in tissue engineered grafts resulting in improper integration or death of encapsulated cells remain current limitations for clinical translation. [1] One strategy to overcome this is the controlled release of proangiogenic growth factors to induce vascularization and thereby bio-integration of the scaffold. Here we describe the preparation and characterization of hydrogels with tunable physico-chemical properties based on radical cross-linkable gelatin and heparin with application perspective as a tissue engineering scaffold allowing for storage and controlled release of proangiogenic growth factors. Methods Cross-linkable gelatin and heparin derivatives were prepared by reaction of the biopolymers with methacrylic anhydride. [2] Their degrees of methacrylation were quantified by nuclear magnetic resonance spectroscopy. Chemical cross-linking of hydrogels based on gelatin methacrylamide (9 % w/w) and heparin methacrylate (1 % w/w) was achieved by thermal-induced radical cross-linking in the presence of a water-soluble redox initiator system (APS Ammoniumperoxodisulfate / TEMED N,N,N',N'-Tetramethylethylenediamine). Hydrogels were characterized concerning gel yield and equilibrium degree of swelling by gravimetric analysis. Viscoelastic properties were tuned by degree of modification of gelatin and characterized using rheological measurements. Release of VEGF165 (vascular endothelial growth factor) from the hydrogels was quantified by ELISA (enzyme-linked immunosorbent assay). Furthermore cytocompatibility of hydrogels was investigated with human primary endothelial cells. Results We will present results on biopolymer derivatization and physico-chemical properties of hydrogels of different compositions. Three gelatin derivatives with methacrylation-degrees between 0.78 mmol/g and 1.45 mmol/g and methacrylated heparin with 0.17 mmol methacrylate groups per gram were prepared. A suitable initiator concentration range for cross-linking of hydrogels consisting of the different derivatives was found by varying the TEMED / APS ratio between 0.0 and 0.5 and the APS concentration between 0.025 M and 0.100 M. We show results on the impact of gel composition, cross-linking time, initiator concentration, and composition on gel yield, equilibrium degree of swelling, and viscoelastic properties. The gels had lower swelling capacities and higher mechanical stiffness's with higher methacrylation degrees of gelatin, whereas gel yield of all preparations were comparable. Furthermore we observed a significant effect of hydrogel drying on its swelling behavior. The swelling capacity was on average halved for all dried hydrogels compared with hydrogels swollen directly after preparation. We will also show effects of gel composition on release of VEGF. Finally, we aim to show results on the reaction of HDMVEC (human dermal microvascular endothelial cells) towards the hydrogel system. Conclusion Chemically cross-linked hydrogels composed of gelatin methacrylamide and heparin methacrylate have tunable physico-chemical properties. Hence this system might be suitable as a tissue engineering scaffold for storage and controlled release of growth factors.