Modulation of GAG-sulfation pattern in biohybrid hydrogels to tune growth factor release

Abstract

Event Abstract Back to Event Modulation of GAG-sulfation pattern in biohybrid hydrogels to tune growth factor release Passant Atallah1, Andrea Zieris1, Uwe Freudenberg1 and Carsten Werner1, 2 1 Max Bergmann Center of Biomaterials, Leibniz Institute of Polymer Research Dresden, Germany 2 Center for Regenerative Therapies Dresden, Technische Universität Dresden, Germany Introduction: GAG based hydrogels that modulate key parameters of the extracellular matrix were previously developed showing long term delivery of signaling molecules[1]. In here, the sulfation patterns of the GAG building blocks govern the electrostatic complexation of biomolecules and thus gradual desulfation should allow for modulating the release profiles of cytokines from the hydrogels. To explore options related to this, selectively desulfated heparin derivatives were prepared, thoroughly characterized, and covalently converted with star-shaped poly (ethylene glycol) into binary polymer networks and analyzed for cytokine release and cellular response in vitro and in vivo. Methods: Different solvolytic desulfation procedures yielded N-desulfated, 6O- N- heparin, 6O-desulfated and 2O-6O-N- (completely) desulfated heparin. All the heparin derivatives were thoroughly analyzed for regioselectivity of desulfation by 1H NMR, particle size analysis by asymetrric field flow fractionation, residual sulfate content by elemental analysis and anticoagulant activity by factor Xa based heparin assay. Hydrogels with graded sulfation pattern were subsequently prepared and the impact of the variation of the net negative charge on network properties was evaluated by rheometry and swelling measurements. Vascular endothelial growth factor (VEGF 165) and fibroblast growth factor 2 (FGF-2) were non-covalently conjugated to the heparin component of the hydrogel and the initial binding affinity of cytokines and their subsequent release was analyzed by ELISA[2]. The influence of the different functionalized matrices to migration and morphology of human umbilical vein endothelial cells (HUVEC) and the modulation of wound healing in diabetic mice has been analyzed. Results and discussion: Desulfated heparin chains were systematically characterized to confirm the chemical properties of the precursors complexed with the hydrogels. Analysis of physical network properties showed the influence of the charge density as well as the sulfation site specific effect on the elasticity and network defects of the formed hydrogels. Hydrogels with variable sulfation pattern show widely tunable release profiles for FGF-2 and VEGF165. In-vitro studies showed that the increased VEGF release from desulfated heparin hydrogels resulted in more pronounced proangiogenic response of HUVECs by enhanced cell migration and tubular formation. Similarly, hydrogels transplanted in db/db mice promoted wound angiogenesis, a critical stage in the wound healing process. Conclusion: Tailoring the sulfation pattern of GAG based hydrogels enables control over cytokine delivery and therefore cellular response in vitro and in vivo. The resulting new class of cell-instructive polymer matrices with tunable GAG sulfation will be instrumental for multiple applications in biotechnology and medicine. Figure 1: Schematic design of biohybrid GAG- star PEG hydrogels with regioselective desulfation resulting in modulation of cytokine release. This work was supported by the Deutsche Forschungsgemeinschaft through grant nos. SFB-TR 67/A10