Sulfation of Thiol-Maleimide Crosslinked Hydrogel Modulates Material Properties and Cell Biocompatibility

Abstract

Sulfation has become a critical but still underexplored factor influencing hydrogel properties and regulating cell behaviour. In this study, a thiol-maleimide crosslinked hydrogel system based on a poly(N-isopropylacrylamide) (pNIPAM) copolymer and dendritic polyglycerol (dPG) was developed to systematically investigate how sulfation modulates material properties, material affinity and cell viability. Sulfated (dPGS) and non-sulfated dPG were crosslinked with pNIPAM copolymers via thiolmaleimide coupling to form hydrogels under temperature suitable for cell cultivation. Sulfation increased the pore size (2.3fold), enhanced gel strength and (G′ 466 vs. 334 Pa at 37 °C) and retardation in enzymatic degradation, while maintaining similar thermoresponsive behaviour. FRAP revealed size-selective transport, with unchanged diffusion of small dextran (4 kDa) but 21% reduction of albumin diffusion efficient nutrient exchange with controlled protein retention for growth factor delivery. Biological evaluation demonstrated enhanced cell viability of A549 and HeLa cells in the sulfated hydrogel, while MCF-7 cells showed cell-type-specific response to sulfation. The dPGS-based hydrogel enhanced cell attachment and viability in a planar hydrogel substrate, and promoted spheroid formation and proliferation when cells were embedded within the 3D hydrogel network, mimicking a more physiologically relevant microenvironment. Taken together, the sulfated thiolcrosslinked hydrogel system offers precise control over thermoresponsive properties, molecular transport, and cell-typespecific interactions establishing hydrogel sulfation as a key design parameter for hydrogels in tissue engineering and regenerative medicine.