Sulfation of thiol–maleimide crosslinked hydrogels 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 thiol–maleimide coupling to form hydrogels at temperatures suitable for cell cultivation. Sulfation increased the pore size (2.3-fold), enhanced gel strength (G′ 466 vs. 334 Pa at 37 °C) and retarded enzymatic degradation, while maintaining similar thermoresponsive behaviour. FRAP revealed size-selective transport, with unchanged diffusion of small dextran (4 kDa) but a 21% reduction in albumin diffusion after sulfation, indicating efficient nutrient exchange while enabling partial retention of larger proteins. Biological evaluation demonstrated the enhanced cell viability of A549 and HeLa cells in the sulfated hydrogel, while MCF-7 cells showed a 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. Overall, our findings identify sulfation as an effective handle to adjust thermoresponsive behaviour, macromolecular transport and cell-type-specific responses in thiol-crosslinked hydrogels, laying the basis for their use in advanced cell culture systems and, in the longer term, tissue-engineering-oriented applications.