Maleimide cross-linked bioactive PEG hydrogel exhibits improved reaction kinetics and cross-linking for cell encapsulation and in situ delivery.

Hydrogels, highly hydrated cross-linked polymer networks, have emerged as powerful synthetic analogues of extracellular matrices for basic cell studies as well as promising biomaterials for regenerative medicine applications.[1] A critical advantage of these artificial matrices over natural networks is that bioactive functionalities, such as cell adhesive sequences and growth factors, can be incorporated in precise densities while the substrate mechanical properties are independently controlled.[2–7] Polyethylene glycol (PEG) hydrogels represent the “gold standard” in this field due to their intrinsic low-protein adsorption properties, minimal inflammatory profile and history of safe in vivo use, ease in incorporating various functionalities, and commercial availability of reagents. In the present study, we explored the use of an alternative reactive cross-linking moiety for PEG hydrogels, the maleimide functional group. We demonstrate several advantages over other cross-linking chemistries, namely stoichiometric hydrogels with improved cross-linking efficiency, bio-ligand incorporation, and reaction time scales appropriate for clinical use with in situ gelation. The maleimide reactive group is extensively used in peptide bioconjugate chemistry because of its fast reaction kinetics and high specificity for thiols at physiological pH.[8] For these experiments, we used a 4-arm PEG-maleimide (PEG-4MAL) macromer and compared it to 4-arm PEG-acrylate (PEG-4A), 4-arm PEG-vinylsulfone (PEG4VS), and UV photo-cross-linked PEG-diacrylate (PEG-DA). Various cross-linking chemistries have been described to create bioactive hydrogel networks of PEG macromers, with Michael-type addition reactions and acrylate polymerization being the most widely utilized.[4] Cross-linking chemistry, gelation time, polymer network structure, and buffer conditions are important considerations when selecting a hydrogel crosslinking format for basic cell biology studies or regenerative