Modular plug-and-play crosslinking platform for precision-engineered hydrogels
SUMMARY Hydrogels are promising for biomedical applications due to their tunable properties and extracellular matrix (ECM)-mimicking capabilities, but available chemistries limit customization potential. We present a modular bioconjugation system using polyphenol radical crosslinking, where gallic acid (GA)-modified glycosaminoglycans rapidly crosslink with diverse biomolecules under blue light and riboflavin. This method enables the creation of customizable, cell-compatible matrices. We validate conjugation chemistry using 1 H NMR, size-exclusion chromatography, and gel electrophoresis. As proof-of-concept, we have created injectable hydrogels from GA-modified hyaluronic acid (HA-GA) crosslinked with various proteins. Mechanical properties are tunable via riboflavin concentration and light exposure. HCT116 colon cancer cells form spheroids in HA-GA-BSA, while other protein-containing hydrogels support dispersed growth, indicating enhanced cell adhesion. HA-GA hydrogels upregulate stemness markers compared to 2D culture, while protein incorporation reduces this effect. Matrix-activated Wnt signaling confirms functional biomolecule integration using HA-GA-Wnt3A matrices. This strategy enables tailored ECM design for tissue engineering and 3D cell culture.