Radical-free photopolymerizable composites of hyaluronic acid and gelatin for tissue engineering

Photopolymerization is widely used in tissue engineering and biofabrication to pattern specific geometries and modulate physical properties. Commonly employed photochemistries rely on a photoinitiator that generates reactive free radicals when exposed to light, which can lead to cytotoxic effects due to interactions with biomolecules and cellular components. To mitigate these issues, we have developed hyaluronic acid and gelatin derivatives of umbelliferone, which can form dimers thanks to cyclobutene ring formation when exposed to long-wavelength UV light (365 nm). These reactions occur efficiently with reduced cytotoxicity and without the need of a photoinitiator. Ligation to the biopolymers was carried out with the incorporation of a triethylene glycol or n-octyl linker that enhances the conformational flexibility of umbelliferone and contributes to improve the rheological properties. By heat annealing these derivatives, we produced double network hydrogels with various compositions. We assessed their physical properties using rheological and uniaxial compression tests, evaluated their cytocompatibility by encapsulating articular chondrocytes, and conducted preliminary printability tests to determine their suitability for injection and extrusion-based biofabrication. The materials exhibited good cytocompatibility and cell adhesion, were successfully extrudable using a pneumatic bioprinter while maintaining cell viability and were compatible with regulatory-approved steam sterilization. Due to their unique properties, these umbelliferone derivatives are well-suited for tissue engineering and biofabrication applications, offering crucial advantages for future clinical translation. Statement of significance: This study introduces a method for preparing novel bioinks from coumarin derivatives of hyaluronic acid and gelatin, key biopolymers in tissue engineering. These derivatives enable photoinitiator-free photocrosslinking without generating free radicals, thereby reducing cytotoxic risks and facilitating easier clinical translation compared to existing approaches. They are compatible with steam sterilization and show promise for extrusion-based techniques like bioprinting and injectability. Through mechanical characterization and biological assessments, the interactions between the biopolymers at different ratios and their effect on encapsulated cells were studied, providing insights for optimizing future tissue engineering applications.