Directing Phase Separation to Harness Biomaterial Microstructures for Controlled Cellular Behavior

The compositional heterogeneity and structural diversity of native extracellular matrix are essential in regulating cell behavior and promoting tissue regeneration. Thermoresponsive polysaccharide-based materials with tunable transition temperatures and phase-separated microstructure offer substantial opportunities in tissue engineering, drug delivery, and wound healing applications. To develop novel synthetic thermoresponsive polysaccharides, we employed versatile chemical routes to attach intrinsically hydrophobic adducts to the backbone of hydrophilic dextran and developed protocols to form hydrogels with defined microstructures. Systematically conjugating methacrylate moieties to the dextran backbone yielded a continuous increase in macromolecular hydrophobicity that induced a reversible phase transition whose lower critical solution temperature can be systematically modulated <i>via </i>variations in polysaccharides concentration, molecular weight, degree of methacrylation, ionic strengths, and Hofmeister salts. Photo-initiated radical polymerization permits facile chemical crosslinking and kinetic capture of phase separation, enabling the formation of hydrogels with defined microdomains. The resulting heterogeneous hydrogels feature tunable microstructures and exhibited both microspheres and continuous phases that promoted enhanced cell adhesion in 2D and interfacial-driven cell migration in 3D. Engineering macromolecular hydrophobicity with temperature-triggered phase separation of conventional hydrophilic, non-phase separating polysaccharides to generate heterogeneous hydrogels with controlled microstructures will find potential applications in chronic wound healing.