Controlled delivery of proteins is a highly promising strategy to treat pathological conditions, including myocardial infarcts, bone defects, and skin lesions, as well as inflammatory, metabolic and neurological disorders. A major roadblock to effective protein therapeutics is the lack of biocompatible and injectable carriers that deliver proteins with high bioactivity and suitable release profiles. The objective of this proposal is to engineer hydrogel microspheres with controlled size and protease-dependent degradation profiles via microfluidics for therapeutic protein delivery. The maleimide-functionalized poly(ethylene glycol) (PEG-MAL) hydrogel system is a modular platform that incorporates protease-degradable or non-degradable cross-links, bioactive ligands, and therapeutic growth factors in a plug-and-play fashion allowing significant material tunability. In addition, PEG elicits minimal inflammatory reactions and has an excellent safety record. Importantly, the release rate of encapsulated proteins is controlled by protease-dependent cleavage of the hydrogel cross-links, providing a specific and triggerable release mechanism. Because the maleimide cross-linking reaction occurs much more rapidly than other hydrogel cross-linking chemistries, novel microfluidic-based polymerization approaches to generate microspheres of controlled size will be applied. By designing particles with different hydrogel cross-links and encapsulated proteins, a robust and flexible protein delivery platform with tuneable dosing and release profiles will be established. The overall objective will be accomplished via two specific aims Engineer PEG-MAL hydrogel microspheres with tuneable dosing and release profiles for BMP2 and VEGF, and evaluate in vivo delivery profiles and bone repair in a murine segmental defect model. This model will be also used in return phase to evaluate in vivo the role of the synergistic integrin and growth factor signalling in bone repair and vascularization.