PROJECT SUMMARY Chronic wounds are a serious problem that affects the health and quality of life of patients with diabetes and cardiovascular diseases. These non-healing ulcers are characterized by prolonged inflammation, poor vascularization, callus formation, and infection. Advanced wound care is estimated to cost over $10 Billion annually in the U.S. and is expected to increase as our population ages. A variety of advanced wound care products have been developed to deliver cellular growth factors, particularly fibroblast growth factors (FGFs), to the wound site to treat debilitating non-healing ulcers. However, poor efficacy has frequently been reported in clinical studies involving FGF delivery to wounds due to the following major obstacles: 1) the poor stability and proteolytic susceptibility of growth factors in vivo, 2) the difficulty of providing a controlled and sustained delivery of the growth factors, and 3) a lack of biomarkers to evaluate efficacy and to guide the development of new therapies. The overall objective of the proposal is to address these challenges through an integrative effort to develop a hyperstable and bioactive wt-FGF1-FGF2 dimer for effective healing of diabetic skin wounds. The efficacy of this new therapeutic approach will be monitored by quantitative in vivo metabolic imaging. The overall objective of this proposal is achieved by: 1) designing of a hyperstable and bioactive FGF1-FGF2 dimer and its hyperstable variants; 2) developing the injectable anionic hydrogels made of gelatin-modified poly(oligo ethylene glycol monoacrylate-co-acrylic acid) – P(OEGA-co-AA), which can provide sustained release of the different variants of the FGF1-FGF2 dimer; and, 3) to monitor metabolic of wt-FGF1-FGF2 dimer and its hyperstable variants using simultaneous acquisition of multiphoton images of NADH and FAD auto fluorescence, as well as collagen SHG intensity. Further, non-invasive Diffuse Reflectance Spectroscopy (DRS) will be used to evaluate angiogenesis activity, plausibly triggered by the FGF2 component of the designed FGF1-FGF2 dimer and its variants.