Studies in our laboratory indicate that collagenase from Clostridium histolyticum promotes endothelial cell and keratinocyte responses to injury in vitro and wound healing in vivo. We postulate that matrix degradation by Clostridial collagenase creates bioactive fragments that can stimulate cellular responses to injury and angiogenesis. To test this hypothesis, we performed limited digestion of defined capillary-endothelial-derived extracellular matrices using purified human or bacterial collagenases. Immunoprecipitation with antibodies recognizing collagens I, II, III, IV, and V, followed by mass spectrometry reveals the presence of unique fragments in bacterial, but not human-enzyme-digested matrix. Results show that there are several bioactive peptides liberated from Clostridial collagenase-treated matrices, which facilitate endothelial responses to injury, and accelerate microvascular remodeling in vitro. Fragments of collagen IV, fibrillin-1, tenascin X, and a novel peptide created by combining specific amino acids contained within fibrillin 1 and tenascin X each have profound proangiogenic properties. The peptides used at 10–100 nM increase rates of microvascular endothelial cell proliferation by up to 47% and in vitro angiogenesis by 200% when compared with serum-stimulated controls. Current studies are aimed at revealing the molecular mechanisms regulating peptide-induced wound healing while extending these in vitro observations using animal modeling.