An injectable hydrogel is a powerful carrier for therapeutic bioactive molecules. Here, an injected concentrated conditioned medium (CCM)-silk nanofiber composite hydrogel was developed to achieve the sustained release of multiple proteins and better wound healing. All the encapsulated proteins showed slow delivery for more than 9 days in vitro. Bioactive molecules such as transforming growth factor-beta1 (TGF-β1), insulin-like growth factor binding protein-1 (IGFBP-1), and platelet-derived growth factor-AB (PDGF-AB) were successfully released from CCM-loaded hydrogels, and they induced the proliferation and migration of fibroblasts and endothelial cells in a dose- and time-dependent manner. The differentiation of fibroblasts into myofibroblasts was also inhibited, implying less scar formation in vivo. Skin wound regeneration results indicated that the CCM-loaded hydrogel enhanced neovascularization, accelerated wound closure, and promoted hair follicle regeneration. The injectable multiple protein delivery system shows a promising application in skin wound repair.
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