Constructing EGF mRNA-Enriched Extracellular Vesicles Based on the AAVS1 Safe Harbor Site to Promote Skin Wound Healing

Skin wound healing is a complex biological process that requires the coordinated regulation of cell proliferation, migration, and extracellular matrix (ECM) remodeling. Epidermal growth factor (EGF) plays a key role in this process, but its clinical application is limited by its rapid degradation at the wound site. Extracellular vesicles (EVs), as natural nanocarriers, can protect nucleic acids from degradation and enhance their bioavailability. In this study, using CRISPR/Cas9 technology, we site-specifically integrated the EGF gene carrying the TPA signal peptide into the AAVS1 safe harbor site of 293F cells, generating a cell line that stably secretes EVs enriched in EGF mRNA. Characterization and in vitro and in vivo functional evaluation of these engineered EVs (293F-EGF-EV) demonstrated that they significantly promoted fibroblast proliferation and migration and inhibited excessive Collagen production. In a rat skin defect model, 293F-EGF-EV promoted wound recovery. High-concentration 293F-EGF-EV focused on "high-quality repair," such as promoting angiogenesis, hair follicle regeneration, and epidermal structural remodeling. Low-concentration 293F-EGF-EV favored "high-efficiency closure", such as reducing scar area. This study offers new insights into skin wound treatment.