Three-dimensional gelatin sponge culture potentiates MSC secretome to enhance full-thickness wound healing and induce hair follicle neogenesis via Wnt/β-catenin and TLR3/STAT3 activation in rats.

This study aimed to produce a high-quality mesenchymal stem cell-derived secretome (MSC-sec) using a three-dimensional culture system to enhance its therapeutic potential for skin wounds. Human umbilical cord-derived MSCs were cultured on optimally modified gelatin sponge (GS) scaffolds (ethanol-plus-polylysine), which maintained high cell viability. The secretome from these GS-cultured MSCs (GS-MSC-sec) contained significantly higher concentrations of key growth factors (KGF, VEGF, PDGF) compared to conventional 2D culture. In vitro, GS-MSC-sec enhanced fibroblast proliferation, migration, and angiogenesis. In a rat full-thickness skin wound model, concentrated GS-MSC-Section (5 ×) not only accelerated wound healing but also promoted wound-induced hair neogenesis. This enhanced regeneration was associated with upregulation of Wnt/β-catenin and TLR3/STAT3 signaling and downregulation of BMP/Smad signaling. These findings demonstrate that GS-MSC-sec possesses significant therapeutic potential for promoting skin wound healing and regeneration, with implications for treating alopecia.