Platelet-derived extracellular vesicles reprogram the wound microenviroment to promote pressure ulcer repair.

Pressure ulcers (PUs) are a major clinical challenge due to their chronic nature and limited treatment options. Platelet-derived extracellular vesicles (pEVs) have emerged as promising cell-free therapeutic agents due to their regenerative potential. This study investigates the therapeutic efficacy and safety of allogeneic pEVs isolated from pooled human platelet lysates (PL) in PUs healing.pEVs were enriched by size exclusion chromatography and characterized by dot blot, NTA, and super-resolution nanoimaging. Their safety and efficacy was assessed using an <i>in vitro</i> human 3D skin model, an <i>in vitro</i> permeation test, and an <i>in vivo</i> PU rat model. Tissue response was evaluated through histological scoring, immunohistochemistry, proteomics, and bioinformatic pathway analysis. pEV treatment enhanced cell viability, increased keratinocyte thickness, and modulated inflammatory profiles on the <i>in vitro</i> 3D skin model without triggering significant IL-1α release. <i>In vivo</i>, pEVs promoted skin thickness, granulation, early collagen deposition, angiogenesis, and M2 macrophage polarization in a dose- and sex-dependent manner. The <i>in vitro</i> permeation test revealed effective and time-dependent pEV skin penetration, especially in female rat skin explants. Although wound healing–related genes were detected in both control and treated samples, pathway enrichment analysis revealed distinct activation patterns, with treated samples showing enrichment in anti-inflammatory and regenerative pathways. pEVs derived from pooled allogeneic platelets represent a safe and standardized therapeutic approach for pressure ulcer repair. While effects on wound closure were variable, pEVs promoted granulation, angiogenesis, collagen deposition, and anti-inflammatory responses in a dose- and sex-dependent manner. Delivery optimization and investigation of sex-specific responses are essential for clinical translation.