Exosomes derived from fibroblasts enhance skin wound angiogenesis by regulating HIF-1α/VEGF/VEGFR pathway

Abstract

Background Angiogenesis is vital for tissue repair but insufficient in chronic wounds due to paradoxical growth factor overexpression yet reduced neovascularization. Therapeutics physiologically promoting revascularization remain lacking. This study aims to investigate the molecular mechanisms underlying fibroblast-derived exosome-mediated angiogenesis during wound repair. Methods To assess the effects of fibroblasts derived exosomes on wound healing and angiogenesis, a full-thickness mouse skin injury model was established, followed by pharmacological inhibition of exosome secretion. The number and state of blood vessels in wounds were assessed by immunofluorescence, immunohistochemistry, hematoxylin–eosin staining, and laser Doppler imaging system. The high-throughput miRNA sequencing was carried out to detect the miRNA profiles of fibroblast-derived exosomes. The roles of candidate miRNAs, their target genes, and relevant pathways were predicted by bioinformatic online software. The knockdown and overexpression of candidate miRNAs, co-culture system, matrigel assay, pharmacological blockade, cell migration, EdU incorporation assay, and cell apoptosis were employed to investigate their contribution to angiogenesis mediated by fibroblast-derived exosomes. The expression of vascular endothelial growth factor A (VEGFA), vascular endothelial growth factor receptor 2 (VEGFR2), hypoxia-inducible factor 1α (HIF-1α), von Hippel–Lindau (VHL), and proline hydroxylases 2 was detected by western blot, co-immunoprecipitation, immunofluorescence, real-time quantitative polymerase chain reaction, flow cytometry, and immunohistochemistry. Furthermore, a full-thickness mouse skin injury model based on type I diabetes mellitus induced by streptozotocin was established for estimating the effect of fibroblast-derived exosomes on chronic wound healing. Results Pharmacological inhibition of exosome biogenesis markedly reduces neovascularization and delays murine cutaneous wound closure. Topical administration of fibroblast-secreted exosomes rescues these defects. Mechanistically, exosomal microRNA-24-3p suppresses VHL E3 ubiquitin ligase levels in endothelial cells to stabilize hypoxia-inducible factor-1α and heighten vascular endothelial growth factor signaling. MicroRNA-24-3p-deficient exosomes exhibit attenuated pro-angiogenic effects. Strikingly, topical application of exosomes derived from fibroblasts onto chronic wounds in diabetic mice improves neovascularization and healing dynamics. Conclusions Overall, we demonstrate central roles for exosomal miR-24-3p in stimulating endothelial HIF-VEGF signaling by inhibiting VHL-mediated degradation. The findings establish fibroblast-derived exosomes as promising acellular therapeutic candidates to treat vascular insufficiency underlying recalcitrant wounds.