Peritoneal M2 Macrophage-Derived Extracellular Vesicles From db/+ Heterozygous Mice Enhance Diabetic Wound Healing in db/db Homozygous Mice Model.

Diabetic wounds pose a major clinical challenge due to persistent inflammation, impaired angiogenesis and defective tissue remodelling. These defects stem partly from dysregulated macrophage polarisation, with sustained M1 dominance and impaired M2 transition hindering resolution and repair. This study evaluates extracellular vesicles (EVs) from IL-4-polarised M2 macrophages of db/+ heterozygous mice for promoting healing in db/db type 2 diabetic wounds. M2 macrophages showed reduced IL-6, elevated IL-10, downregulated pro-inflammatory genes (iNOS, TNF-α, IL-6) and upregulated anti-inflammatory markers (IL-10, ARG-1, TGF-β). EVs, isolated by precipitation, displayed characteristic cup-shaped morphology (mean diameter 100.5 ± 31.47 nm) by SEM/TEM and CD63, CD9 and CD81 expression. Following in vitro migration and viability assessments, EVs were coated onto 3D-printed carboxymethyl cellulose (CMC) scaffolds to enhance wound healing efficacy. In full-thickness dorsal wound model (n = 24 mice; groups: NC, CMC, CMCE, PC), 1 day post-treatment, CMCE treatment markedly increased NO in serum and wound fluid and elevated IL-10 concentrations in wound exudate. By Day 20, CMCE treated wounds exhibited complete closure, with extensive re-epithelialisation, collagen deposition, neovascularisation (> 10 vessels/HPF), reduced inflammation depth and well-organised crust formation. These results indicate that M2-macrophage-derived EVs from heterozygous donors, delivered via bioactive scaffolds, rapidly modulate early inflammatory signalling and drive regenerative healing in homozygous diabetic mice' wounds, supporting a scalable cell-free therapeutic approach.