HucMSC-derived extracellular vesicles promote peripheral nerve regeneration through direct axonal support and enhanced MerTK‑associated myelin debris clearance

Peripheral nerve injury (PNI) often leads to poor functional recovery, and effective treatments remain limited. Human umbilical cord mesenchymal stem cell-derived extracellular vesicles (hucMSC-EVs) have shown promise in promoting nerve repair, but the underlying mechanisms are not fully understood. A 10-mm sciatic nerve defect model was established in rats to evaluate the therapeutic potential of hucMSC-EVs. Single-cell RNA sequencing (scRNA-seq) was performed to identify responsive cell populations. In vitro, macrophages and Schwann cells were exposed to EVs with or without the MER proto-oncogene tyrosine kinase (MerTK) inhibitor UNC2250 or the MER/FLT3 dual inhibitor UNC2025. Phagocytosis assays were performed to evaluate myelin debris (MD) uptake, Western blot was employed to detect autophagosome formation and PI3K-Akt activation, and qRT-PCR together with immunofluorescence were used to assess M2 polarization and the repair phenotype. In vivo UNC2025 was carried out to confirm its functional necessity in the regenerative process. HE staining of major organs was performed to evaluate biosafety. HucMSC-EVs significantly promoted axonal regeneration and improved functional recovery. scRNA-seq revealed that hucMSC-EVs expanded MerTK⁺ macrophage subpopulation and repair Schwann cell subpopulation. In vitro, hucMSC-EVs upregulated MerTK in both cell types and enhanced M2 polarization, autophagosome formation, and phagocytosis in macrophages, while promoting the repair phenotype and MD interaction in Schwann cells. The selective MerTK inhibitor UNC2250 significantly attenuated EVs-enhanced phagocytosis in both cell types, as well as the EVs-induced repair phenotype of Schwann cells. HucMSC‑EVs increased PI3K‑Akt signaling; this effect was partially reversed by UNC2250 and more strongly by the dual MER/FLT3 dual inhibitor UNC2025, suggesting that MerTK contributes to but may not be the sole mediator of Akt activation. In vivo, UNC2025 attenuated the EVs-induced improvements in axonal regeneration and functional recovery. MD clearance was further confirmed by MBP immunofluorescence, which showed reduced MBP⁺ debris in the EVs group. HucMSC-EVs promote peripheral nerve regeneration and functional recovery through combined direct neurotrophic effects and enhanced MerTK-associated MD clearance in macrophages and Schwann cells. These effects are associated with activation of the PI3K-Akt pathway. Our findings support hucMSC-EVs as a promising therapeutic strategy for PNI, although the causal contribution of MerTK in vivo requires further validation.