Abstract
Previous studies showed that acellular nerve allografts (ANAs) have been successfully utilized in repairing peripheral nerve defects, and exosomes produced by stem cells are useful in supporting axon regrowth after peripheral nerve injury. In this study, exosomes from hair follicle epidermal neural crest stem cells (EPI-NCSCs-Exos) combined with ANAs were used to bridge facial nerve defects. EPI-NCSCs-Exos were isolated by ultracentrifuge, and were identified. After coculture, EPI-NCSCs-Exos were internalized into dorsal root ganglions (DRGs) and schwann cells (SCs) in vitro, respectively. EPI-NCSCs-Exos elongate the length of axons and dendrites of DRGs, and accelerated the proliferation and migration of SCs, and increased neurotrophic factor expression of SCs as well. The next step was to assign 24 Sprague Dawley male rats randomly and equally into three groups: the autograft group, the ANA group, and the ANA + EPI-NCSCs-Exos group. Each rat manufactured a 5-mm gap of facial nerve defect and immediately bridged by the corresponding transplants, respectively. After surgery, behavioral changes and electrophysiological testing of each rat were observed and assessed. At 90 days postoperatively, the retrogradely fluorescent tracer-labeled neurons were successfully observed on the injured side in the three groups. Morphological changes of facial nerve regeneration were evaluated by transmission electron microscopy and semithin toluidine blue staining. The results showed that nerve fiber density, nerve fiber diameter, and myelin sheath thickness in the ANA group were significantly worse than those in the other two groups (P < 0.05). No significant difference in nerve fiber density and myelin sheath thickness was observed between the autograft group and the ANA + EPI-NCSCs-Exos group (P = 0.14; P = 0.23). Our data indicated that EPI-NCSCs-Exos facilitate ANAs to bridge facial nerve defects and have the potential to replace autograft therapy in clinic.