Exosomes derived from menstrual blood-derived mesenchymal stem cells and treadmill training restore motor function following SCI by activating PI3K/Akt pathway

Abstract

Background: Spinal cord injury (SCI) remains a global challenge due to limited neural regeneration and functional recovery. Emerging therapies, such as mesenchymal stem cell-derived exosomes and exercise training, have shown promise, but their individual efficacy is insufficient. The synergistic effects of menstrual blood-derived mesenchymal stem cell-derived exosomes (MenSCs-Exo) and weight-supported treadmill training (WSTT) in SCI repair remain unclear. This study investigated their combined therapeutic potential and underlying mechanisms in SCI rats. Methods: A T10 spinal cord hemisection model was conducted in adult male Sprague-Dawley rats, which were randomized into five groups: Sham, SCI, Exo (200 µg MenSCs-Exo via tail vein injection every 48 h for 4 doses), TT (WSTT starting on day 3 post-SCI), and Exo + TT. Motor function was evaluated using the Basso, Beattie, and Bresnahan (BBB) scale and CatWalk XT® gait analysis. Histological assessments included hematoxylin and eosin (H&E) staining, Masson’s trichrome staining, immunofluorescence for β-tubulin III (Tuj1) and myelin basic protein (MBP), and transmission electron microscopy (TEM). Western blot analyzed fibrosis-related proteins (COL1, COL3, α-SMA) and PI3K/AKT pathway activation (p-AKT, PI3K, β-catenin, LEF1). Results: Combined Exo + TT significantly improved motor function compared to monotherapies. BBB scores in the Exo + TT group were higher than SCI controls from day 7, with marked differences at 4 weeks (P < 0.05). CatWalk analysis revealed enhanced hindlimb coordination, reduced dragging, and improved paw print parameters in Exo + TT rats. Histologically, Exo + TT reduced spinal cord cavitation, inflammation, and fibrosis (P < 0.01 vs. SCI), while promoting axonal (Tuj1) and myelin (MBP) regeneration with ordered structure. TEM showed preserved myelin lamellae and reduced axonal degeneration. Western blot confirmed decreased COL1, COL3, and α-SMA expression, along with upregulated p-Akt, PI3K, β-catenin, and LEF1 in Exo + TT rats (P < 0.01). Conclusion: MenSCs-Exo combined with WSTT synergistically enhances motor recovery after SCI by promoting tissue repair, reducing fibrosis, and activating the PI3K/Akt pathway. This cell-free therapy paired with rehabilitation exercise offers a novel strategy for SCI treatment.