Vein transplantation of exosomes released from bone marrow stem cells in repairing of spinal cord injury

English Keywords:Exosomes  Spinal cord injury  Bone marrow stem cells

English Abstract:

  【Abstract】 Objectives: To investigate whether the exosomes derived from bone marrow stem cells(BMSCs) by intravenous infusion can promote the regeneration of injured spinal cord. Methods: BMSCs from adult rats were enriched and expanded by whole bone marrow culture in vitro, collecting the cell supernatant of P2 generation, by using the ExoQuick Precipitation for separation and purification of exosomes, the exosome markers, CD9, CD63 were analyzed by using Western blot. The MSCs-Exosomes morphology was assessed by transmission electron microscopy(TEM). The rats were induced by spinal cord strike machine to establish the spinal cord models． Exosomes of 500μl(final concentrate of 200μg/ml) were infused into tail vein． The rats were randomly divided into the following three groups(n=10/group): sham group, control group(SCI+PBS) and exosomes group(SCI+exosomes). Neurological functions were evaluated by the Basso-Beattie-Bresnahan(BBB) score and the slope plate experiment score at 1, 3, 7, 14, 21, 28 days after modeling. Histomorphological changes of spinal cord were observed at 28 days after modeling by hematoxylin-eosin(HE) staining and luxol fast blue(LFB) staining, Survivor number of nerve cells was observed at 28 days after modeling by Nissl staining. Results: TEM observation of MSCs-Exosomes revealed the presence of spherical vesicles with a typical cup-shape. The size distribution profile displayed a homogeneous population from 40-100nm, which was consistent with the literature description, MSCs-Exosomes were found to express high levels of CD9, CD63 by using Western blot. After modeling, BBB score and sloping plate experiment score in each time point were normal in sham group, but in control group and exosomes group were significantly lower than those in sham group(P<0.05), the scores of BBB in exosomes group were 6.30±0.95, 12.70±1.57, 16.60±1.08, 17.00±0.67 at 7, 14, 21, 28 days respectively after modeling, which were higher than those in control group of 2.50±1.08, 6.90±0.99, 10.50±0.85, 12.50±1.08(P<0.05) respectively, the scores of slope plate experiment in exosomes group were 43.00±3.50, 55.50±4.38, 62.50±2.64, 65.00±3.33 respectively at 7, 14, 21, 28 days after modeling, which were higher than those in control group of 34.00±3.16, 43.00±4.22, 49.00±4.59, 52.50±4.25(P<0.05) respectively． HE, LFB and Nissl staining showed nerve cells with normal morphology in sham group at 28 days after modeling, in control group， spinal cord porosis， disorder of myelin sheath and necrosis of nerve cells were found. The injured spinal cord was restored better and the number of neuronal survival was better in exosomes group than that in control group． TEM observation of MSCs-Exosomes revealed the presence of spherical vesicles, with a typical three-dimensional circular or cup-shape. The size distribution profile displayed a homogeneous population from 40-100nm with a complete lipid membrane structure in the peripheral region with low density content. Conclusions: Intravenous administration of MSCs-Exosomes promotes the restoration of injured spinal cord and improves motor functions， which plays a protective role in the repair of spinal cord injury．