A Study On The Gene-Adjustment Mechanism Of The Combination Of Electroacupuncture And Transplanted BMSCs On Neural Repairation And Regeneration After Spinal Cord Injury

Objectives : To explore the feasibility of electroacupuncture promoting induction of bone marrow stromal cells(BMSCs) into neural cells; The combination of electroacupuncture and transplanted BMSCs initiate a new, safe, effective treatment and repairation of CNS diseases and to reveal the mechanism of gene adjustment.Animal: Eight young SD rats, weighing 100g each, were used for the cultivation of BMSCs.Eighty-one healthy adult SD rats, weighing 250±20g each, were used for the experiments. Female and male rats all can be concluded. The rats are randomly divided into five groups: (1) 9 rats, sham operation group; (2) 18rats, injection of phosphate buffered saline after Allen's method injury; (3)18. rats, transplantation of BMSCs after Allen's method injury; (4)18 rats, electroacupuncture treatment after Allen's method injury 24 hours later; (5) 18 rats, electroacupuncture treatmentafter transplantation of BMSCs 24 hours later. There were 3 time point in each group, 3d,7d,14d.Methods:1. BMSCs were cultured in vitro, observation the cells' morphology under the microscope.2. Functional outcome measurements using the combine behavioral score(CBS) performed 3d,7d and 14d post-transplantation.3. Light microscope were used to identify the morphology of the sections of injured spinal cord tissue; Immunohistochemical staining were used to identify the BrdU-labeled BMSCs.4. At 14 days of induction, immunohistochemical staining to detect whether the BrdU-labeled BMSCs expressed the astrocytic marker-GFAP(glial fibriliary acidic protein) or the neuron marker-NSE(neuro-specific enolase).5. Situ hybrization were used to identify the expression of BDNF,NGFmRNA after 3d,7d and 14d post-transplantation.Results:1. Original cultivation and subculturing of BMSCs:when original cells were cultured, cluster-like clones appeared, and maldistribution .After subculturing , BMSCs proliferated quickly.BMSCs were almost full of the culture flask 7days later, and the cells were spindle in shape and well-distributed.2. Neurological function improved in every group of every time point; BMSC group,electroacupuncture group and PBS group had significant difference(P<0.05); The difference between combination of electroacupuncture and transplanted BMSC group and PBS group was obvious(P<0.01); Neurological function improved better between combination of electroacupuncture and transplanted BMSC group and BMSC group,electroacupuncture group(P<0.05); BMSC group and electroacupuncture group had no significant difference(P>0.05).3. BrdU-labeled BMSCs were localized to the injury core and its boundary zone after transplantation; The local lesion of injury spinal cord was smaller in BMSC group and electroacupuncture group than in PBS group, moreover, the local lesion was the smallest in combination of electroacupuncture and transplanted BMSC group.4. The percentage of expression NSE and GFAP of BrdU-labeled BMSCs was: BMSC group was 7.2% and 1.5%, combination of electroacupuncture and transplanted BMSC group was 7.9% and 2.1%.5. Expression of BDNF,NGFmRNA in BMSC group and electroacupuncture group was higher than that in PBS group(P<0.05); The expression in combination of electroacupuncture and transplanted BMSC group was higher significantly than PBS group(P<0.01); Compared with BMSC group and electroacupuncture group, expression of BDNF,NGFmRNA was higher in combination of electroacupuncture and transplanted BMSC group(P<0.05); There was no difference between electroacupuncture group and BMSC group(P>0.05).Conclusion:Transplanted BMSCs can survive and localize to the injured area of spinal cord. Electroacupuncture can promote induction of transplanted BMSCs into neural cells. The combination of electroacupuncture and BMSCs transplantation can improve neurological function of spinal cord injuried rats.The combination of electroacupuncture and BMSCs transplantation can change the microenviroment of the injured area of spinal cord. Maybe it is the mechanism of decreasing the spinal cord injury, promoting restoration of injured spinal cord and improving motor function.