Experimental Research On The Treatment Of Spinal Cord Injury In Rats By Transplantation Of Bone Marrow Mesenchymal Stem Cells Over-expressing SDF-1?

ObjectiveTo further study the migration experiments in vivo of BMSCs overexpressing SDF-1?and the effect on the treatment of SCI,which were based on its migration in vitro in the early stage of our research.Observing the migration of BMSCs overexpressing SDF-1?to the injured area of spinal cord,the recovery of the motor function in SCI rat hind limbs and the repair of pathological structure of the damaged spinal cord through the establishment of SCI rat models.Methods1.Primary BMSCs were attained from density gradient ultracentrifugation,and the cell phenotype was performed by flow cytometry in the second generation;BMSCs stably overexpressing SDF-1?system?SDF-1?-BMSCs?was constructed,and BMSCs with empty vector?null-BMSCs?and SDF-1?-silent?siRNA-BMSCs?were established as controls.2.The SCI rat models were builded by reformative Allen's method.The experimental animals were divided into six groups:Control group,Sham group,PBS group,null-BMSCs transplantation group,SDF-1?-BMSCs transplantation group and siRNA-BMSCs transplantation group.After successful building,the recovery of motor function in both hind limbs was performed by the BBB scale score.3.?1?The effects of BMSCs overexpressing SDF-1?on the morphology of injured rat spinal cord were analyzed by the HE staining of histopathology.?2?The proteins of SDF-1?and CXCR4 in the spinal cord area were determined by Western blot.?3?Under the fluorescence microscope,the migration of BMSCs to the damaged area of the spinal cord was observed.?4?Under the laser confocal microscopy,the spatial distribution of SDF-1?and CXCR4 labeled positive cells in the spinal cord was observed before and after SCI.Results1.BMSCs stably overexpressing SDF-1?system?SDF-1?-BMSCs?was successfully constructed.2.The SCI rat model was successfully constructed.On the 28^th day after SCI,the BBB scores of the null-BMSCs and the SDF-1?-BMSCs transplantation group were higher than the PBS group and the siRNA-BMSCs transplantation group,while SDF-1?-BMSCs group was higher than null-BMSCs group.3.?1?In the SDF-1?-BMSCs transplantation group,the number of neurons and glial cells in the injured spinal cord increased,the myelin regenerated,and the structure of white matter improved.?2?Compared with Control group and Sham group,the protein content of SDF-1?and CXCR4 were lower in PBS group,null-BMSCs transplantation group and siRNA-BMSCs transplantation group,but increased in SDF-1?-BMSCs transplantation group,and SDF-1?-BMSCs transplantation group was more than that in null-BMSCs transplantation group.?3?Compared with the null-BMSCs group and the siRNA-BMSCs group,the number of GFP?+?cells in the SDF-1?-BMSCs group was higher.?4?Under the laser confocal microscopy:1)The protein of SDF-1?was mainly expressed in the cytoplasm.In normal spinal cord area,SDF-1?positive cells were premarily distributed in the anterior horn;after SCI,they were finded in the white matter of the damaged area with the transplantation of BMSCs overexpressing SDF-1?.At the same time,the distribution could be revisibled in the gray.2)The protein of CXCR4was mainly expressed on the cell membrane.In normal spinal cord area,CXCR4positive cells were concentrated in the ependymal layer around the central canal of spinal cord;after SCI,they were finded in the gray and white through the transplantation of BMSCs overexpressing SDF-1?.Conclusions1.With the transplantation of BMSCs overexpressing SDF-1?after SCI,the recovery of the double hind limbs was the fastest and the effect was better,with the damaged spinal cord tissue better repaired.2.The high expression of SDF-1?and CXCR4 in the damaged spinal cord and the highest number of GFP?+?cells under the fluorescence microscope in the SDF-1?-BMSCs transplantation group are suggesting that BMSCs overexpressing SDF-1?may increase the homing rate of BMSCs through the SDF-1?/CXCR4biological axis.3.The spatial distribution of SDF-1?and CXCR4 before and after SCI was observed by laser confocal microscopy,suggesting that overexpression of SDF-1?may promote the multi-directional differentiation of BMSCs in vivo.However,whether the transplanted BMSCs differentiate into neurons and glial cells in the injured area of the spinal cord needs further verification.