| Objective: To create the relative traumatic chronic spinal cord injury（SCI） rodent animal modelling. To evaluate both Diffusion Tensor Imaging（DTI） and Diffusion Tensor Tracking（DTT） on exploring the animal modellings and the effect of the Bone marrow derived mesenchymal stem cells（BMSCs） transplantation. To invent the precise delivery strategy for transplantation into Post traumatic syringomyelia（PTS）. To evaluate the effect of different numbers of BMSCs on repairing chronic SCI. Methods: PSI-IH Impactor was used to initiate the contused SCI at T10 level. BBB score, MRI scanning, DTT and DTI were weekly used for evaluating. Magnetic nanoparticles, used as the marker for transplantation, were delivered into PTS under the guidance of coordinate. 8μl magnetic nanoparticles, twice and separately, were delivered.Complete bone marrow separation method was to apply for the isolation of BMSCs. Both flow cytometry and light microscope observation were used for the identification of BMSCs. DMEM, 4×105, 8×105 and 1×106 BMSCs were transplanted into the injury area of the modellings. BBB score, rotarod test, visual evaluation, MRI scanning, DTT, DTI, IF（immunofluorescence） and H&E staining were weekly used for determining the effect of BMSCs. After 8 weeks post transplantation, MAP2, Nestin, GFAP, Tublin, VEGF and BDNF were chosen as the IF marker for evaluating different numbers of BMSCs’ effect on restorating nerve cells, eliminating glial scar, rebuilding vascularization, trophic factors paracrine.Results: The modellings were successfully created without any sign of death or wound infection. The BBB score after 8w SCI showed4.21±0.35, 5.08±0.72, 5.79±0.88, 6.12±0.69, 6.39±0.55, and 6.44±0.62. MRI scanning suggested the formation of three types of PTS. The volume of of PTS post SCI in 6w were 0 mm3,（0.21±0.35）mm3,（2.21±0.44）mm3,（3.18±0.82）mm3,（3.27±0.79）mm3, and（3.31±0.73）mm3. DTI suggested both ADC and FA post SCI in 6w were（2351.91 ±780.73）×10-6mm2/s,（2033.22±720.11）×10-6mm2/s,（1972.10±710.31）×10-6mm2/s,（1604.22±720.51）×10-6mm2/s,（1550.62±656.11）×10-6mm2/s,（1549.71±660.73）×10-6 mm2/s, and 0.30±0.06, 0.31±0.07, 0.32±0.07, 0.33±0.09, 0.34±0.08, 0.34±0.08.The magnetic nanoparticles transplantation was successfully performed based on the calculated coordinates. The volume of PTS decreased from（5.71±0.21） to（3.23±0.36） mm3, after repeating the process, the PTS volume further decreased from（3.23±0.36） to（1.48±0.72） mm3.The result of flow cytometry suggested that the obtained cells are positive for specific MSCs markers CD105, CD90, CD44, and negative for hematopoietic markers（CD45, CD34）.The aforementioned BMSCs were successfully transplanted into the local injury area of the SCI animals. MRI scanning was performed to prove the successful performance. None of the rats died or showed the sign of infections or tumorigenesis. The functional test suggested all the cell transplanted groups showed the improved situation compared with DMEM group, the 1×106 BMSCs group showed the best results among four groups. All the cell transplanted groups showed the decreased injury area compared with DMEM group through the evaluation of visual evaluation, MRI scanning, DTT, DTI, HE and IF, the 1×106 BMSCs group showed the best results among four groups. Different numbers of BMSCs showed the different improved situation on restorating nerve cells, eliminating glial scar, rebuilding vascularization, trophic factors paracrine. The 1×106 BMSCs group showed the best results among BMSCs transplantation groups.Conclusions: The traumatic chronic SCI modelling is mature at 4w post injury. Both DTI and DTT are quantitative strategy for SCI diagnosis and evaluating SCI degree. Local BMSCs transplantation post chronic SCI is an effective treatment. BMSCs are able to versatilely repair SCI through restorating nerve cells, eliminating glial scar, rebuilding vascularization, trophic factors paracrine. The present study suggested the optimal cell numbers for repairing traumatic chronic rodent SCI is 5×106/kg. |
PDF Full Text Request