Experimental Study On Transplantation Of Bone Marrow Mesenchymal Stem Cells For Spinal Cord Injury

The treatment of spinal cord injury is always a stubborn problem for neurosurgeons because nerve cell cannot regenerate by itself and the glia scar can prevent the axonal regeneration. Therefore, to decrease the neuron death and formation of scar and to increase axonal regeneration are the keys for clinical treatment. Mesenchymal stem cell mainly included the advantage such as, convenience to draw the materials, simple cultivation,and could autograft without the advantage of immunological rejection etc,and convert toward nerve cells under special condition in vivo or in vitro.Transplantation of bone marrow mesenchymal stem cell into injured spinal cord tissue can reduce the defection degree of neural function and promote the recovery of neural function. Thus, it may become a new approach for clinical treatment of spinal cord injury.The study is composed of three parts:1. Experimental study on targeted inducement of human bone mesenchymal stem cells to neurons2. The effects of different revulsants on the differentiation of human bone mesenchymal stem cells into neurons3. The Research on the treatment of transplantation of induced differentiated Human Bone Marrow Mesenchymal Stem Cells on spinal cord injuryPart 1. Experimental study on targeted inducement of human bone mesenchymal stem cells to neurons[Objective]: To establish a systemic method of isolation, culture and inducement of human bone mesenchymal stem cells to neurons in vitro.[Methods]: The human bone mesenchymal stem cells, which were isolated from bone marrow with the lymphocyte separate liquid, cultured and expanded in vitro, were induced by combined neuro-revulsant. The morphologic change of cells was being observed, and the expression of surface sign of induced cell was identified by immunocytochemistry during inducing period. [Results]: Human bone mesenchymal stem cells could be separated, cultured and expanded in vitro. After being induced by combined neuro-revulsant 48h, some of cells expressed neuronal marks: cellular shrinkage and neurite growth. By immunochemical staining cells showed nestin(+) or NF(+), and GFAP(-).[Conclusion]: hMSC of isolated culture in vitro can soon differentiate into sufficient neuron-like cell in the nontoxic effect of complex nerve inductor. Thus a systemic empirical method of extrasomatic isolation, culture and induction of hMSC is established in this experiment.Part.2 The effects of different revulsants on the differentiation of human bone mesenchymal stem cells into neurons[Objective]: To find a relatively ideal revulsant and method of differentiation of human bone mesenchymal stem cells into neurons.[Methods]: The human bone mesenchymal stem cells, which were isolated from bone marrow with the lymphocyte separate liquid, cultured and expanded in vitro, were induced by different combined neuro-revulsant. The morphologic change of cells was being observed during inducing period, the differentiation of induced cells was identified by immunocytochemistry, and data was analyzed by SPSS10.0.[Results]: Human bone mesenchymal stem cells, which were induced by different combined neuro-revulsant, exhibited neuronal morphological characteristics: cellular shrinkage and neurite growth. The result of immunocytochemistry showed that the positive rates of nestin and NF of different groups were dissimilar, and the group E had the tiptop positive rate in all groups: nestin (5.653±1.228) %, NF (74.724±3.651) %. The results of GFAP immunocytochemistry stain were negative in all groups. The results of data analysis by SPSS10.0 showed that the difference between the different induced groups had prominence signification.[Conclusion] A relatively ideal combined neuro-revulsant and method of inducement of human bone mesenchymal stem cells into neurons were found. Part.3 The Research on the treatment of transplantation of induced differentiated Human Bone Marrow Mesenchymal Stem Cells on spinal cord injury[Objective]: To research whether the transplantation of oriented-induced Human Bone Marrow Mesenchymal Stem Cells is helpful to the functional recovery of animals with spinal cord injury and the influence of different transplantation time on the recovery.[Method]: 36 SD rats were made into models of spinal cord injury, and then divided into three groups of A,Band C at random. Group A were transplanted immediately after the injury of spinal cord, Group B one week after the injury while Group C were the control group. Use BBB scale to evaluate each group 1-6 weeks after the injury, and the data were analyzed by SPSS12.0. Immunofluorescence technique was used to detect the survival of the cells marked by Brdu and its integration with the spinal cord.[Results]: The mark of Group B was much higher in the BBB scale. Compared with the other two groups , the differences had statistical significance. Under the Immunofluorescence, we observed that in Group B most of the transplanted cells were survival which formed reticular structure with the regenerating host neuropil, bridging at the two sides of the injured area.[Conclusion]: Delayed transplantation of the oriented-induced hMSCs can promote the functional recovery of the animals with spinal cord injury.