The Study Of Recovery Of Chronic Spinal Injury Procedured By Endothelial Progenitor Cells

Objective1.To select a ideal device, by which the model of chronic compression of spinal cord was made, via designing3types of compressing-device to contribute to further studies on path physiologic mechanism of chronic compression spinal cord and its recoverment.2.Bone marrow mononuclear cells (BMMNCs) were from rats bone marrow isolated, and the BMMNCs were to endothelial progenitor by inducing-culture medium differentiated. Then they were injected into the model of chronic compression of spinal cord, to evaluate the impact of EPCs on recoverment after decompression of the model of the chronic compression of spinal cord.Method1.Three types of fixing compression-device were designed via the software AutoCAD201064x and Solidworks201064x. Twenty nine Wistar rats were enrolled in this part of experiment, which were randomly distributed into sham operation group (n=5),SP group(n=8),EB compressing group(n=8) and D-H group(n=8). Rats, whose lamina were resected only, are from the sham operation group, whereas the experimental group were implanted respectively the3types of device. withdrawal force were respectively measured after1week of the implantation. after6weeks of compression the status of apoptosis were observed via TUNEL stain of cells.2.Bone marrow mononuclear cells (BMMNCs) were obtained from the medulla ossium of4-6w Wistar rats by flushing rat’s femurs and tibiae and were isolated by density gradient centrifugation and then cultured in inducing culture medium,EGM-2MV. The growing statuses were morphologically observed under inverted microscope every day. The cultured cells at the7th day of the cultuvation were identified by Flow Cytometry, which performed to analyze the expression of CD133, KDR and CD34of the cultured cells,meanwhile by the experirment of uptaking DiL-acLDL and combining FITC-UEA-1under fluorescence microscope.3. Ten models of chronic compression of spinal cord, whose spinal cord were compressed for6weeks,were distributed into model control group (n=5), simple decompression group,SD group(n=5)and decompresion combined with the injection of EPCs group,DCE group (n=5). The devices were uninstalled after the BBB evaluation. The spinal marrow of DCE group were injected with EPCs with105cells(10u), whereas the SD group were given with10u PBS. After4weeks BBB evaluation is given, and then the specimens of compressional segments were havested, to which immunofluorescence stain of ChAT were procedured. The numbers of positive cells were analyzed by statistics.Result1.SP and D-H group showed no dislocation from the fixed location, according to the X-ray, after1week of implantation. The caudal wings of EB showed its tendency of dislocation to the posterior. Measured value of force in the process of5mm pull out shows that the maximal withdrawal force of D-H,SP and EB are respectively equal to245.4±12.83,7.67±0.70and25.77±21.55,which were statistically analyzed via ANOVA. Difference among them was statistically significant(P<0.05).in the D-H group it emerge a rapid reduction of withdraw force in the process of pull out. the specimens of spinal segments, which are after6weeks of compression, were procedured TUNEL stain, which shows that the numbers of TUNEL positive cells of the device implanted3groups were statistically varied(P<0.05).2.The successful induction to endothelial progenitor cells was vertified by observation of morphological characteristic of cells, which were cultured by EGM-2MV,also by Flow Cytometry, that measured the expression of CD133,CD34and KDR, which were respectively equivalent to57.5%、11.4%and19.2%,and by identification via the experiment of uptaking DiL-acLDL and combining FITC-UEA-1by fluorescence microscope.3.BBB evaluation of SD group and DCE group,which were preformed decompression4weeks later, were both significantly different from theirself before decompression (P<0.05).Between the two experimental group(SD and DCE), the D-values of BBB,which are between decompression4weeks later and before, were significantly of difference(P<0.05).The number of ChAT positive cells of the control group is4.4±1.67, SD7.0±1.22and DCE17.4±3.58,among which the significant variation existed(P<0.05).Conclusion1. The type of D-H compressive device,which was manufactured and designed via AutoCAD as well as Solidworks, was a ideal apparatus, which was controllable strong and accorded with the requirements of the model of chronic compression of spinal cord in rats.2.The recovery degree of motor neuronal function in4weeks after decompression in DCE group was superior to SD.