| Spinal cord ischemia-reperfusion injury (SCII) is a common complication of surgery, particularly after aortic aneurysms. sacral tumor surgery, and these surgeries need to block the aorta, which leads to spinal cord injury, especially paraplegia caused by spinal cord ischemia-reperfusion after surgical treatment of thoracic aorta, the incidence rate is up to 3% to 18%. The mechanisms are complex, oxidative stress and apoptosis are the main mechanisms of SCII. Preventive methods of SCII are not ideal, and the therapy of SCII is mainly symptom and supportive treatment currently, which can't fundamentally prevent the occurrence of paralysis. Long hospital stay, expensive cost of patients bring heavy burden to family and society, and the therapeutic effect is not ideal. Mesenchymal stem cells (MSCs) are a kind of pluripotent stem cells derived from mesoderm, which have many characteristics including strong proliferative ability and multi-directional differentiate potential, weak immunogenicity, convenient source, easy to be isolated and cultured, and localization in the damaged area and differentiation into the cell types of the located tissue after entering into body. But stem cell viability could be influenced by the unfavorable factors such as ischemia, inflammatory factors, this leads to death of some stem cells and then impacts on treatment effects. Zinc is an essential trace element in human or animal body, has stabilizing effect on various kinds of biomembranes, can alleviate a variety of organ ischemia reperfusion injury and plays the roles of anti-oxidative stress, anti-apoptosis and improving local damaged micro-environment.This study is planned to pretreat rats with Zine and prepare SCII rat model, then transplant mesenchymal stem cells, to investigate the preventive and therapeutical effects of Zine and MSCs on spinal cord ischemia-reperfusion injury and the increased effect of Zine on repair of spinal cord ischemia-reperfusion injury by BMSCs. In this study, BMSCs were isolated. purified and expanded by the method of adherent culture, the cell cycle was measured by flow cytometry. The third passage BMSCs suffered from serum starvation, and then were cultured in the medium with presence of bFGF, EGF and RA for 6 d to be induced to differentiate into neurons, the methods of immunohistochemistry and immunofluorescence were used to detect the expression of neuronal surface markers, like NSE and NeuN. BMSCs were labeled by CM-Dil, mark rate was detected by flow cytometry. Adult female SD rats were divided into cell therapy group (0.1 mL.108·mL-1 BMSCs labeled by CM-Dil were administrated by retro-orbital intravenous injection), the Zine group (10 mg·kg-1 body weight ZnSO4 was given to rats by intraperitoneal injection every day before establishing model for 5 continuous days), Zine and cells group (rats were given both Zine pretreatment and cell transplantation), model group, sham operation group and normal control group. SCII model was prepared by occlusion of the abdominal aorta. Motor function assessment of rats were performed by Basso Beattie Bresnahan (BBB) Motor Function Assessment in the four groups of models, respectively at 1d,2d,3d, 4d,5d,6d after reperfusion. 7 days after ischemia reperfusion injury, rat spinal cord tissue were collected and detected by HE staining. Nissl body staining, Tunel method, immunofluorescence and immunohistochemistry to observe the changes of morphology and apoptosis of the spinal cord and the expression of the apoptosis-related genes. On the 4th day after ischemia-reperfusion, the distribution of BMSCs in rats was observed by Maestro in vivo imaging system; on the 7th day after ischemia-reperfusion, the distribution and differentiation of BMSCs in spinal cord were observed by using laser scanning confocal microscope.The results of this study showed that BMSCs cultured by adherent culture method had good cell viability and fast proliferation after subculture. After induction and differentiation, differentiated cells of BMSCs expressed the neuron specific markers of NeuN and NSE, which meant they had differentiated into neuron-like cells. Labeling with 4μg·mL-1 CM-Dil was not toxic to BMSCs. and the labeling rate is up to 93.9%. BBB motor function assessment of cell therapy group, Zine group. and Zine and cell group were significantly higher than that of model group (P<0.05). HE staining and Nissl body staining showed that the morphological changes of pathological tissue in all the intervention groups were significantly better than those in the model group. The results by Tunel method showed that the apoptosis rates of the intervention group were significantly lower than that in model group (P<0.05). Bcl-2/Bax ratio in intervention groups was significantly higher than that in model group (P<0.05), Caspase-3, Caspase-9, C-fos protein expressions in intervention groups were significantly lower than those in model group (P<0.05). Maestro in vivo imaging systems result showed that the BMSCs aggregated at the injury site on the third day of reperfusion. Homing of BMSCs, and differentiation of BMSCs into neurons and astrocytes in the injuried spinal cord were observed by laser scanning confocal microscopy on the 7th day of reperfusion.Conclusion:Zine can prevent spinal cord ischemia-reperfusion injury; BMSCs can promote the repair of spinal cord ischemia-reperfusion injury; Zine can enhance the therapeutic role of BMSCs on spinal cord ischemia-reperfusion injury; Homing of BMSCs. and differentiation of BMSCs into nerves and astrocytes in injuried spinal cord may be one of the mechanisms of repair of SCII by BMSCs transplantation; Anti-apoptosis may be one of the mechanisms that BMSCs Zine and BMSCs prevent and treat SCII, and Zine enhance the repair of SCII by BMSCs transplantation.
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