The Research Of The Treatment Of Co-transplanted Umbilical Cord Mesenchymal Stem Cells And Cytokines In Traumatic Brain Injury

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The cultivation and induced differentiation of umbilical cord-derived mesenchymal stem cellBackgroudUmbilical cord-derived mesenchymal stem cells(UC-MSCs)are obterned from the umbilical cord after rat gestation.UC-MSCs have the ability to change into different type of cells and to proliferate continuously.The resource of UC-MSCs is rich in umbilical cord,and UC-MSCs are safe to transplantation.UC-MSCs could be induced to the cells with neurological function what were transplanted into the stroke models,as the result,the motor behavior of models were notable improvement.But the molecule mochanism of the treatment with UC-MSCs is unkown.Object1.We want to obtain and indentify the UC-MSCs in vitro.2.We take the plan to induce UC-MSCs differentiation to the cells with neurological function.3.We model the inflammatory cells in vitro to explore the roles of co-transplaned cytokines to UC-MSCs.Method1.The isolation and culture of rat UC-MSCs.2.The indentification of UC-MSCs with flow cytometry3.The differentiation of UC-MSCs to the cells with neurological function.4.The immunofluorescent staining of the induced differentiation cells.5.The cytoactive of UC-MSCs what was stimulated with IL-1?.Result1.UC-MSCs were isolated successfully and which could be induced to express neurogliocyte's marker GFAP and Tuj 1.2.The co-transplanted cytokines could protect UC-MSCs from inflammation by IL-1? through promoted cytoactive.3.IL-1? could inhabit the expression of Pi3k and AKT,but up-regulate the expression of caspase 3 which could lead UC-MSCs to apoptosis.However,the co-transplanted cytokines could reduce the effect of IL-1?to promote UC-MSCs survival.Conclution1.The isolated UC-MSCs could be induced to differentiate to neurological cells which express neurogliocyte's marker GFAP and Tuj1.2.The co-transplanted cytokines could promote UC-MSCs survival through up-regulated the cytoactive and inhibited apoptosis.Section ? The building and UC-MSCs transplatation of Traumatic brain injurie rat modelsBackgroundTraumatic brain injurie(TBI)is the disease of neurologicl dysfunction in brain injury area which is caused by traumatism.The acute injury and the second injury are the important factor to lead the wounded person disability.The inflammatory,angiorrhexis and thrombogenesis after TBI could lead the stroke and hypoxic ischemic brain damage which caused nerve apoptosis.The neurological dysfunction is hard to be recovery because the inhibitory regeneration of neuron.UC-MSC transplantation is a novel technical to treat nerve injury,so we plan to explor the treatment of UC-MSC transplantation in TBI model.ObjectIn this experiment,we plan to build the TBI rat model and transplant UC-MSCs to dectect and grade the change of TBI model behavior.Method1.We builded the TBI model with free fall method and divided into 4 groups:co-transplantation group,stem cell group,cytokines group and PBS group.2.The detection and Neurological impairment score(NSS)of TBI models after TBI operation 1,2,3,4 weeks.3.We obtained and adjusted the concentration of UC-MSCs to 1x106/uL.After that,we injected the UC-MSCs at the damaged area,the total volume is 5uL and the transplanted UC-MSC number is 5x106.At last,the needle which was used to transplant UC-MSC should be maintained 10 min after the transplantation finished.4.The detection and Neurological impairment score of TBI models after UC-MSCs transplantation 1,2,3,4 weeks.ResultThe survival rate of TBI model is 98%,and the results of behavior and NSS were homogeneous highly.In the results,the NSS of co-transplantation group was lower than stem cell group(P<0.05),stem cell group was lower than sytokines group and PBS group(P<0.01).ConclutionWe builded the TBI models with rat successfully,and the survival rate of TBI models was homogeneous highly.The results of behavior and NSS revealed that the treatment effect of co-transplantation group was better than stem cell group,but stem cell group was better than cytokines group and PBS group.The transplantation with cytokines and UC-MSCs would be an novel and effective treatment in TBI.Section ? The research of the mechanism of transplanted UC-MSC and cytokines in TBI modelsBackgroundTBI is a serious disease to affect the lives of patients,but there are no effective treatment to treat the patients.The transplantation of UC-MSC disease is a novel method to treat the nerve injury,but the mechanism of the treatment is still unkown.So we take this project to explore the molecular mechanism and the therapeutic mechanism of UC-MSC and cytokines.Method1.The detection about the survival of the transplanted UC-MSC with immuuofluorescent staining.2.The detection about the expression of the cytokines by the transplanted UC-MSC with ELISA3.The detection about the expression of the molecular about proliferation and apoptosis with western blot.Result1.The survival and the differentiation of UC-MSCs in TBI model We took the frozen section and immunoflurorscent staining after UC-MSCs were transplanted 4 weeks.The staining result revealed that UC-MSCs could differentiated to the neuroglia like cells which expression GFAP.However,we did not find the neuroglia like cells in the model of the groups cytokines and PBS.2.The expression of neurotrophic factors and inflammatory factors after UC-MSC transplantationThe expression of GDNF and BDNF were enhanced in stem cell group and co-transplantation group compaired with cytokines group and PBS group(P<0.05).But the level of GDNF and BDNF were significant higher after UC-MSCs were transplanted 4 weeks(P<0.001),and the co-transplantation group was higher than stem cell group(P<0.05).In contrast,the level of inflammatory factors were lower in stem cell group and co-transplantation group than other groups after UC-MSCs were transplanted 2 weeks(P<0.05),and the inflammatory factors were significant reduce after UC-MSCs were transplanted 4 weeks(P<0.001).3.The expression of the protein about proliferation and apoptosis In our results,the expression of p-Pi3k and p-AKT were enhanced in stem cell group and co-transplantation group(P<0.05),and the level of co-transplantation was higher(P<0.05).In contrast,the expression of caspase what involved in cell apoptosis was reduced in stem cell group and co-transplantation group(P<0.01),and the level of co-transplantation group was lower than stem cell group(P<0.05).Conclution1.The transplanted UC-MSCs could differentiate to neuroglia like cells in the damaged area,and the surviving and differentiated UC-MSCs could release neurotrophic factors and enhance the expression of Pi3k and AKT to promote the damaged neuron survival;meanwhile,they could inhibit the expression of apoptotic protein caspase3 to reduce the apoptosis of neuron in damaged area.2.The cytokines were transplanted into TBI model with UC-MSCs could promote UC-MSCs survival and differentiation to mediate the transplantation therapy.