Effects Of Low Dose Irradiation On Distribution Of Bone Marrow Mesenchymal Stem Cells In Vivo

Part one:Fetal bone marrow-derived mesenchymal stem cells——basic and biological charactersObjective:To explore the condition of the isolation and culture of fetal bone marrow-derived mesenchymal stem cells(FMSCs)in vitro and to observe the biological characteristics of FMSCs.Methods:Bone marrow mononuclear cells(BMMNCs)were harvested from bone marrow by flushing femurs and tibias of four-month old human fetales.The resulting cells were purified via density gradient centrifugation and cultured in vitro.The proliferation and growth characteristics of FMSCs were analyzed during primary and passage cultures.FMSCs cell growth curve and divisional index curve was detected by MTT.The well-grown FMSCs of the third generation were cultured in the DMEM-LG medium containing new-born calf serum,dexamethasone,β-glycerophosphate, antiscorbic acid to induce the ostroblasts;in the DMEM-LG containing new-born calf serum,dexamethasone,insuline to induce adipocytes.Results:After seeded 48 hours,the adherent cell showed spindle shape, polygonal shape and fibroblast-cell-like shape.FMSCs was passaged after one week, the FMSCs were still in latent phase after being passaged 24-36 hours,2-3 days later, cells were in log phase,6 days later,and cells came into platform phase.Until 15 passages,the growth rate of FMSCs decreased.The phenotype analysis with FMC showed that FMSCs expressed highly CD29,CD44 and CD49e.They also expressed low level of CD106,CD54,CD11b,but FMSCs did not express CD34.FMSCs which cultuered in osteogenic inducer could change into black when stained by von Kossa. And FMSCs which cultuered in adipogenetic inducer could become jacinth when stained by oil red. Conclusion:FMSCs isolated,cultured,proliferated applying density gradient centrifugation,attachment method combining with monoclone cultivation have high autoploidy,and FMSCs can be induced into adipocytes and ostroblasts successfully in vitro.Part two:Role of low dose irradiation on homing and distribution of bone marrow mesenchymal stem cells in vivoObjective:To investigate the role of low dose irradiation on homing and distribution of bone marrow mesenchymal stem cells in vivo.Methods:According to Allen's punch method,the spinal cord injury mice model was successfully established in our lab.The models were randomly divided into two group,irradiated group and control group.Primary BMMSCs tagged with CFDA SE were injected at injured position of SCI mice in 30 minutes after mice were irradiated. The animals were sacrificed 24 and 96 hours after injection and the distribution characteristic of BMMSCs was studied through fluorescence microscope.Results:24 hours after BMMSCs injection,BMMSCs engraftment at the sites of injured position could be detected in both groups,but irradiated group as compared to control group increased significantly.96 hours after transplantation,BMMSCs engraftment still could be detected at the sites of injured position,cannot at other organs;we did not find the BMMSCs in the SCI of control group.Conclusion:In model of SCI,after transplantation,BMMSCs could immigrate to the position of spinal cord injury;after irradiated the model of SCI,BMMSCs could maintain stabilization of organism internal environment by enhancing reproductive activity.