| bjective:Study on human umbilical cord blood mesenchymal stem cell trans-plantation in the repair of bone marrow hematopoietic damage,promote the re-covery of hematopoietic function of bone marrow.Methods:1.Culture,identification and CM-Di L markers of human umbilical cord blood mesenchymal stem cells:Resuscitation of human umbilical cord blood mesenchymal stem cells,and then the cells were cultured in 20%FBS89%DMEM/F-12 and 1%penicillin streptomycin medium.24h remove all themedium.Then medium was changed about 2 to 3 days.Observing the chang-es in cell morphology and growth under optical microscope.The cells are pas-saged when the cells are filled with the 70%-80%of the culture bottle.Identifi-cation and CM-Di L markers of the sixth generation cells and the labeling effect and cell morphology were observed under the fluorescence microscope.2.BALB/c mice were fed and established animal models of bone marrow hematopoietic damage:BALB/c mice were irradiated with X rays with a total dose of 6Gy,and an animal model of bone marrow injury was established.3.Experimental study of human umbilical cord blood mesenchymal stem cells transplantation to repair hematopoietic injury in bone marrow:The experi-mental group,control group and normal group were set up.The experimental group and control group were the mice with bone marrow injury,and the nor-mal group was normal mice.In the experimental group,CM-Di L labeled hu-man umbilical cord blood mesenchymal stem cells were injected into the tail vein of each mouse with 5x10~6(0.2ml).The control group and the normal group entered the normal saline 0.2ml through the tail vein.The general situa-tion of mice’s activities,feeding and body weight changes were observed,and the hematologic changes,bone marrow and homing of human umbilical cord blood mesenchymal stem cells were collected at 1,5,7,14 and 21 days after cell infusion.Results:1.Culture,identification and CM-Di L markers of human umbilical cord blood mesenchymal stem cells:Under the inverted microscope,human umbilical cord blood mesenchymal stem cells adhered and grew in a spindle shape,arranged orderly,the cell morphology was uniform,the refraction was good,the boundary was clear,and the cell morphology was stable until the 11generation.The sixth generation of h UBMSC,were used to identify cell surface markers,the results showed cells with high expression of CD105,CD29,CD90,the positive rates were 100%,99.9%and 66.9%,almost no expression of CD34,CD45.CM-DiL labeled human umbilical cord blood mesenchymal stem cells.Under fluorescence microscope,the cell membrane and cytoplasm showed red fluorescence.After first,seventh,fourteenth days and 21 days,the positive rate of CM-Di L marker in h UBMSCs cells was 100%,66.9%,31.7%,4.1%,respec-tively.2.BALB/c mice were fed and established animal models of bone marrow hematopoietic damage:The total dose of 6Gy X ray irradiated mice.On the first,fifth,seventh and 14 day after irradiation,the three blood system decreased,the myelodysplasia decreased,and the seventh day descended most obviously.On the fourteenth day,the peripheral blood and bone marrow hematopoiesis began to recover.On the twenty-first day,the peripheral blood and bone marrow hem-atopoiesis returned to normal.3.Experimental study of human umbilical cord blood mesenchymal stem cells transplantation to repair hematopoietic injury in bone marrow:(1)Blood rou-tine:On the first day after transplantation,there was no significant difference in the count of white blood cells,platelets,red blood cells and hemoglobin in the three groups of mice.Fifth days after transplantation,white blood cells,red blood cell count and hemoglobin concentration in experimental group com-pared with the control group and the experimental group compared with the normal group,the difference was not statistically significant(P>0.05),but the control group compared with normal group,the difference was obviously(P<0.05),platelet count of three group had a significant difference(P<0.05);Seventh days after transplantation,white blood cell,platelet count and hemo-globin concentration in the experimental group compared with control group,the difference was obviously(P<0.05),red blood cell count of experimental group compared with control group,the difference was not obviously(P>0.05),white blood cells,platelets,red blood cell count of experimental group and control group compared with the normal group,the difference was obvi-ously(P<0.05),but no significant difference was found in hemoglobin con-centration when experimental group compared with the normal control group(P>0.05).Fourteenth days after transplantation,white blood cells,red blood cell,platelet and hemoglobin concentration in experimental group compared with the control group,the difference was obviously significant(P<0.05),white blood cells,platelets,red blood cell count of experimental group and control group compared with the normal group,the difference was obviously significant(P<0.05),compared with the normal group but the hemoglobin concentration in the experimental group,the difference was not obviously(P>0.05);Twenty-first days after transplantation,compared to white blood cells,red blood cell count and hemoglobin concentration in the experimental group and the control group,the difference was not obviously(P>0.05),but the white blood cells,red blood cell count,the control group compared with the normal group,the difference was obviously significant(P>0.05),platelet count in the experimental group compared with control group and the control group compared with the normal group,the differences were statistically sig-nificant(P<0.05).(2)Bone marrow smear:Bone marrow smears showed first,fifth,seventh,14 days after transplantation,the hematopoietic function of the mice in the experimental group and the control group was inhibited.In mice,the myelodysplastic decreased and the nucleated cells decreased significantly.and the seventh day descended most obviously.Fourteenth days after trans-plantation,bone marrow hyperplasia was recovered,the experimental group better than the control group;In the twenty-first day after transplantation,the hematopoietic function of the mice in the experimental group and the control group was recovered,and there was no difference compared with the normal group.(3)Pathological section of bone marrow:The bone marrow biopsy showed that the hematopoietic function of the mice in the experimental group and the control group was inhibited on the fifth day after the transplantation.The number of nucleated cells decreased and the proportion of non hematopoi-etic cells increased.On the seventh day after transplantation,the number of nu-cleated cells was significantly reduced,adipose tissue increased,and a large number of vacuoles were seen.However,in the control group,the number of nucleated cells was more obvious than that of the experimental group,and the adipose tissue was more.After fourteenth days after transplantation,the hema-topoietic function of the experimental group and the control group were recov-ered and the nuclear cells were distributed in clusters,but the proliferation of bone marrow in the experimental group was better than that of the control group.(4)The tracer of human umbilical cord blood mesenchymal stem cells:On day first,fifth,seventh,14,and 21 days after transplantation,human umbilical cord blood mesenchymal stem cells labeled with CM-Di L were found in mice bone marrow smears.Conclusion:(1)Human umbilical cord blood mesenchymal stem cells were spindle shaped and adhered to the wall.The cells expressed CD105,CD29,and CD90,and almost did not express CD34 and CD45;CM-Di L could be used as a tracer for human umbilical cord blood mesenchymal stem cells.(2)The total dose of 6Gy X ray irradiated BALB/c mice,and the mouse model of bone marrow hematopoietic injury could be established.(3)The transplantation of human umbilical cord blood mesenchymal stem cells can repair the injury of bone marrow hematopoietic and promote the recovery of hematopoietic func-tion of bone marrow. |
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