An Experimental Study In Bone Marrow Mesenchymal Stem Cells In Treatment Of Acute Radiation-induced Intestine Injury In Rats

Part One: A study on the isolation, culture, purification of bone marrow-derived mesenchymal stem cells in sprague dawley rats and identification of its biological characteristics.Objective:To explore the method of isolation, culture, purification of SD rat bone marrow mesenchymal stem cells in vitro and identification of its biological characteristics. To establish an ideal isolation and culture system of SD rat bone marrow mesenchymal stem cells for further study and clinic application.Method: The SD rBMSCs were isolated from density gradient centrifugation combined with attachment culture method from bone marrow of the young SD rats.The shape was observed and the growth curve was detected.The cell membrane antigens were identified.The potency that BMSCs are induced and differentiated into osteoblasts and lipoblasts was observed.Result: It was found that rBMSCs were spindle shape and fibroblast-cell-like shape and the size of rBMSCs was homogeneous under imicroscope.The growth curve show the"S". CD44, CD29 merebrane antigen could be detected in 98% of the third generation of rBMSCs,but not CD34 membrane antigen.The rBMSCs are induced and differentiated into osteoblasts and lipoblasts.Conclusion: An effective and ideal isolation and culture system of SD rat bone marrow mesenchymal stem cells in vitro has been established.The rBMSCs were isolated and purified by density gradientcentrifugation combined with attachment culture method have high purity and proliferation activity.The biological properties are stabile. It applicates for further research on BMSCs. Part Two: An experimental study in bone marrow mesenchymal stem cells in treatment of acute radiation-induced intestine injury in ratsObjective: To observe the effect of the BMSCs on repairment of intestinal radiation damage and to explore the implanted mechanism of the BMSCs to provide theoretical and principles for its clinic use.Methods: Thirty female Sprague Dawley rats were randomly divided into 2 groups (each 15), control group and experimental group (BMSCs treatment group). They received medical linear accelerato a single dose of 12 Gay at whole abdominal irradiation. BMSCs treatment group were infused 4 hours before with a dose of 2×106 /ml BMSCs via the vena caudalis; Control group were infused a dose of 1ml normal saline. The animals were sacrificed three and fourteen days after irradiation. Plasma and terminal ileum specimens were collected. Plasma citrulline was detected by ELISA and pathologieal section was obversed in microscope. the expression of SDF-1 of terminal ileum was obversed by immunohistochemistry. To determine the implantation of BMSCs, the sex determing gene Sry was detected by PCR in recipients.Results: Three days after irradiation, BMSCs treatment group necrosis of epithelial cells and infiltration of inflammatory cell was less, intestinal mucosal villi and glands were more affluent compared with the control group. Fourteen days after irradiation, BMSCs treatment group intestinal structural was more integrity, and intestinal mucosal villi and glands were more affluent compared with the control group; Three and fourteen days after irradiation, in microscope , BMSCs treatment group, the measurements of ileum villus height were (245.54±12.75)μm and (321.36±18.21)μm. Control group, the measurements of that were (215.46±13.52)μm and (303.3±16.65)μm, P <0.05. BMSCs treatment group, the measurements of crypt depth were (148.82±10.12)μm and (204.25±13.58)μm. Control group, the measurements of that were (132.72±11.96)μm, (192.5±12.23)μm, P <0.05; Three and fourteen days after irradiation, the plasma citrulline of BMSCs treatment group was (11.32±1.42)μg/ml and(17.66±2.39)μg/ml. the plasma citrulline of control group was (9.15±1.56)μg/ml and (14.48±2.10)μg/ml, P <0.05; Three days after irradiation, the expression of SDF-1 of terminal ileum of BMSCs treatment group had a singnificant increase when compared with irradiation control group, P <0.05; The Sry gene was detected in recipient rats of BMSCs treatment group, which confirmed that BMSCs participate in the repair of rats intestinal tract damage.Conclusion: BMSCs can implant radiant injury tissue and involve the repairment of intestinal mucosal structure and function. Implantation of BMSCs may have close relation with SDF-1 / CXCR4.