大鼠骨髓间充质干细胞与脊髓脱-第三军医大学学报.doc

大鼠骨髓间充质干细胞与脱细胞脊髓支架体外共培养 王开见，任先军，蒋 涛，周长立，阴 洪（ ） [摘要] 目的 评价脱细胞脊髓支架在大鼠骨髓间充质干细胞(one marrow mesenchymal stem cells，BMSCs)BMSCs分离，培养及鉴定；BMSCs向神经元诱导；脱细胞脊髓支架的制备；将第3代BMSCs同脱细胞脊髓支架共培养，MTT法检测细胞增殖活性并描绘细胞增殖曲线，比较与普通培养的差异；取第3代BMSCs以不同浓度[（0.5、1、2、3、4）×106/mL]接种于支架（脊髓支架修整0.5 cm小段），检测细胞在支架上的粘附率及上架细胞数，建立接种浓度与细胞粘附率、上架细胞数的关系回归方程；扫描电镜观察脱细胞脊髓支架形态以及细胞与支架的粘附情况。结果 成功实现BMSCs的分离及培养，BMSCs流式鉴定CD29、CD90阳性表达，向神经元诱导胶质纤维酸性蛋白（GFAP）、巢蛋白BMSCs在支架上细胞增殖活力显著提高（P＜0.05）。细胞与支架的粘附率在接种度为2×106/mL最高，达到(79.6±2.7)%，单位体积上架细胞数达到最大值（1.364±0.047）BMSCs粘附、存活、增殖，该支架是良好的天然脊髓组织工程材料。当接种浓度为2×106/mL，粘附率及细胞上架数液基本达到最大，此浓度为最佳接种浓度。 [关键词] 骨髓间充质干细胞An experimental study of rat bone marrow mesenchymal stem cells co-cultured on the acellular scaffold of spinal cord in vitro Wang Kaijian, Ren Xianjun, Jiang Tao, Zhou Changli, Yin Hong (Department of orthopaedics ,Xinqiao Hospital ,Third Military Medical University , Chongqing,400037 ,China) [Abstract] Objective To evaluate the effect of the acellular scaffold of spinal cord on the rats bone marrow mesenchymal stem cells cultured in vitro and explore the best inoculation concentration. Methods:SD rat BMSCs were separated，cultivated and identified；BMSCs induced to neurons；The acellular scaffold of spinal cord was prepared ；The third generation of BMSCs were co-cultured with the acellular scaffold of spinal cord and MTT method was applied to assay the cell proliferation rates and to describe cell proliferation curve. Differences with common culture were explored.；The third generation of BMSCs in different concentrations (0.5, 1, 2, 3, 4 x106 /ml，respectively) were seeded on scaffolds (with spinal cord scaffolds trimmed as 0.5 cm small section) to detect cell adhesion rates and cell numbers on scaffolds，and to establish regression equation of inoculation concentration and cell adherence rate , cell numbers on scaffold；Scanning electron microscopy was also used to check shape of acellular scaffold of spinal cord and the adhesion between cells and scaffold. Results: we have success