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目的通过制备大鼠心肌梗死模型,分别移植经纯化了的人源骨髓单克隆间充质干细胞(SCMSCs)、未纯化的间充质细胞和单个核细胞及外周血单个核细胞,旨在比较SCMSCs移植是否更有利于心脏功能改善。方法应用磁珠分选和极限稀释细胞培养方法获取SCMSCs,利用贴壁培养方法得到未纯化的骨髓间充质细胞,利用梯度密度离心法得到骨髓单个核细胞和外周血单个核细胞。流式细胞仪分析所得细胞特性后,将其移植到梗死心脏。术后1个月,应用血流动力学技术检测大鼠心脏功能,随后取材,以免疫荧光检测移植细胞的分化情况,用碱性磷酸法计算血管密度。结果流式分析结果显示,SCMSCs99%以上表达间充质干细胞标记蛋白,阴性表达造血细胞标记蛋白。功能检测显示,移植SCMSCs心脏收缩功能(LVdP/dtmax)较明显高于其他各组。血管计数显示,SCMSC组血管密度明显高于其他各组。免疫荧光结果显示,移植的SCMSCs向心肌细胞和血管内皮细胞转化效率明显高于其他骨髓细胞,外周血细胞并不发现分化。结论经纯化均一的SCMSCs移植较目前常用的未纯化、不均一的干细胞移植更利于梗死心脏收缩功能恢复。
OBJECTIVE: To prepare purified rat bone marrow mesenchymal stem cells (SCMSCs), unpurified mesenchymal cells, mononuclear cells and peripheral blood mononuclear cells by rat myocardial infarction model. Is transplantation more conducive to improving cardiac function? Methods SCMSCs were obtained by magnetic bead sorting and limiting dilution cell culture. Bone marrow mesenchymal cells were obtained by adherent culture method. Bone marrow mononuclear cells and peripheral blood mononuclear cells were obtained by gradient density centrifugation. After the cell characteristics were analyzed by flow cytometry, they were transplanted to the infarcted heart. One month after operation, the hemodynamics was used to detect the cardiac function in rats, then the material was collected and the differentiation of the transplanted cells was detected by immunofluorescence. The blood vessel density was calculated by alkaline phosphatase. Results The results of flow cytometry showed that more than 99% of SCMSCs expressed mesenchymal stem cell marker protein and negatively expressed hematopoietic cell marker protein. Functional tests showed that cardiac contractile function (LVdP / dtmax) in SCMSCs was significantly higher than that in other groups. Vascular count showed that the vascular density in SCMSC group was significantly higher than the other groups. Immunofluorescence results showed that the transplanted SCMSCs transformed into cardiomyocytes and vascular endothelial cells were significantly higher than other bone marrow cells, peripheral blood cells and found no differentiation. CONCLUSIONS: Purified and homogenized SCMSCs are more conducive to the resuscitation of systolic function in infarction than the unpurified and heterogeneous stem cell transplantation currently used.