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目的 探讨自体骨髓间充质干细胞 (MSCs)体外扩增后经心内膜移植治疗急性心肌梗死的安全性及治疗效果。方法 开胸结扎再通形成小型猪急性心肌梗死再灌注损伤的动物模型 ,抽取骨髓 ,1.0 77g ml密度梯度离心法分离骨髓单个核细胞 ,扩增培养后 2～ 3周经导管心内膜移植 ,移植前后心功能及血生化检查 ,3个月时取梗死心肌、肝、肾、脾等组织冰冻切片及HE染色 ,免疫荧光法鉴定结蛋白desmin和心肌特异性肌钙蛋白I (cTnI)的表达。结果 小型猪MSCs为形态均一的梭形细胞 ,每 4 0ml骨髓液经 2～ 3周传 3代平均获得 (3.81± 0 .0 9)× 10 7个细胞 ,移植后 6周及 3个月LVEF较心梗后明显改善 (P <0 .0 5 ) ,肝肾功能、血糖、动脉血气结果正常 ,3个月荧光显微镜下可见移植细胞 ,免疫组化提示desmin及cTnI表达阳性 ,HE染色提示肝、脾、肾等主要脏器结构正常。结论 体外扩增MSCs体内移植安全 ,各重要脏器功能及结构正常 ,心内膜注射移植操作方便 ,移植后的MSCs体内定位明确 ,心功能改善 ,并有心肌相关蛋白的表达。
Objective To investigate the safety and therapeutic effect of autologous bone marrow mesenchymal stem cells (MSCs) expanded after endocardial endocardial transplantation in the treatment of acute myocardial infarction. Methods Thoracotomy was performed to establish an animal model of reperfusion injury in mini-pigs with acute myocardial infarction. Bone marrow was sampled and bone marrow mononuclear cells (BMMNCs) were isolated by density gradient centrifugation (1.077g ml). After 2 to 3 weeks of culture, Cardiac function and blood biochemical examination before and after transplantation. Frozen section of infarcted myocardium, liver, kidney, spleen and other tissues were harvested at 3 months, HE staining and immunohistofluorescence were used to detect the expression of desmin and cardiac troponin I (cTnI) . Results The miniature porcine MSCs were spindle-shaped cells with uniform morphology. The average number of MSCs per 100 ml of bone marrow fluid was (3.81 ± 0.09) × 10 7 cells after 2 to 3 weeks of passage, and 3 weeks after transplantation (P <0.05), liver and kidney function, blood glucose and arterial blood gas were normal. Transplanted cells were observed under fluorescence microscope at 3 months. Immunohistochemistry showed that the expression of desmin and cTnI was positive. HE staining showed that liver , Spleen, kidney and other organs of normal structure. Conclusion MSCs transplantation in vitro is safe in vivo, and the function and structure of all the important organs are normal. The endocardial injection and transplantation are easy to operate. The MSCs possessed in vivo were well localized in vivo, the cardiac function improved and the expression of myocardial associated protein was observed.