摘要
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 evaluated the safety and effectiveness of autologous mesenchymal stem cell (MSCs) transplantation into endocardium in treatment of acute myocardial infarction. Methods Acute myocardial infarction was induced by reperfusion after ligating the left anterior descending coronary artery (LAD) for 1 hour in swines. Mononuclear cells (MNCs) were freshly isolated from the bone marrow blood by 1.077 g/ml density gradient centrifugation. Two to three weeks after cultured in the complete cell culturing medium, the purified MSCs or medium without serum were autologously transplanted into the target ischemic endocardium by injection catheter. The myocardiom around infarcted region was examined under fluorescence microscope with immunohistochemistry for desmin and myocardium specific troponin I (cTnI) 3 months after transplantation. LV function and blood biochemistry were examined at 6 weeks and 3 months after transplantation. Results The swine MSCs showed a fibroblast like morphology with vortex like distribution at their peak propagation and (3.81±0.09)×10 7 cells were harvested after culturing three generations over 2～3 weeks. DAPI labeled MSCs in 1ml of medium and medium only were injected into many sites of the ischemic LAD territory with injection catheter. Left ventricular function was improved significantly and the LVEFs were 46.6± 3.5% and 56.7±0.8% vs 34.9±0.9% ( P <0.05) at 6 weeks and 3 months vs post AMI respectively. The functions of liver and kidney, blood glucose and arterial blood gas analysis were normal. The blue fluorescent nuclear cells expressed markers of myocardium phenotype which showed positive stain of desmin and cTnI. The function and microstructure of important organs were normal. Conclusions The in vitro propagated MSCs were safe for regeneration therapy in vivo . The experimental endocardium injection catheter can be used in experimental study due to its convenient manipulation. The transplanted MSCs are located definitely and express markers of myocardium phenotype, resulting in the significant improvement of heart function.
出处
《中华老年多器官疾病杂志》
2004年第4期276-280,共5页
Chinese Journal of Multiple Organ Diseases in the Elderly
基金
国家 973计划 (2 0 0 1CB5 0 990 6)
国家 863计划重大专项 (2 0 0 2AA2 0 5 0 5 1
2 0 0 3AA2 0 5 160 )
国家自然科学基金 (0 3 0 5 0 40 2 )
北京市科委重大项目 (H0 2 0 2 2 0 0 10 190 )。
