曲美他嗪联合骨髓间质干细胞移植治疗急性心肌梗死的实验研究

目的探讨曲美他嗪(TMZ)能否改善骨髓间质干细胞(MSCs)在体外缺氧模型及急性心肌梗死(AMI)大鼠心脏的存活。方法加入或未加入TMZ的MSCs在无血清培养基培养并缺氧暴露12h,采用透射电子显微镜和流式细胞仪检查第3代MSCs的活力和凋亡。30只Wistar大鼠随机分为AMI对照组、MSCs组及(MSCs+TMZ)组,结扎左冠状动脉前降支制备AMI模型。将MSCs注入梗死心肌边缘[(MSCs组和(MSCs+TMZ)组)]。(MSCs+TMZ)组的大鼠在AMI前3d开始至AMI后28d加喂TMZ。移植28d后,超声心动图评估心脏结构和功能。免疫荧光染色检测移植细胞在体内的存活和分化。TUNEL法检测细胞凋亡。收集TMZ治疗开始前和AMI后24、48h的血液样本,测量C反应蛋白(CRP)与肿瘤坏死因子-α(TNF-α)的变化。结果缺氧培养下,TMZ处理过的MSCs细胞凋亡降低了一半。在体内与AMI对照组相比,MSCs组和(MSCs+TMZ)组的心肌梗死面积显著缩小,心功能明显改善。与单纯MSCs移植相比,TMZ与MSCs移植的组合治疗表现出了更低的干细胞凋亡、更高的干细胞存活、更小的心肌梗死面积和进一步改善的心功能。各组之间CRP、TNF-α的基线水平并无显著差异,然而24h时(MSCs+TMZ)组的所有参数均低于MSCs组。结论 MSCs移植添加TMZ治疗AMI增加MSCs存活和心脏功能的恢复上优于单纯MSCs移植,抑制炎症因子表达可能是其机制之一。

The experimental research on the treatment of acute myocardial infarction by trimetazidine and bone marrow mesenchymal stem cells transplantation

Objective To investigate the effects of trimetazidine(TMZ) on the survival of mesenchymal stem cells(MSCs) in an ex-vitro model of hypoxia and subsequent activities of transplanted MSCs in rat hearts with acute myocardial infarction(AMI).Methods MSCs were cultured in serum-free medium and exposed to hypoxia for 12h with or without TMZ.The viability and apoptosis of MSCs at passage 3 were examined by transmission electron microscope.Thirty wistar rats were divided randomly into 3 groups,including AMI control group,MSCs transplantation group,and TMZ+MSCs group.MSCs were injected into peri-infarct myocardium(MSCs and TMZ+MSCs groups) thirty minutes after coronary artery ligation.The rats in TMZ+MSCs group were additionally fed TMZ from 3 days before AMI to 28 days after AMI.Cardiac structure and function were assessed by echocardiography 28 days after transplantation.The survival and differentiation of transplanted cells were detected by immunofluorescent staining.The cellular apoptosis in the peri-infarct region was detected with TUNEL assay.Blood samples were collected before the start of TMZ therapy and 24h,48h after AMI,and inflammatory cytokines(CRP,TNF-α) were measured.Results In hypoxic culture,the TMZ-treated MSCs displayed a two-fold decrease in apoptosis under serum-free medium and hypoxia environment.In vivo,cardiac infarct size was significantly smaller,cardiac function significantly improved in the MSCs and TMZ+MSCs groups than those in the control group.Combined treatment with TMZ and MSCs implantation demonstrated a further decrease in MSCs apoptosis,a further increase in MSCs viability,a further decrease in infarct size,and a further improvement in cardiac function,compared with those of MSCs group.The baseline levels of inflammatory cytokines(CRP,TNF-α) were not significantly different among the groups.However,all parameters at 24h were lower in TMZ+MSCs group than those in MSCs group.Conclusion Implantation of MSCs combined with TMZ treatment is superior to MSCs monotherapy for MSCs viability and cardiac function recovery.The inhibition of inflammatory cytokines expression may be the potential mechanism.