曲美他嗪预处理BMSCs移植对心肌缺血再灌注大鼠心功能改善的影响

Effect of trimetazidine preconditioning BMSCs transplantation on myocardial ischemia and reperfusion improved cardiac function in rats

目的探讨曲美他嗪(TMZ)预处理骨髓间充质干细胞(BMSCs)移植到缺血再灌注(I/R)微环境下细胞生存及对心功能改善的影响。方法将TMZ预处理或未预处理的BMSCs与心肌细胞H/R上清液共培养,采用CCK-8法检测细胞活性、RT-PCR检测Bcl-2、Bax的基因表达水平。32只SD大鼠随机分为假手术组(Sham组)、心肌I/R组、干细胞移植组(BMSCs组)及TMZ预处理干细胞移植组(BMSCs+TMZ组)。建立大鼠心肌缺血再灌注模型,当再灌注平稳后于损伤心肌边缘注入干细胞(细胞数5×105)。1 w后用荧光显微镜观察BMSCs存活数,4 w后应用超声心动图评估心功能,TTC法检测梗死面积,Western印迹分析移植周围区域Bcl-2及Bax的蛋白表达。结果 BMSCs与心肌细胞H/R上清液共培养后细胞活性显著下降,但TMZ预处理后使其下降趋势得到明显逆转。RT-PCR结果表明,与未预处理BMSCs相比,在TMZ预处理BMSCs中,Bcl-2基因表达水平上调,Bax基因表达水平下调。在体内,术后1 w,BMSCs+TMZ组每个高倍镜下双标记细胞数显著高于BMSCs组。术后4 w,BMSCs+TMZ组和BMSCs组均较I/R组心功能明显改善,心肌梗死面积缩小;两组间进一步比较,BMSCs+TMZ组表现出了更进一步的心功能改善与更小的心肌梗死面积。Western印迹结果表明,与未预处理BMSCs相比,在TMZ预处理BMSCs组中,Bcl-2表达上调,Bax蛋白下调。结论血运重建联合TMZ预处理BMSCs移植治疗急性心肌梗死增加BMSCs存活和心功能的恢复上优于单纯BMSCs移植,可能通过上调Bcl-2及下调Bax的表达而发挥抗凋亡作用。

Objective To investigate the survival of cells under the microenvironment after trimetazidine( TMZ) pretreatment bone marrow mesenchymal stem cells( BMSCs) transplanted to ischemia-reperfusion( analog revascularization) and the influence on cardiac function,and the mechanism. Methods The TMZ pretreatment or pretreatment hypoxia / reoxygenation( H / R) BMSCs supernatant of cultured myocardial hypoxia,CCK-8 was used to assay cell activity,RT-PCR was used to detect levels of Bcl-2,Bax gene expression. 32 SD rats were randomly divided into Sham,myocardial ischemia and reperfusion( I / R),stem cell transplantation( BMSCs) and TMZ pretreated stem cell transplantation( BMSCs+TMZ) groups. BMSCs survival was observed by fluorescence microscope,four weeks later,the heart function was assessed by echocardiography,infarct size was tested by TTC,the protein expressions of Bcl-2 and Bax around the transplanted area were analyzed by Western blot. Results The activities of cultured cells with BMSCs and cardiomyocytes after H / R supernatants were significantly decreased,but it declined after TMZ pretreatment significantly reversed. Compared with that of non-pretreated BMSCs group,after TMZ pretreatment,the Bcl-2 gene expression level was increased,while Bax gene expression levels was decreased. In vivo,after one week treatment,each high magnification double labeled cells was significantly higher in BMSCs+TMZ group than that of BMSCs group. After four weeks,the heart function was significantly improved in BMSCs+TMZ group and BMSCs group than that of I / R group,myocardial infarct size was reduced. Compared with non-pretreated BMSCs,Bcl-2 expression of BMSCs at TMZ pretreatment group was upregulated,whereas the expression of Bax was lower. Conclusions Revascularization combined trimetazidine pretreatment BMSCs transplantation on acute myocardial infarction could increase survival of BMSCs and improve recovery of cardiac function. It may be related with upregulating the expression of Bcl-2 and down regulating the expression of Bax.