Livin基因修饰的骨髓间充质干细胞移植治疗对急性心肌梗死大鼠心功能的影响

Effect of livin-modified BM-MSCs transplantation on cardiac function following acute myocardial infarction in a rat model

目的:探讨livin修饰的骨髓间充质干细胞(MSCs)移植改善急性心肌梗死大鼠心功能的作用及livin和促凋亡蛋白caspase-3、caspase-7、caspase-9在感染后的表达情况。方法:通过全骨髓培养法分离培养骨髓间充质干细胞,感染携带livin基因的重组腺病毒后用流式检测其对MSCs凋亡的影响。通过Western blot法分别检测livin、caspase-3、caspase-7和caspase-9的蛋白表达情况。采用冠状动脉左前降支结扎法建立心肌梗死模型,然后实验分4组进行:无胎牛血清的DMEM组;单纯干细胞治疗组(MSCs组);空载腺病毒转染干细胞组(r Ad-control/MSCs)组;livin基因转染干细胞组(r Ad-livin/MSCs组)。1个月后采用生理仪记录各组大鼠左室收缩压(LVSP)、左室舒张末压(LVEDP)、左室内压最大上升速率(+dp/dt_(max))和左室内压最大下降速率(-dp/dt_(max))来评价大鼠心功能。结果:r Ad-livin/MSCs组凋亡率较MSCs组和r Ad-control/MSCs组显著降低(P<0.05);r Ad-livin/MSCs组抗凋亡蛋白livin明显上升(P<0.05),促凋亡蛋白caspase-3、caspase-7及caspase-9表达显著下降(P<0.05);细胞移植后1个月,r Ad-livin/MSCs组心功能比MSCs组改善更明显(P<0.05)。结论:r Ad-livin感染MSCs后可促进抗凋亡蛋白livin的表达,同时降低促凋亡蛋白caspase-3、caspase-7和caspase-9的表达及细胞凋亡率。r Ad-livin/MSCs移植能进一步改善心肌梗死大鼠心功能。

AIM：To study the effect of livin gene-modified bone marrow mesenchymal stem cells（ BM-MSCs）transplantation on the cardiac function following acute myocardial infarction in a rat model and the expression of livin,caspase-3,caspase-7 and caspase-9 in the livin gene-modified BM-MSCs.METHODS：The MSCs were obtained by the whole bone marrow culture method,and the apoptosis of the MSCs after infection with adenovirus vector carrying enhanced green fluorescent protein（ EGFP） gene and livin recombinant vector（ r Ad-livin） were detected by flow cytometry.The expression of livin,caspase-3,caspase-7 and caspase-9 was detected by Western blot.After permanent left anterior descending artery occlusion,the rats were randomized to receive intramyocardial injection of DMEM without cells（ vehicle group）,or containing MSCs（ MSCs group）,MSCs（ EGFP）（ r Ad-control/MSCs group） or MSCs（ livin）（ r Ad-livin/MSCs group）.Left ventricular systolic pressure（ LVSP）,left ventricular end-diastolic pressure（ LVEDP）,the maximum increased rate of left ventricular pressure（-dp/dt_（max）） and the maximum decline rate of left ventricular pressure（ ＋ dp/dt_（max）） were recorded for evaluating the cardiac functions.RESULTS：The apoptosis of r Ad-livin/MSCs was significantly decreased as compared with MSCs and r Ad-control/MSCs（ P〈 0.05）.Meanwhile,the expression of caspase-3,caspase-7and caspase-9 was significantly downregulated as compared with the other 2 groups（ P〈 0.05）.The cardiac function in r Ad-livin/MSCs group was significantly improved as compared with DMEM group,and those in the other 2 groups got the similar results,but the function in r Ad-livin/MSCs group was better improved.Meanwhile,the number of surviving cells in r Ad-livin/MSCs group was significantly improved as compared with the other 2 groups.CONCLUSION：The apoptosis of MSCs is decreased after r Ad-livin transfection,and the expression of caspase-3,caspase-7 and caspase-9 is also significantly downregulated while the expression of livin is significantly upregulated.Transplantation of livin-modified BM-MSCs by lentiviral vector results in better prognosis for treating myocardial infarction by enhancing cell survival.