转染Akt基因对心肌细胞I/R损伤的机制探讨

Experimental study of protective effect of cationic liposome mediated transfecting human Akt gene into cardiac myocytes against ischemia/reperfusion in vitro

目的探讨转染基因Akt对心肌细胞缺血/再灌注(I/R)损伤的机制。方法采用新生SD大鼠进行心肌细胞培养,建立模拟心肌I/R模型;提取人Akt质粒,用阳离子脂质体介导的基因转染法将人Akt质粒转染心肌细胞。实验分5组:空白对照组(Control组)、缺血/再灌注损伤对照组(I/R组)、转染Akt基因组(Akt组)、转染阳离子脂质体空载体组(Vehicle control组)和Akt抑制剂组(Akt blockade组);各组进行I/R后测定乳酸脱氢酶(LDH)活性、四唑盐比色法检测细胞活性、免疫印迹法(Western blot法)检测细胞内Akt、低氧诱导因子-1α(hypoxia-inducible factor-1α,HIF-1α)蛋白表达。结果 Akt组较I/R组、Ve-hicle control组和Akt blockade组其LDH含量明显减少(P<0.05),细胞活性高(P<0.05),Akt与HIF-1α蛋白表达增加(P<0.05)。结论转染Akt基因可减少缺血性心脏病心肌细胞I/R损伤,对缺血性心脏病具有保护作用。

Objective To explore the effect and mechanism of transfecting human Akt gene against ischemia/reperfusion（I/R） in cardiac myocytes in vitro.Methods Cardiac myocytes from new born Sprague-Dawley rats were isolated and cultured in vitro and were randomly divided into five groups：control group,ischemia/ reperfusion（I/R）group,Akt group,vehicle control group and Akt blockade group.The levels of lactate dehydrogenase（LDH）and the cell activity in the medium were tested.The expression levels of Akt,HIF-1α protein were measured by Western blot.Results In comparison with I/R group,Vehicle control group and Akt blockade group,the levels of LDH in Akt group were significantly decreased（P0.05）,together with higher cell activity（P0.05）with the highest protein expression levels of Akt and HIF-1αwere found by western blot（P0.05）.Conclusion Transfecting plasmid Akt gene reduces injury in cultured cardiomyocytes against I/R and shows protection effects on ischemic heart disease.