siRNA靶向干扰IL28RA基因对缺氧复氧心肌细胞损伤的保护作用

Protective effects of siRNA targeted IL28RA gene on hypoxia reoxygenation cardiomyocy-tes injury

目的:探讨小干扰RNA(small interference RNA,siRNA)靶向干扰白细胞介素28受体α(interleukin 28 receptor alpha,IL28RA)基因对心肌细胞缺氧复氧损伤的保护作用。方法 :设计合成干扰IL28RA基因表达的3对siRNA(siRNA-6158、siRNA-6160、siRNA-6162),采用脂质体转染原代乳大鼠心肌细胞,分为正常对照组、单纯缺氧复氧组、缺氧复氧+阴性对照转染组、缺氧复氧+siRNA-6158转染组、缺氧复氧+siRNA-6160转染组、缺氧复氧+siRNA-6162转染组,探索siRNA转染心肌细胞的合适浓度,检测心肌细胞存活率、培养液中LDH的水平及心肌细胞IL28RA蛋白表达,观察siRNA干扰IL28RA基因对心肌细胞缺氧复氧过程中的保护作用。结果:采用脂质体转染法,siRNA浓度在80 nmol/L对心肌细胞具有较高的转染效率。与单纯缺氧复氧组和缺氧复氧+阴性对照转染组比较,缺氧复氧+siRNA-6158转染组和缺氧复氧+siRNA-6160转染组的LDH活性明显降低(P<0.05),心肌细胞存活率明显升高(P<0.05),IL28RA蛋白表达明显减少(P<0.05)。结论 :干扰IL28RA基因的表达有望成为保护缺氧复氧心肌损伤的重要手段。

Objective：To explore the protective effects of small interference RNA（siRNA） targeted interleukin 28 receptor alpha（IL28RA） gene on the hypoxia reoxygenation cardiomyocytes injury. Methods：A complex was designed and synthesized,which consisted of three pairs of siRNA（siRNA-6158,siRNA -6160,siRNA-6162） interfering IL28 RA gene expression. Liposome transfection method was used to transfect the siRNA into primary neonatal rat cardiomyocytes. The cells were divided into six groups including the normal control group,hypoxia and reoxygenation group（H / R group）,H / R group ＋negative control transfection group,H / R ＋siRNA-6158 transfection group,H / R ＋siRNA-6160 transfection group,and H / R ＋siRNA-6162 transfection group. The cell survival rate and lactate dehydrogenase（LDH）level in the supernatant were detected. The appropriate transfection concentration was explored. The IL28 RA protein expression was observed in the H / R process by using Western blot method. Results：The siRNA of 80 nmol / L is the appropriate transfection concentration. Compared with the H / R group and H / R group ＋negative control transfection group,the LDH activity in the H / R ＋siRNA-6158 group and H / R ＋siRNA-6160 group was significantly decreased（P 〈0.05）, the survival rate in the both groups was significantly increased（P〈 0.05）, and the IL28 RA protein in the both groups was significantly reduced（P〈 0.05）.Conclusion：Inhibiting IL28 RA gene expression was expected to be an important method of protecting anoxic myocardial disease.