精氨酰-tRNA合成酶在局灶性脑缺血再灌注大鼠中的表达及机制研究

摘要

背景目前，临床上针对哺乳动物氨基酰-tRNA合成酶的研究主要集中在精氨酰-tRNA合成酶、亮氨酰-tRNA合成酶、甲硫氨酰-tRNA合成酶、丙氨酰-tRNA合成酶的结构和功能方面，对哺乳动物尤其是大鼠脑缺血与氨基酰-tRNA合成酶关系的研究报道极少。目的探讨精氨酰-tRNA合成酶在局灶性脑缺血再灌注大鼠中的表达及机制。方法选取清洁级健康雄性 SD 大鼠60只，其中4只大鼠进行预实验，采用线栓法栓塞大脑中动脉建立局灶性脑缺血再灌注模型。然后将剩余的56只大鼠随机分为正常组8只、假手术组8只、脑缺血再灌注组40只。正常组不做任何外科处理；假手术组大鼠外科操作步骤与脑缺血再灌注组相同，只是线栓不进入颈内动脉、不造成脑缺血；脑缺血再灌注组大鼠建立局灶性脑缺血再灌注模型。采用反转录-聚合酶链反应（RT-PCR）测定正常组、假手术组、脑缺血再灌注组大鼠再灌注后不同时间点精氨酰-tRNA合成酶 mRNA 相对表达量，采用 Western Blotting 法测定正常组、假手术组、脑缺血再灌注组大鼠再灌注后不同时间点精氨酰-tRNA合成酶蛋白相对表达量。结果假手术组与脑缺血再灌注组大鼠再灌注2 h、脑缺血再灌注组再灌注48 h 精氨酰-tRNA合成酶 mRNA 相对表达量及蛋白相对表达量比较，差异无统计学意义（P ﹥0.05）；假手术组大鼠精氨酰-tRNA合成酶 mRNA 相对表达量及蛋白相对表达量均低于脑缺血再灌注组再灌注6 h、12 h 及24 h（P ﹤0.05）；脑缺血再灌注组大鼠再灌注6 h 精氨酰-tRNA合成酶 mRNA 相对表达量及蛋白相对表达量均高于再灌注2 h 和12 h（P ﹤0.05）；脑缺血再灌注组大鼠再灌注24 h 精氨酰-tRNA合成酶 mRNA 相对表达量及蛋白相对表达量均高于再灌注12 h 和48 h（ P ﹤0.05）。结论大鼠局灶性脑缺血再灌注后6 h 及24 h 精氨酰-tRNA合成酶表达明显升高，可能与缺血和再灌注两次损伤有关。%Backgrounds At present,aminoacyl - tRNA synthetase related clinical studies mostly concentrate on the structure and function of arginyl - tRNA synthetase,leucyl - tRNA synthetase,methionyl - tRNA synthetase and alanyl - tRNA synthetase,research reports about relationship between aminoacyl - tRNA synthetase and cerebral ischemia is relatively rare. Objective To explore the expression of arginyl - tRNA synthetase in rats with focal cerebral ischemia reperfusion and the mechanism. Methods At first,4 out of 60 clean healthy male SD rats were selected to carry out the preliminary experiment, then the other 56 rats were randomly divided into A group(n = 8),B group(n = 8) and C group(n = 40);focal cerebral ischemia reperfusion model was prepared by suture - occluded method(by embolization of middle cerebral artery). Rats of A group did not received any surgical intervention,rats of C group prepared for focal cerebral ischemia reperfusion model,while rats of B group received same surgical intervention without line - lock of internal carotid or cerebral ischemia. RT - PCR was used to detect the relative expression quantity of arginyl - tRNA synthetase mRNA,Western Blotting method was used to detect the relative expression quantity of arginyl - tRNA synthetase protein. Results After 2 hours and 48 hours of reperfusion,no statistically significant differences of relative expression quantity of arginyl - tRNA synthetase mRNA or arginyl - tRNA synthetase protein was found between B group and C group(P ﹥ 0. 05);after 6 hours,12 hours and 24 hours of reperfusion,the relative expression quantity of arginyl - tRNA synthetase mRNA and arginyl - tRNA synthetase protein of B group were statistically significantly lower than those of C group(P ﹤ 0. 05). After 6 hours of reperfusion,the relative expression quantity of arginyl -tRNA synthetase mRNA and arginyl - tRNA synthetase protein of C group were statistically significantly higher than those after 2 hours and 12 hours of reperfusion(P ﹤ 0. 05);after 24 hours of reperfusion,the relative expression quantity of arginyl - tRNA synthetase mRNA and arginyl - tRNA synthetase protein of C group were statistically significantly higher than those after 12 hours and 48 hours of reperfusion(P ﹤ 0. 05). Conclusion After 6 hours and 12 hours of reperfusion,the expression of arginyl -tRNA synthetase is significantly elevated in rats with focal cerebral ischemia reperfusion,its mechanism is possibly related with the double damage of ischemia and reperfusion.