缺血预处理通过抑制精氨酰-tRNA合成酶和Caspase-3表达减少脑梗死大鼠脑细胞的凋亡

Ischemic precondition reduces apoptosis of brain cells in rats after cerebral infarction by inhibiting arginyl-tRNA synthetase and Caspase-3 expressions

目的探讨缺血预处理对脑梗死大鼠脑细胞的保护作用及其机制。方法120只SD大鼠按随机数字表法分为缺血预处理组(50只)、脑梗死组(50只)、假手术组(10只)、正常组(10只),前2组大鼠采用改良Longa线栓法制备大脑中动脉闭塞(MCAO)致脑梗死模型(缺血预处理组大鼠在造模前将线栓置入大脑中动脉阻断血流15 min),假手术组大鼠行假缺血预处理和假阻断大脑中动脉血流,正常组大鼠不做任何处理。分别于正常组和假手术组造模后24 h,缺血预处理组和脑梗死组造模后2 h、6 h、12 h、24 h、48 h,每组取10只大鼠,采用TUNEL染色检测小鼠缺血半暗带区细胞凋亡率,免疫组化染色检测小鼠缺血半暗带区含半胱氨酸的天冬氨酸蛋白水解酶-3(Caspase-3)的表达,RT-PCR和Western blotting法分别检测小鼠缺血半暗带区精氨酰-tRNA合成酶(ArgRs)基因和ArgRs蛋白的表达。结果与正常组和假手术组比较,脑梗死组大鼠造模后不同时间点缺血半暗带区细胞凋亡率、Caspase-3阳性表达率、ArgRs基因和ArgRs蛋白的表达均明显升高,差异均有统计学意义(P<0.05)。与脑梗死组比较,缺血预处理组大鼠造模后不同时间点缺血半暗带区细胞凋亡率、Caspase-3阳性表达率、ArgRs基因和ArgRs蛋白的表达均明显降低,差异有统计学意义(P<0.05)。结论脑缺血预处理可以抑制脑梗死大鼠脑缺血半暗带区ArgRS基因及ArgRS蛋白、Caspase-3的表达,减少脑细胞的凋亡,从而起到脑保护作用。

Objective To study the protective role of ischemic precondition in brain cells and its mechanism in rats after cerebral infarction.Methods According to random number table method,120 SD rats were divided into ischemic precondition group(n=50),cerebral infarction group(n=50),sham-operated group(n=10),and normal control group(n=10);cerebral infarction models of middle cerebral artery occlusion(MCAO)in the former two groups were induced by modified Longa thread method;precondition(blood flow in the middle cerebral artery was blocked for 15 min before model making)was given to the ischemic precondition group;rats in the sham-operated group were performed sham ischemic precondition and sham occlusion of middle cerebral artery blood;rats in the normal control group did not receive any treatment.At 24 h after model making,rats in the normal control group(n=10)and sham-operated group(n=10)were sacrificed,and 2,6,12,24,and 48 h after model making,rats in the ischemic precondition group(n=10)and the cerebral infarction group(n=10)were sacrificed;the Caspase-3 expression in the ischemic penumbra was detected by immunohistochemistry,and the apoptosis rate in the ischemic penumbra was determined by TUNEL.Real-time PCR and Western blotting were used to detect the mRNA and protein expressions of arginyl-tRNA synthetase(ArgRS).Results As compared with those in the normal control group and sham-operated group,the apoptosis rate,positive expression rate of Caspase-3,and ArgRs mRNA and protein expressions in the ischemic penomere zone of the rats in the cerebral infarction group were significantly increased at different time points after modeling(P<0.05).As compared with those in the cerebral infarction group,the apoptosis rate,positive expression rate of Caspase-3,and ArgRs mRNA and protein expressions in the ischemic penomere zone at different time points after modeling in the ischemic precondition group were significantly decreased(P<0.05).Conclusion Cerebral ischemic precondition can inhibit the mRNA and protein expressions of ArgRS and Caspase-3 expression,reduce the apoptosis of brain cells,and thus play a neuroprotective role in brain tissues.