ApoE肽段预处理通过调节c-jun氨基端激酶阻抑小鼠脑缺血再灌后神经细胞的凋亡

Role of apoE-mimetic peptide preconditioning in regulation of c-jun N-terminal protein kinase on the neuronal apoptosis following focal cerebral ischemia reperfusion in mice

目的:为探讨外源性载脂蛋白E(apoE)肽段对局灶性脑缺血再灌(I/R)的保护机制,观察apoE-1410拟肽对I/R小鼠脑组织磷酸化c-jun氨基端激酶(JNK)表达变化的影响。方法:实验选用160只雄性ICR小鼠,随机分为3组:假手术对照组(sham组),模型组(I/R组),apoE治疗组(apoE组),采用可逆性大脑中动脉栓塞(MCAO)模型,术后行神经功能评分。I/R3、6、12、24、48h行HE染色观察皮质区神经细胞的形态变化,并计算存活神经元的数目。利用WesternBlot法、免疫组织化学法检测p-c-jun的表达,行TUNEL法检测凋亡细胞数。结果:与对照组比较,模型组小鼠神经功能评分均降低,I/R12h后存活神经元数目明显减少;p-c-jun阳性反应术后3、6、12、24h显著增高(P<0.05);随缺血时间延长,凋亡细胞增多,并于48h达高峰(P<0.05)。与模型组比较,治疗组小鼠神经功能评分增加,各时相点p-c-jun和TUNEL表达均不同程度下调(P<0.05)。I/R3h至48h皮质区p-c-jun表达与TUNEL呈正相关(r=0.716,P<0.01)。结论:缺血侧皮质区p-c-jun表达的增强可诱导I/R后神经细胞凋亡;拟apoE-1410肽对I/R具有一定的治疗作用,其机制之一可能是通过抑制JNK活化实现的。

Objective：In order to study the mechanism unerlying the protection of brain by apoE-mimetic peptide in case of focal cerebral ischemia reperfusion（I/R）,and investigate the effect of apoE-1410 peptide on the changes of c-jun N-terminal protein kinase（JNK） expression in cortex in mice.Methods：A total of 160 male ICR mice were randomly divided into sham operation group（sham group）,model group（I/R group） and apoE treatment group（apoE）.The middle cerebral artery occlusion（MCAO） model of mice were performed,and then the neurological function scores were examined.The tissues of cortex were sampled at specified time points（I/R 3,6,12,24,48 h）,and changes were observed by the HE staining method,the number of living/apoptotic neurons was counted.The expression of p-c-jun was determined by immunohistochemical method and Western Blot assay,the apoptosis of neurons was detected by TUNEL method.Results：In I/R group,the neurological function scores were significantly lower,the living neurons were much less than those in sham group,the p-c-jun level was significantly enhanced at 3,6,12 and 24 h（P0.05）,the number of apoptosis cells increased gradually following the ischemic reperfusion time-proloned and were peaked at 48 h（P0.05）.The neurological function scores of apoE group were increased,and the expression of p-c-jun and TUNEL were lower than those in the I/R group.The quantification of p-c-jun expression in cortex was positively correlated with TUNEL positive cells in I/R 3 h to 48 h（r=0.716,P0.01）.Conclusion：The increased expression of p-c-jun in cortex may induce neuronal apoptosis after I/R,apoE-1410 peptide may possess the treatmental role for I/R by inhibiting the activation of the JNK signaling pathway.