一种新的载脂蛋白E拟肽对小鼠局灶性脑缺血再灌后c-Jun氨基端激酶通路的影响

Role of a novel apoE-mimetic peptide in regulation of JNK signaling pathway on the nerve cell apoptosis of mice following focal cerebral ischemia reperfusion

目的：研究小鼠局灶性脑缺血再灌（Ⅰ/R）后c-Jun氨基端激酶（JNK）通路在皮质神经元凋亡中的作用,探讨新的拟apoE-1410肽通过此途径发挥的保护机制.方法：建立小鼠可逆性大脑中动脉栓塞（MCAO）模型,在预定时间点观察皮质区组织学变化,采用免疫组织化学法检测p-c-Jun、 caspase-3的表达;免疫印迹法检测p-c-jun的表达,TUNEL法检测皮质区凋亡细胞数.结果：与对照组比较,模型组12h后存活神经元数目明显降低;各组小鼠p-c-Jun、 caspase-3阳性反应均有不同程度增强;随缺血时间延长,凋亡细胞增多,并于24h达高峰.与模型组比较,治疗组各时相点3项指标的表达均不同程度下调. Ⅰ/R 3h～24h皮质区p-c-Jun表达与caspase-3呈正相关;caspase-3与TUNEL呈正相关.结论：缺血侧皮质区p-c-Jun表达的增强可能通过激活caspase依赖的凋亡途径诱导Ⅰ/R后神经元凋亡;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通路活化实现的.

Objective： To investigate the expression and mechanism of c-Jun N-terminal protein kinase （JNK） signaling pathway in the nerve cell apoptosis in the cortex in mice after focal cerebral ischemia reperfusion （ I/R）, and to approach the role of apoE-1410 in nervous system protection and molecular mechanism of JNK signaling pathway. Methods： To replicate the middle cerebral artery occlusion model of mice, the mice were fixed at specified time points, the tissues of the cortex were sampled, the histopathologic changes were observed with HE staining, and the number of living/apoptotic neurons was counted separately. The expression of p-c-Jun and caspase-3 was determined by immunohistochemical method, Western blot assay was used to examine the p-c-Jun, and the apoptosis of nerve cells was detected with TUNEL method. Results： In the I/R group, the living neurons were much less than those in the sham group, the p-c-Jun, caspase-3 level and the number of apoptosis cells was significantly enhanced, and a lot of apoptosis cells were observed and peaked at 24 h. The expression of p-c-Jun, caspase-3 and the number of apoptosis cells were lower in the apoE group than those in the I/R group. The quan- tification of p-c-Jun expression was positively correlated with caspase-3, and the expression of p-c-Jun was positively correla- ted with the TUNEL positive cells after I/R. Conclusion： The increased expression of p-c-Jun in the cortex may induce neuronal apoptosis by activation of caspase signaling pathway after I/R. ApoE-1410 reduced early brain injury and apoptosis of nerve cells by inhibiting the activation of the JNK signaling pathway.