依达拉奉对惊厥持续状态大鼠海马IRE1 mRNA表达及神经元凋亡的影响

Effects of edaravone on IRE1 mRNA expression and neuronal apoptosis in the hippocampus of rats with status convulsivus

摘要:

目的：探讨依达拉奉（EDA）对惊厥持续状态（SC）后大鼠海马内质网应激关键标志分子IRE1表达及神经元凋亡的影响。方法：将19~21 d日龄的Sprague-Dawley大鼠随机分为SC组、EDA组、对照组。其中SC组、EDA组再按SC后处死时间点不同分别分为4，12，24，48，72 h 5个亚组；每组均8只。用RT-PCR检测SC后海马IRE1 mRNA的表达。用原位细胞凋亡检测法（TUNEL）观察神经元凋亡细胞数。电镜观察海马CA1区神经元超微结构变化。结果：①RT-PCR 结果：SC组4 h和12 h时间点IRE1 mRNA含量较对照组明显升高，差异有非常显著性（P<0.01）；EDA组4，12，24 h时间点IRE1 mRNA含量显著高于SC组对应时间点及对照组的含量，差异有非常显著性（P<0.01）。②TUNEL结果：SC组12~72 h时间点TUNEL阳性细胞数多于对照组，差异有显著性（P<0.01），且SC组TUNEL阳性细胞数随惊厥持续时间延长而增加。EDA组12~72 h时间点TUNEL阳性细胞数较SC组明显减少，差异有显著性（P<0.05或P<0.01），但仍高于对照组，差异有显著性（P<0.05或P<0.01）。③ 电镜观察结果：SC组及EDA组4 h后即开始出现核周细胞器的改变，12 h后出现明显细胞核的改变。其中SC组细胞凋亡严重，EDA组凋亡较轻。结论：EDA可能通过上调IRE1的表达及维持其活性而缓解内质网压力而发挥对神经元的保护作用,有望成为在体抑制内质网应激的有效药物。 ［中国当代儿科杂志，2009，11（6）：471-475］

Abstract:

OBJECTIVE: To investigate the expression of the key marker of endoplasmic reticulum stress (ERS) IRE1 mRNA and neuronal apoptosis in the rat hippocampus after status convulsivus (SC), and the intervention effects of edaravone, a novel free radical scavenger. METHODS: Sprague-Dawley (SD) rats aged 19-21 days were randomly assigned to three groups: normal control, SC and edaravone-treated SC. SC was induced in the later two groups. The two groups were subdivided into 5 groups sacrificed at 4, 12, 24, 48, and 72 hrs after SC induction. IRE1 mRNA expression in the hippocampus was detected by RT-PCR. Neuronal apoptosis was observed by TdT-mediated dUTP nick end labeling (TUNEL). The ultramicrostructural changes of neuron were observed by electron microscopy. RESULTS: IRE1 mRNA expression was obviously up-regulated 4 and 12 hrs after SC compared with the normal control group (P<0.01). IRE1 mRNA expression in the edaravone-treated SC group was notably higher than the untreated SC group 4, 12 and 24 hrs after SC and the normal control group (P<0.01). TUNEL positive cells in the hippocampus in the untreated SC group were significantly more than those in the normal control group (P<0.01). The number of TUNEL positive cells increased with the prolonged convulsion time. TUNEL positive cells in the edaravone-treated SC group were significantly reduced compared with those in the untreated SC group 12, 24, 48 and 72 hrs after SC (P<0.05 or P<0.01), but remained higher than the normal control group (P<0.05 or P<0.01). The peri-nucleus cell organ injuries were observed 4 hrs after SC and karyopycnosis and cytoplasm condensation were observed 12 hrs after SC in the SC and the edaravone-treated SC groups. The edaravone-treated SC group demonstrated less severe apoptosis than the untreated SC group. CONCLUSIONS: Edaravone may have neuroprotections against SC by an up-regulation of IRE1 expression. It might serve as an effective agent for reducing ERS in vivo.［Chin J Contemp Pediatr, 2009, 11 (6):471-475］