Z-ATAD-FMK预处理对癫痫持续状态后大鼠海马Caspase-12表达的影响和神经元的保护作用

Effects of Z-ATAD-FMK pretreatment on the expression of Caspase-12 and neuronal protection in hippocampus of rats after status epilepticus

目的:探讨Caspase-12抑制剂(苄氧羰-丙氨酰-苏氨酸-丙氨酰-天冬氨酰-氟甲基酮,Z-ATAD-FMK)对大鼠癫痫持续状态(status epilepticus,SE)后海马Caspase-12表达及神经细胞损伤的影响。方法:80只雄性SD大鼠,随机分成正常对照组(A组,8只)、二甲基亚砜(DMSO)对照组(B组,24只)、SE组(C组,24只)、Z-ATAD-FMK预处理组(D组,24只)。后3组按时间点分成3个亚组,分别为12 h组、24 h组、72 h组,每亚组各8只。采用氯化锂-匹罗卡品法制作SE大鼠模型,B、C、D组均于造模前2 h分别于右侧脑室内注入DMSO 5μL、DMSO 5μL、Z-ATAD-FMK 5μL。采用免疫组化法检测大鼠海马Caspase-12蛋白的表达变化;光镜和电镜下分别观察大鼠海马病理学和神经细胞超微结构的改变情况。结果:A组和B组大鼠海马CA1区均有少量Caspase-12蛋白表达,但A组与B组之间差异无统计学意义(P>0.05);C组大鼠海马CA1区Caspase-12蛋白表达在SE后12 h开始增加,24 h进一步升高,于72 h达最高点,且均较A和B组各时间点显著增高(P<0.01);D组24 h、72 h时间点Caspase-12蛋白表达的动态变化与C组相似,但均较C组明显降低(P<0.05或P<0.01)。光镜和电镜观察结果显示SE后D组海马神经细胞的损伤情况较C组减轻。结论:Z-ATAD-FMK能选择性地阻断Caspase-12的活性,并使神经元损伤减轻,提示Z-ATAD-FMK对惊厥性脑损伤可能有保护作用。

Objective： To investigate the effect of Caspase-12 inhibitor（Z-Ala-Thr-Ala-Asp-fluoromethyl ketone,Z-ATAD-FMK）on expression of Caspase-12 and neuronal damage in the rat hippocampus after status epilepticus（SE）.Methods： Eighty male Sprague-Dawley（SD）rats were randomly enrolled into normal control group（A,8 rats）,Dimethyl Sulfoxide（DMSO）control group（B,24 rats）,status epilepticus group（C,24 rats）and Z-ATAD-FMK treatment group（D,24 rats）.The last three groups were further divided into three subgroups at different time point,which were executed respectively at 12 h,24 h and 72 h after status epilepticus discontinued.The SE models were constructed using lithium-pilocarpine.5μL DMSO,5μL DMSO,5μL Z-ATAD-FMK were separately administered into the right lateral ven-tricles of rats in the last three groups 2 h before the beginning of SE.The expression levels of Caspase-12 protein in the hippocampus CA1 region were detected with immunohistochemistry（IHC）;HE staining was used to view the histopathological changes in hippocampus and the ultrastructural changes were obserred under electron microscope.Results： The immunohis-tochemistry results indicate that a little expression of Caspase-12 protein in group A and B,which had no statistically significant each other.In the group C,the expressions of both protein begin to increase markedly at 12 h after SE,and elevated gradually after the time passed,reached the maximum at 72 h,and both significantly upregulated compared with group A and B（P 0.01）,nevertheless,the expression tendency in the group D was similar to group C,the expression of Caspase-12 in group D was notably lower than that in group C at 24 h and 72 h（P 0.05 or P 0.01）.The neuron changes of subgroups of group D were more lighten than that the corresponding group C.Conclusion： The activation of Caspase-12 is selectively blocked by the Caspase-12 inhibitor（Z-ATAD-FMK）,which decreases the degree of neuronal damage,indicat-ing that Z-ATAD-FMK may have protective effect against the damage caused by status epilepticus.