创伤后神经元自噬水平下降及谷氨酸介导的凋亡增多可导致创伤后癫痫易患性增加  

Decrease in autophagy and increase in glutamate-mediated apoptosis after trauma may lead to increased incidence of post-traumatic epilepsy

作　　者：王洁[1] 柴晓洋 李昱晨 陆克义[3] WANG Jie;CHAI Xiao-yang;LI Yu-chen;LU Ke-yi(Department of Neurology,Fisrt Hospital of Shanxi Medical University,Taiyuan 030001,China;Beijing Luhe Hospital,Capital Medical University,Beijing 100069,China;Department of Nuclear Medicine,First Hospital of Shanxi Medical University,Taiyuan 030001,China)

机构地区：[1]山西医科大学第一医院神经内科,山西太原030001 [2]首都医科大学附属北京潞河医院,北京101149 [3]山西医科大学第一医院核医学科,山西太原030001

出　　处：《中国病理生理杂志》2022年第11期1945-1951,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81501132);山西省卫生计生委科研项目(No.2014034)。

摘　　要：目的:研究创伤后癫痫(post-traumatic epilepsy,PTE)潜伏期内的神经组织凋亡水平改变对PTE易患性增加的作用,进一步探讨谷氨酸(glutamate,Glu)及自噬改变在该过程中的作用。方法:利用Noble-Collip鼓制备轻型创伤性脑损伤(traumatic brain injury,TBI)大鼠模型,4 d后大脑皮层下注射小剂量FeCl_(2)观察其对PTE发生率的影响;通过ELISA法检测轻型TBI后大脑皮层神经细胞凋亡相关蛋白水平,预先给予凋亡抑制剂zVAD-FMK之后观察皮层神经细胞凋亡变化及其对大鼠PTE发生率的影响;检测PTE潜伏期内脑组织中Glu含量,通过干预Glu水平观察其对PTE易患性的影响;通过Western blot法检测脑组织中自噬相关蛋白水平,观察其在这一过程中的作用。结果:与sham组相比,轻型TBI后大鼠7~90 d内PTE的发生率为11.2%,差异有统计学意义(P<0.01),提示PTE模型制备成功。在造模后4 d向大鼠大脑皮层注射明显低于致痫剂量的FeCl_(2),结果显示TBI+FeCl_(2)组73.2%大鼠出现癫痫发作,提示轻型TBI后PTE易患性增加。TBI后给予FeCl_(2)注射的大鼠海马神经细胞凋亡明显增多,预先给予凋亡抑制剂zVAD-FMK后,TBI+FeCl_(2)+zVAD-FMK组PTE发生率下降至32.6%,与TBI+FeCl_(2)组相比差异有统计学意义(P<0.01)。与sham+FeCl_(2)组相比,TBI+FeCl_(2)组脑内Glu含量增多(P<0.01);预先给予Glu释放抑制剂riluzole,可以显著减少神经组织凋亡并降低PTE发生率;预先给予线粒体膜稳定剂cyclosporine A可以显著降低caspase-3及caspase-9活性(P<0.01);而给予自由基清除剂超氧化物歧化酶(superoxide dismutase,SOD)后,caspase-3和caspase-9活性均显著降低(P<0.05或P<0.01)。TBI+FeCl_(2)组大脑皮层组织的微管相关蛋白1轻链3-Ⅱ(microtubule-associated protein 1 light chain 3-Ⅱ,LC3-Ⅱ)及beclin-1蛋白水平显著下降(P<0.01);预先给予自噬激动剂rapamycin后,自噬水平升高,且caspase-3水平下降。结论:创伤后神经元自噬水平下降及谷氨酸介导�AIM:To detect the changes of neuronal apoptosis level during the latent period of post-traumatic epilepsy(PTE),and to explore the roles of apoptosis,glutamate(Glu)and autophagy in the increased sensitivity of PTE.METHODS:Mild traumatic brain injury(TBI)rat model was prepared by Noble-Collip drum.After 4 d,small dose of FeCl_(2) was injected into the subcortical cortex to observe the incidence of PTE.The apoptosis of cerebral cortex neurons after mild TBI was detected.After injection of zVAD-FMK,an inhibitor of apoptosis,the changes of apoptosis and its effect on the incidence of PTE was observed.The content of Glu in brain tissues was detected during the latent period of PTE,and its effect on the increased susceptibility of PTE was observed.The level of autophagy in brain tissues was detected by Western blot.RESULTS:Compared with sham group,the incidence of PTE in 7 to 90 d after mild TBI was 11.2%(P<0.01),suggesting that the PTE model was successfully constructed.On the 4th day after modeling,FeCl_(2) was injected into the cerebral cortex at a dose lower than the epileptic dose.The results showed that 73.2% of the rats in the experimental group had seizures in TBI+FeCl_(2) group(P<0.01),which suggested that PTE susceptibility was increased after mild TBI.After TBI,the apoptosis of hippocampal neurons was significantly increased by giving FeCl_(2).Compared with TBI+FeCl_(2) group,the incidence of PTE in the TBI+FeCl_(2)+zVAD-FMK group was decreased to 32.6%(P<0.01).Compared with TBI+FeCl_(2) group,Glu content in brain tissues was increased in TBI+FeCl_(2) group(P<0.01).After the administration of Glu-releasing inhibitor riluzole,the incidence of PTE and neuronal apoptosis were decreased significantly.The activity of caspase-3 and caspase-9 was significantly decreased after treatment with cyclosporine A(P<0.01).After administration of superoxide dismutase(SOD),both caspase-9 and caspase-3 were decreased.The protein levels of microtubule-associated protein 1 light chain 3-Ⅱ(LC3-Ⅱ)and beclin-1 were significantly

关 键 词：创伤后癫痫 谷氨酸 细胞凋亡 自噬 

分 类 号：R741.02[医药卫生—神经病学与精神病学] R363.2[医药卫生—临床医学]