左乙拉西坦通过抑制NLRP3炎症小体的活化和氧化应激改善APP/PS1小鼠的认知障碍

Levetiracetam Ameliorates Cognitive Impairment in APP/PS1 Mice by Inhibiting NLRP3 Inflammasome Activation and Oxidative Stress

关键词:  左乙拉西坦  阿尔茨海默病  神经炎症  炎症小体  认知障碍

Key words:  levetiracetam  Alzheimer's disease  neuroinflammation  inflammasome  cognitive impairment

摘要:

目的 以氧化应激和神经炎症为切入点，考察左乙拉西坦治疗阿尔茨海默病的药效和潜在机制。方法 淀粉样蛋白前体蛋白/I型早老素双转基因（amyloid precursor protein-swe/presenilin-1-1dE9，APP/PS1）模型小鼠按照体质量每2 d通过尾静脉注射不同剂量左乙拉西坦（5，10，20 mg·kg^-1），连续给药30 d。自主活动试验考察实验动物自主活动能力；Morris水迷宫试验考察实验动物认知能力；免疫组化试验考察实验动物海马脑区小胶质细胞活化程度；半胱天冬酶-1（caspase-1）活力检测试剂盒考察实验动物海马脑区caspase-1活力；蛋白免疫印迹试验考察实验动物海马脑区白介素-1β（interleukin-1β，IL-1β）、核苷酸结合寡聚化结构域样受体家族pyrin结构域蛋白3（NOD-，LRR-and pyrin domain-containing protein 3，NLRP3）、凋亡相关斑点样蛋白（apoptosis-associated speck-like protein containing a CARD，ASC）、caspase-1蛋白表达水平；比色法试剂盒检测实验动物海马脑区超氧化物歧化酶（superoxide dismutase，SOD）活力、谷胱甘肽（glutathione，GSH）水平、过氧化氢（H₂O₂）含量和丙二醛（malondialdehyde，MDA）水平。结果 与空白组小鼠相比，APP/PS1模型组小鼠的逃避潜伏期显著延长，且海马脑区小胶质细胞活化程度明显加剧，IL-1β水平明显增加，NLRP3和caspase-1蛋白水平明显增加，SOD活力显著下降，GSH水平显著减少，MDA水平显著增加。与模型组相比，左乙拉西坦10 mg·kg^-1和20 mg·kg^-1可以显著缩短APP/PS1小鼠逃避潜伏期，显著减轻海马脑区小胶质细胞活化程度，显著降低IL-1β水平，显著减少NLRP3和caspase-1蛋白水平，显著增加SOD活力和GSH水平、显著减少MDA水平。结论 左乙拉西坦可以显著改善APP/PS1小鼠的认知功能，其作用机制可能与抑制炎症小体的活化和氧化应激密切相关。

Abstract:

OBJECTIVE To explore the efficacy and mechanism of levetiracetam in the treatment of Alzheimer's disease from the perspective of oxidative stress and neuroinflammation. METHODS Amyloid precursor protein-swe/presenilin-1- 1dE9(APP/PS1) model mice were injected with different doses of levetiracetam(5, 10, 20 mg·kg^-1) through tail vein every two days for a total 30 consecutive days. The locomotor activity experiment was used to investigate the locomotor ability of APP/PS1 mice. Morris water maze experiment was used to investigate the spatialcognitive ability of APP/PS1 mice. Immunohistochemistry experiment was used to evaluate the degree of microglia activation in hippocampus of APP/PS1 mice. Caspase-1 activity of hippocampus in APP/PS1 mice was tested by kit. The expressionlevel of caspase-1 was detected by Western blotting. The expression levels of interleukin(IL)-1β, NOD-, LRR- and pyrin domain-containing protein 3(NLRP3), apoptosis-associated speck-like protein containing a CARD domain (ASC) and caspase-1 activityin hippocampus were detected by colorimetric assay. The superoxide dismutase(SOD) activity, glutathione(GSH) level, H₂O₂ level and malondialdehyde(MDA) level in hippocampus were detected by colorimetric kit. RESULTS Compared with control group, the escape latency in Morris water maze and the activation degree of microglia in hippocampus was significantly increased, and the protein level of IL-1β, NLRP3 and caspase-1 was obviously increased in APP/PS1 group. In addition, the SOD activity and GSH level was significantly reduced with MDA level increased in APP/PS1 model group. Compared with APP/PS1 group, levetiracetam 10 mg·kg^-1 and 20 mg·kg^-1 significantly reduced the escape latency and microglia activation, and obviously decreased the protein level of IL-1β, NLRP3 and caspase-1, significantly enhanced SOD activity and elevated GSH level while decreased MDA level. CONCLUSION Levetiracetam can significantly improve the cognitive function of APP/PS1. Its mechanism may be related to inhibiting the NLRP3 inflammasome activation and oxidative stress.