雷帕霉素通过激活LC3介导的细胞自噬改善ApoE4/5×FAD小鼠的认知功能  

Activation of LC3-mediated autophagy by rapamycin improves cognitive function of ApoE4/5×FAD mice

作　　者：崔晓丽 魏振 沈辉[3] 张静 戴晓曼 张健[5] 陈晓春[5] 曾育琦[5] CUI Xiao-li;WEI Zhen;SHEN Hui;ZHANG Jing;DAI Xiao-man;ZHANG Jian;CHEN Xiao-chun;Zeng Yu-qi(Department of Geriatric,The People's Hospital Affiliated to Fujian University of Traditional Chinese Medicine;Depart-ment of Neurology,Fujian Provincial Hospital,Shengli Clinical Medical College of Fujian Medical University;Department of Geriatrics,Fujian Medical University Union Hospital;Fujian Institute of Geriatrics,Fujian Medical University Union Hospital;Department of Neurology,Fujian Medical University Union Hospital,Fuzhou 350001,China)

机构地区：[1]福建中医药大学附属人民医院老年病科暨干部病房 [2]福建省立医院神经内科,福建医科大学省立临床医学院 [3]福建医科大学附属协和医院干部病房 [4]福建医科大学附属协和医院福建省老年医学研究所 [5]福建医科大学附属协和医院神经内科,福建福州350001

出　　处：《中国病理生理杂志》2021年第8期1400-1408,共9页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81601116);福建省卫生健康科技计划项目(No.2016-ZQN-23;No.2017-ZQN-22);福建省自然科学基金(No.2016J01554;No.2020J05116)。

摘　　要：目的:探讨表达不同人载脂蛋白E(ApoE)亚型的5个家族性阿尔茨海默病(FAD)突变基因(5×FAD)转基因小鼠(EFAD小鼠)的海马组织细胞自噬功能是否存在差异,以及雷帕霉素能否通过增强细胞自噬减少β-淀粉样蛋白(Aβ)沉积而改善EFAD小鼠的认知功能。方法:6月龄ApoE3/5×FAD(E3FAD)小鼠和ApoE4/5×FAD(E4FAD)小鼠每日给予雷帕霉素(1 mg/kg)灌胃,持续6周,给药结束后通过Morris水迷宫检测小鼠的学习记忆能力,透射电镜观察各组自噬体超微结构特点,免疫组织化学染色法观察脑内淀粉样斑块沉积情况,Westernblot方法检测海马内自噬相关蛋白p-mTOR和LC3-II的表达,ELISA法测定海马组织可溶性和不溶性Aβ_(1-42)水平。结果:与溶媒组E3FAD小鼠相比,溶媒组E4FAD小鼠Morris水迷宫训练潜伏期延长(P<0.05),探索实验时穿越平台次数少(P<0.05),距离平台平均距离更长(P<0.05)。雷帕霉素干预可改善E3FAD和E4FAD小鼠的认知功能,降低2组小鼠海马组织可溶性Aβ_(1-42)水平和E4FAD小鼠海马组织不溶性Aβ_(1-42)水平(P<0.05),缩小了E3FAD和E4FAD小鼠之间的认知差距(P<0.01)。同时,溶媒组E4FAD小鼠马组织自噬标志物LC3-II表达比E3FAD小鼠低(P<0.05),雷帕霉素可降低E3FAD和E4FAD小鼠海马组织p-mTOR水平(在E4FAD小鼠中作用更为明显)、升高LC3-II表达、增强细胞自噬功能(P<0.05)。结论:ApoE4加重了5×FAD转基因小鼠的认知功能障碍,增加了脑内Aβ沉积和细胞自噬。雷帕霉素通过抑制mTOR、激活LC3介导的自噬减轻Aβ沉积,从而改善EFAD小鼠的认知功能。AIM:To investigate whether there are differences in autophagy in the transgenic mice that coexpress 5 familial Alzheimer disease(FAD)mutations(5×FAD)and different human apolipoprotein E(ApoE)isoforms(EFAD mice),and whether rapamycin reduces the deposition of amyloidβ-protein(Aβ)and improves the cognitive function of EFAD mice by enhancing autophagy in hippocampus.METHODS:Six-month-old ApoE3/5×FAD(E3FAD)mice and ApoE4/5×FAD(E4FAD)mice were administered intragastrically with rapamycin(1 mg/kg)or vehicle every day for 6 weeks.Spatial learning and memory of EFAD mice were eveluated by the modified Morris water maze test.Transmission electron microscope was used to observe ultrastructural characteristics of autophagy.Immunohistochemistry was used to observe the amyloid plaque deposits in the hippocampus.The protein levels of autophagy-related molecules,p-mTOR/mTOR and LC3-II/LC3-I,in the hippocampus were determined by Western blot.Aβ_(1-42)levels in the hippocampus were measured by ELISA.RESULTS:Compared with vehicle-treated E3FAD mice,the escape latency and the swimming distance toward to the hidden platform were extended in vehicle-treated E4FAD mice,while the number of crossing the previous platform position was reduced(P<0.05).After rapamycin administration,the cognitive function of both E3FAD and E4FAD mice was improved(P<0.01),and the gap of cognition between the 2 groups was narrowed.The accumulation of Aβ,including soluble Aβ_(1-42)in both E3FAD mice and E4FAD mice,and insoluble Aβ_(1-42)in E4FAD mice,were reduced(P<0.05).The autophagy marker LC3-II/LC3-I ratio was reduced in E4FAD-vehicle group compared with E3FAD-vehicle group(P<0.05).Rapamycin inhibited the level of p-mTOR/mTOR,increased the LC3-II/LC3-I ratio,and enhanced autophagy in EFAD mice especially E4FAD mice(P<0.05).CONCLUSION:ApoE4 aggravated cognitive impairment,and increased Aβdeposition and autophagy dysfunction in the brain of 5×FAD transgenic mice.Rapamycin may reduce Aβdeposition and improve the cognitive function of EFAD mice by

关 键 词：雷帕霉素 自噬 阿尔茨海默病 载脂蛋白E4 Β-淀粉样蛋白 

分 类 号：R749.16[医药卫生—神经病学与精神病学] R363.2[医药卫生—临床医学]