摘要
目的:探讨黄连解毒汤对阿尔茨海默病(AD)大鼠学习记忆能力和海马区NLRP3炎症通路中NOD样受体热蛋白结构域3(NLRP3)、天冬氨酸蛋白酶-1(Caspase-1)和白细胞介素-1β(IL-1β)表达的影响。方法:健康雄性SD大鼠70只,通过Morris水迷宫筛选出合格大鼠60只,随机抽取10只作为假手术对照组,其余50只采用双侧海马注射β-淀粉样蛋白1-42(Aβ_(1-42))1μg/μL建立AD模型大鼠,随机分为AD模型组,石杉碱甲2×10^(-5)g/kg组、黄连解毒汤6、3、1.5 g/kg组,每组10只,假手术对照组注射等体积用于配置Aβ_(1-42)的溶剂,造模后连续灌胃4 w,1次/d,灌胃结束后Morris水迷宫法检测大鼠学习记忆能力;苏木素-伊红(HE)染色观察大鼠海马病理改变;酶联免疫吸附法(Elisa)检测大鼠血清IL-1β水平、蛋白免疫印迹法(Western Blot)、实时荧光定量PCR反应(Real-time PCR)检测大鼠海马组织中NLRP3、Caspase-1、IL-1β蛋白和mRNA的表达水平;免疫组化法(IHC)检测大鼠海马组织中NLRP3、Caspase-1、IL-1β、IBA-1蛋白表达。结果:与正常对照组比较,模型对照组大鼠逃避潜伏期显著升高(P<0.05),穿越平台次数、有效区域停留时间和运动距离显著降低(P<0.05或P<0.01);HE染色观察到模型对照组大鼠海马区神经细胞变性坏死,排列紊乱,数量减少;血清IL-1β水平显著升高(P<0.01);海马NLRP3、Caspase-1、IL-1β蛋白和mRNA表达明显上调(P<0.05或P<0.01),IHC检测NLRP3、Caspase-1和IBA-1蛋白表达明显升高(P<0.05或P<0.01)。与模型对照组比较,黄连解毒汤3、6 g/kg组可降低模型大鼠逃避潜伏期,增加模型大鼠穿越平台次数、有效区域停留时间和运动距离(P<0.05或P<0.01),降低血清IL-1β水平(P<0.05);下调大鼠海马NLRP3、Caspase-1、IL-1β蛋白和mRNA表达(P<0.05或P<0.01),保护神经元。结论:黄连解毒汤能改善AD大鼠学习记忆能力,保护神经元,抑制AD大鼠神经炎反应,其机制可能与下调NLRP3炎症通路中NLRP3及下游Caspase-1、IL-1β的表达有关。
Objective:To investigate the effects of Huanglian Jiedu Decoction on learning and memory abilities of rats with Alzheimer’s disease(AD)and the expression of nucleotide binding oligomerization domain like receptor protein 3(NLRP3),cysteine-dependent aspartate-directed protease-1(Caspase-1),and interleukin-1β(IL-1β)in the NLRP3 inflammatory pathway of rat hippocampus.Methods:Seventy healthy male SD rats were subjected to the Morris water maze test,from which 60 eligible ones were selected.Ten rats were randomly selected and classified into the sham operation group,and the remaining 50 were injected with 1μg/μLβ-amyloid 1-42(Aβ_(1-42))into bilateral hippocampi for inducing AD.They were then randomly divided into the AD model group,2×10^(-5)g/kg huperzine A group,and 6,3,and 1.5 g/kg Huanglian Jiedu Decoction groups,with 10 in each group.The ones in the sham operation group were injected with the same volume of Aβ_(1-42)solvent.After modeling,the rats were gavaged with the corresponding drugs,once a day,for four weeks.Then the learning and memory abilities of rats were detected in the Morris water maze test,and the pathological changes in hippocampus were observed by hematoxylin-eosin(HE)staining.The serum level of IL-1βwas detected by enzyme-linked immunosorbent assay(Elisa).The protein and mRNA expression levels of NLRP3,Caspase-1,and IL-1βin rat hippocampal tissue were assayed by real-time fluorescence quantitative PCR(Real-time PCR)and Western blotting,respectively.The protein expression levels of NLRP3,Caspase1,IL-1β,and IBA-1 in rat hippocampal tissue were measured by immunohistochemistry(IHC).Results:Compared with the normal control group,the model group exhibited prolonged escape latency(P<0.05),reduced number of platform crossings and time of stay in the effective area,and shortened distance of movement(P<0.05 or P<0.01).HE staining showed that the nerve cells in the hippocampus were degenerated,necrotic,disorderly arranged,and decreased.The serum level of IL-1βwas significantly increased(P<0.01).The protein and mRNA expression levels of NLRP3,Caspase-1,and IL-1βin the hippocampus were significantly up-regulated(P<0.05 or P<0.01),while the protein expression levels of NLRP3,Caspase-1,and IBA-1 were proved elevated by IHC(P<0.05 or P<0.01).Compared with the model group,Huanglian Jiedu Decoction at 3 and 6 g/kg shortened the escape latency of rats(P<0.05),increased the number of platform crossings and stay time in effective area,extended the movement distance(P<0.05 or P<0.01),lowered the serum IL-1β(P<0.05),down-regulated the mRNA and protein expression levels of NLRP3,Caspase-1,and IL-1βin hippocampus(P<0.05 or P<0.01),and protected the neurons.Conclusion:Huanglian Jiedu Decoction improves the learning and memory abilities of AD rats,protects neurons,and inhibits neuritis,which may be related to the down-regulation of NLRP3 and downstream Caspase-1 and IL-1βin NLRP3 inflammatory pathway.
作者
董秤均
曹蠡馨
屈艳秦
梁雨晴
杨燕
伍文彬
Dong Chengjun;Cao Lixin;Qu Yanqin;Liang Yuqing;Yang Yan;Wu Wenbin(Hospital of Chengdu University of TCM,Chengdu 610072)
出处
《中药药理与临床》
CAS
CSCD
北大核心
2021年第6期7-13,共7页
Pharmacology and Clinics of Chinese Materia Medica
基金
四川省科技厅应用基础项目(编号:2018JY0060)
成都中医药大学“杏林学者”学科人才提升计划(编号:QNXZ2018004)
关键词
黄连解毒汤
阿尔茨海默病
NOD样受体热蛋白结构域3
天冬氨酸蛋白酶-1
白细胞介素-1Β
Huanglian Jiedu Decoction
Alzheimer’s disease(AD)
nucleotide binding oligomerization domain like receptor protein 3(NLRP3)
cysteine-dependent aspartate-directed protease-1(Caspase-1)
interleukin-1β(IL-1β)
