决明子治疗高脂血症的网络药理学研究及斑马鱼实验验证  

Network pharmacology study of Cassiae Semen in treatment of hyperlipidemia and experimental validation in zebrafish

作　　者：周嫦 彭珑萍 董艺丹 马梦娇 周茂琳 李强[2] 王佑华[1] ZHOU Chang;PENG Longping;DONG Yidan;MA Mengjiao;ZHOU Maolin;LI Qiang;WANG Youhua(Longhua Hospital,Shanghai University of Traditional Chinese Medicine,Shanghai 200032,China;Children's Hospital of Fudan University,Shanghai 201102,China)

机构地区：[1]上海中医药大学附属龙华医院,上海200032 [2]复旦大学附属儿科医院,上海201102

出　　处：《上海中医药大学学报》2024年第4期71-80,共10页Academic Journal of Shanghai University of Traditional Chinese Medicine

基　　金：国家自然科学基金资助项目(8220153795,8210152696);上海市科技成果转化与产业化项目(15401902500)。

摘　　要：目的:基于网络药理学探讨决明子治疗高脂血症的作用机制,并通过斑马鱼实验进行验证。方法:在中药分子机制生物信息学注释数据库(BATMAN-TCM)和中药系统药理学分析平台(TCMSP)检索并筛选决明子活性成分及相关靶点,通过GeneCards、DisGeNET、在线人类孟德尔遗传数据系统(OMIM)检索得到与高脂血症有关的靶点,利用Venny 2.1.0平台获得药物与疾病的共同靶点。使用Cytoscape3.8.2绘制决明子-活性成分-作用靶点网络图,采用String数据库构建蛋白质-蛋白质相互作用(PPI)网络图,将决明子-高脂血症共同靶点通过注释、可视化和集成发现数据库(DAVID)进行基因本体(GO)功能及京都基因与基因组百科全书(KEGG)通路富集分析。根据网络药理学研究结果,推测橙黄决明素(AO)为决明子降血脂的有效作用成分,在此基础上进行斑马鱼实验验证。以蛋黄液高脂饮食诱导斑马鱼高脂血症模型,造模后给予AO干预,验证AO治疗高脂血症的效果及作用机制。首先进行AO对高脂血症斑马鱼的毒性实验,确定AO最大给药浓度;后续观察AO对高脂血症斑马鱼总胆固醇(TC)、三酰甘油(TG)含量及肝脏组织形态学的影响;采用实时荧光定量PCR检测核心靶点mRNA在高脂血症斑马鱼中的表达。结果:共获得包括AO在内的13个决明子活性成分,决明子作用于高脂血症的潜在靶点109个,包括核心靶点肿瘤坏死因子(TNF)、白介素(IL)-1β、前列腺素内过氧化物合酶2(PTGS2)、半胱氨酸天冬氨酸蛋白酶3(CASP3)、过氧化物酶体增殖物激活受体-γ(PPAR-γ)等,主要涉及IL-17信号通路、TNF信号通路、脂质和动脉粥样硬化相关作用通路等。斑马鱼验证实验结果显示:造模后斑马鱼出现明显的脂质聚积,AO可显著降低高脂血症斑马鱼组织中的TG、TC含量(P<0.05,P<0.01),减少肝脏组织内脂肪空泡和脂滴形成,并可显著下调核心靶点TNF-α、IL-1βmRNA表达(P<0.01)。Objective:To investigate the mechanisms of Cassiae Semen in treating hyperlipidemia based on network pharmacology and be validated through zebrafish experiments.Methods:The active ingredients and related targets of Cassiae Semen were searched and screened in the Bioinformatics Analysis Tool for Molecular Mechanisms of Traditional Chinese Medicine(BATMAN-TCM)and the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP).Hyperlipidemia-related target genes were obtained through searching GeneCards,DisGeNET and Online Mendelian Inheritance in man(OMIM).The common targets of drugs and diseases were obtained by Venny 2.1.0 platform.The Cassiae Semen-active ingredients-potential target network diagram was drawn by Cytoscape3.8.2,the protein-protein interaction(PPI)network diagram was constructed by String database,and the Cassiae Semenhyperlipidemia common targets were subjected to Gene Ontology(GO)enrichment analysis and Kyoto Encyclopedia of Genes and Genomes Academic Journal of Shanghai University of Traditional Chinese Medicine Vol.38 No.4 Jul.2024(KEGG)pathway enrichment analysis through the Database for Annotation,Visualization and Integrated Discovery(DAVID).According to the results of the network pharmacology study,aurantio-obtusin(AO)was hypothesized to be the effective acting component of Cassiae Semen in lowering blood lipids,and then the zebrafish experiments were carried out to verify based on above.The zebrafish hyperlipidemia model was induced by high-fat diet with egg yolk liquid,and was treated by AO after modeling,in order to verify the effect and mechanisms of AO in treating hyperlipidemia.Firstly,the toxicity experiment of AO on hyperlipidemic zebrafish was performed to determine the maximum administration concentration of AO.Subsequently,the effects of AO on total cholesterol(TC)and triglyceride(TG)contents and liver histomorphometry of hyperlipidemic zebrafish were observed.Quantitative real-time PCR was used to detect the expressions of core target mRNAs in hyperl

关 键 词：高脂血症 决明子 橙黄决明素 网络药理学 斑马鱼 

分 类 号：R285[医药卫生—中药学]