基于网络药理学、分子对接和实验验证探讨加味真武汤延缓慢性肾功能衰竭进展的作用机制  

Mechanism of Modified Zhenwutang in Delaying Progression of Chronic Renal Failure Based on Network Pharmacology,Molecular Docking,and Experimental Verification

作　　者：张圆圆 靳培培 李宛哲 郭登洲[2] ZHANG Yuanyuan;JIN Peipei;LI Wanzhe;GUO Dengzhou(Graduate School,Hebei University of Chinese Medicine,Shijiazhuang 050091,China;The First Affiliated Hospital of Hebei University of Chinese Medicine/Hebei Provincial Hospital of Traditional Chinese Medicine,Shijiazhuang 050011,China)

机构地区：[1]河北中医药大学研究生学院,石家庄050091 [2]河北中医药大学第一附属医院/河北省中医院,石家庄050011

出　　处：《中国实验方剂学杂志》2023年第16期123-133,共11页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：河北省中医药管理局科研计划项目(2023021);河北省自然科学基金项目(H2022423367);河北中医学院博士研究生创新项目(XCXZZBS2023009)。

摘　　要：目的:基于网络药理学初步预测加味真武汤治疗慢性肾功能衰竭(CRF)的活性成分、作用靶点及信号通路,通过分子对接和动物实验验证探讨其延缓疾病进展的可能作用机制。方法:通过本草组鉴(HERB)数据库获取加味真武汤的有效成分及靶点;通过基因数据库(GeneCards)获取CRF疾病靶点;通过Venny 2.1软件获得交集靶点基因,结合STRING数据库绘制蛋白质-蛋白质相互作用网络,并通过Cytoscape 3.9.1软件筛选加味真武汤治疗CRF的核心作用靶点;使用Metascape数据库对交集基因进行基因本体(GO)分析和京都基因与基因组百科全书(KEGG)富集分析。选取关键作用靶点与药物活性成分通过AutoDockTools 1.5.6软件进行分子对接验证;采用腺嘌呤灌胃12周的方法建立实验性CRF大鼠模型,给予加味真武汤和盐酸贝那普利干预4周。给药结束后处死大鼠,通过免疫组织化学(IHC)、免疫荧光(IF)、实时荧光定量聚合酶链式反应(Realtime PCR)、蛋白免疫印迹(Western blot)法检测大鼠肾组织中脯氨酸羟化酶1(PHD1)、脯氨酸羟化酶2(PHD2)、低氧诱导因子-1α(HIF-1α),α-平滑肌肌动蛋白(α-SMA)表达水平进行动物实验验证。结果:通过HERB数据库获得加味真武汤药物靶点基因426个;通过GeneCards获取CRF相关靶点基因2698个;药物疾病交集基因154个;核心靶点8个,其中白蛋白(ALB)、蛋白激酶B1(Akt1)、肿瘤坏死因子(TNF)、白细胞介素-6(IL-6)、胰岛素(INS)、血管内皮生长因子A(VEGFA)、肿瘤蛋白p53(TP53)、白细胞介素-1β(IL-1β)等靶点可能与加味真武汤治疗CRF密切相关;KEGG富集分析预测加味真武汤治疗CRF的机制主要涉及脂质与动脉粥样硬化、HIF-1信号通路、细胞凋亡、核转录因子-κB(NF-κB)信号通路等通路。分子对接结果,加味真武汤的成分与核心作用靶点中的ALB、Akt1、TNF、IL-6、INS、VEGFA、TP53、IL-1β具有较为稳定的结合活性,其可能通过影响ALBObjective:To preliminarily predict the active ingredients,targets,and signaling pathways of modified Zhenwutang in the treatment of chronic renal failure(CRF)based on network pharmacology and explore its potential mechanism for delaying disease progression through molecular docking and animal experiments.Method:The effective ingredients and targets of modified Zhenwutang were obtained from the HERB database.The targets related to CRF were obtained from the GeneCards.The intersection target genes were obtained using Venny 2.1 software and a protein-protein interaction(PPI)network was constructed using the STRING.The core targets for treating CRF with modified Zhenwutang were screened using Cytoscape 3.9.1 software.The intersection genes were analyzed using Metascape database for gene ontology(GO)analysis and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment analysis.Molecular docking validation was performed using AutoDockTools 1.5.6 software for the key targets and active ingredients.An experimental CRF model was established in rats by administering adenine via gavage for 12 weeks,followed by intervention with modified Zhenwutang and benazepril hydrochloride for four weeks.After treatment,the rats were euthanized,and immunohistochemistry(IHC),immunofluorescence(IF),real-time quantitative polymerase chain reaction(Real-time PCR),and western blot were performed to detect the expression levels of prolyl hydroxylase domaincontaining proteins 1(PHD1),prolyl hydroxylase domain-containing proteins 2(PHD2),hypoxia-inducible factor-1α(HIF-1α),andα-smooth muscle actin(α-SMA)in the renal tissues of the rats.Result:A total of 426 drug target genes of modified Zhenwutang were obtained from the HERB database.A total of 2698 target genes related to CRF were obtained from the GeneCards database.There were 154 intersection genes between the drug and the disease.Eight core targets were identified,including albumin(ALB),protein kinase B1(Akt1),tumor necrosis factor(TNF),interleukin-6(IL-6),insulin(INS),vascular endotheli

关 键 词：加味真武汤 CRF 网络药理学 分子对接 动物实验 

分 类 号：R284[医药卫生—中药学] R285[医药卫生—中医学] R289R287R22R2-031R33R24