六味地黄汤干预肾纤维化作用机制的网络药理学分析  

Network Pharmacological Analysis of the Mechanism of Liuwei Dihuang Decoction in Intervening Renal Fibrosis

作　　者：张海英 王晖 高海波 郭金晶 蒋志兵 潘佳俊 孟斌[1] 张熙[1] 唐群[1] ZHANG Haiying;WANG Hui;GAO Haibo;GUO Jinjing;JIANG Zhibing;PAN Jiajun;MENG Bing;ZHANG Xi;TANG Qun(Hunan University of Chinese Medicine,Changsha410208 Hunan,China)

机构地区：[1]湖南中医药大学,湖南长沙410208

出　　处：《中药新药与临床药理》2023年第3期367-375,共9页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：湖南省自然科学基金项目(2021JJ30506);长沙市自然科学基金项目(kq2014089);湖南省教育厅重点项目(21A0242);湖南中医药大学研究生科研创新项目(2021CX32)。

摘　　要：目的基于网络药理学方法及体内外实验探讨六味地黄汤(LWDHD)干预肾纤维化的潜在作用机制。方法(1)通过TCMSP平台检索筛选LWDHD的活性成分及其作用靶点;通过GeneCards、OMIM、TTD数据库检索肾纤维化疾病相关靶点;利用R语言VennDiagram包对LWDHD活性成分靶点和肾纤维化疾病相关靶点取交集,得到共同(交集)靶点,即为LWDHD治疗肾纤维化的关键靶点;运用Cytoscape 3.9.1软件构建LWDHD治疗肾纤维化的“活性成分-关键靶点”网络,分析核心活性成分;通过STRING平台构建共同靶点蛋白互作(PPI)网络;分别通过DAVID数据库、R语言软件的Pathview包对共同靶点进行GO功能及KEGG通路富集分析;采用AutoDock Vina软件对核心活性成分与核心靶点进行分子对接。(2)体内实验:将24只雄性SD大鼠随机分为4组:假手术组、模型组、LWDHD组(6.75 g·kg^(-1))及依那普利组(10 mg·kg^(-1)),每组6只。采用单侧输尿管结扎术(UUO)建立肾纤维化大鼠模型。各组以相应药物灌胃给药(10 mL·kg^(-1)),每日1次,连续7 d。采用HE、MASSON染色法观察大鼠肾脏组织病理变化和胶原沉积情况。(3)体外实验:将体外培养的HK-2细胞分为正常对照组(10%空白血清)、模型组(10%空白血清+200μmol·L^(-1)CoCl_(2))、LWDHD组(10%含药血清+200μmol·L^(-1)CoCl_(2));采用200μmol·L^(-1)CoCl_(2)模拟肾内缺氧环境,同时加入空白或含药血清,培养24 h后收集细胞;采用Western Blot法检测HK-2细胞HIF-1α蛋白表达水平。结果(1)共得到LWDHD活性成分40个,活性成分作用靶点194个,肾纤维化疾病相关靶点5135个,共同(交集)靶点即LWDHD治疗肾纤维化的关键靶点159个。筛选核心活性成分主要有槲皮素、豆甾醇、山柰酚、β-谷甾醇、薯蓣皂苷元等,核心靶点有热休克蛋白90α家族A类成员1(HSP90AA1)、c-Jun氨基末端激酶(JUN)、蛋白激酶B(AKT1)及缺氧诱导因子1α(HIF-1α)等。GO功能主要与细胞因子介导信号通�Objective To explore the potential mechanism of Liuwei Dihuang Decoction (LWDHD)in intervening renal fibrosis based on a network pharmacological approach and in vitro and in vivo experiments.Methods (1)To search and screen the active ingredients of LWDHD and their targets through the TCMSP platform;to screen the targets related to renal fibrosis disease through GeneCards,OMIM and TTD databases;R language VennDiagram package was used to intersect the targets of active ingredients of LWDHD and the targets related to renal fibrosis disease to obtain the common (intersecting) targets,which are LWDHD key targets for the treatment of renal fibrosis;Cytoscape 3.9.1 software was used to construct the"active ingredients-key targets"network for the treatment of renal fibrosis in LWDHD and analyze the core active ingredients;STRING platform was used to construct the common target protein-protein interaction (PPI) network;DAVID database,the R language software,and the R language software were used to analyze the core active ingredients respectively.The DAVID database and Pathview package of R language software were used for GO function and KEGG pathway enrichment analysis of the common targets.AutoDock vina software was used to perform molecular docking between core active ingredients and core targets.(2)In vivo experiments:24 male SD rats were randomly divided into 4 groups:shamoperation group,model group,LWDHD group (6.75 g·kg^(-1))and Enalapril group (10 mg·kg^(-1)),6 rats in each group.A rat model of renal fibrosis was established using unilateral ureteral ligation (UUO).The rats were administered by gavage (10 mL·kg^(-1))once daily for 7 consecutive days.The renal pathological changes and collagen deposition were observed by HE and MASSON staining.(3)In vitro experiments:HK-2 cells cultured in vitro were divided into normal control group (10%blank serum),model group (10%blank serum+200μmol·L^(-1)CoCl_(2))and LWDHD group (10%drug-containing serum+200μmol·L^(-1)CoCl_(2));200μmol·L^(-1)CoCl_(2)was used to simulate

关 键 词：六味地黄汤 肾纤维化 网络药理学 体内外实验 槲皮素 豆甾醇 薯蓣皂苷元 HIF-1信号通路 大鼠 HK-2细胞 

分 类 号：R285.5[医药卫生—中药学] R857.3[医药卫生—中医学]