主 办:北 京 中 医 药 大 学
ISSN 1006-2157 CN 11-3574/R

北京中医药大学学报 ›› 2021, Vol. 44 ›› Issue (1): 60-68.doi: 10.3969/j.issn.1006-2157.2021.01.010

• 中药药理 • 上一篇    下一篇

基于网络药理学探讨肾气丸干预2型糖尿病的分子机制及关键作用通路的验证*

杨金伟1, 喻嵘2,3, 吴勇军4, 刘秀3, 邓奕辉1#   

  1. 1 湖南中医药大学中西医结合学院 湖南 410208;
    2 湖南中医药大学研究生院;
    3 中医方证研究转化医学湖南省重点实验室;
    4 湖南中医药大学药学院
  • 收稿日期:2020-07-29 出版日期:2021-01-30 发布日期:2021-01-18
  • 通讯作者: #邓奕辉,女,博士,教授,博士生导师,主要研究方向:中西医结合防治内分泌疾病,E-mail:644138330@qq.com
  • 作者简介:杨金伟,男,博士,住院医师
  • 基金资助:
    *国家自然科学基金面上项目(No.81573956),国家重点研发计划中医药现代化研究重点专项(No.2018YFC1704300),湖南省研究生科研创新项目(No.CX2018B471),湖南省“国内一流建设学科”中医学开放基金项目(No.2018ZYX44)

Using network pharmacology to study the molecular mechanism and validate the key pathway of Kidney-qi pill against Type 2 diabetes mellitus*

Yang Jinwei1, Yu Rong2,3, Wu Yongjun4, Liu Xiu3, Deng Yihui1#   

  1. 1 College of Integrated Traditional Chinese and Western Medicine,Hunan University of Chinese Medicine,Changsha,Hunan 410208,China;
    2 Graduate School,Hunan University of Chinese Medicine,Changsha,Hunan 410208,China;
    3 Key Laboratory of Colleges and Universities,Traditional Chinese Medicine Prescription and Transformation of Medical Laboratory,Changsha,Hunan 410208,China;
    4 School of pharmacy,Hunan University of Chinese Medicine,Changsha,Hunan 410208,China
  • Received:2020-07-29 Online:2021-01-30 Published:2021-01-18
  • Contact: Prof.Deng Yihui,Ph.D,Doctoral Supervisor.College of Integrated Traditional Chinese and Western Medicine,Hunan University of Chinese Medicine.No.300,Xueshi Road,Hanpu Science and Education Park,Yuelu District,Changsha,Hunan 410208.E-mail:644138330@qq.com.
  • Supported by:
    National Natural Science Foundation of China(No. 81573956),National Key Research and Development Plan of China(No.2018 YFC1704300),Graduate Research Innovation Project of Hunan province(No. CX2018B471),Open Fund for Traditional Chinese Medicine of Hunan Province(No.2018ZYX44)

摘要: 目的 基于网络药理学方法探讨肾气丸治疗2型糖尿病的活性成分、作用靶点及可能作用机制,并通过体外实验进行初步验证。方法 应用TCM database@Taiwan、Chemistry Database及TCMSP数据库获取肾气丸的化学活性成分及相关靶点信息,通过对比GenCards及OMIM 2个数据库,归纳预测肾气丸化学成分中明确对2型糖尿病具有治疗作用的靶点,进行成分-靶点相互作用网络图的构建。运用DAVID 6.8数据库对基因富集分析,获得GO功能注释结果及KEGG通路富集结果,分析通路相关基因及功能。综合药物-疾病靶基因,使用String 10.5工具构建靶点相互作用网络,通过Cytoscape软件进行拓扑分析,对肾气丸关键化学成分及靶点进行评价。验证实验以大鼠胰岛细胞瘤(INS-1)细胞作为研究对象,应用高糖诱导细胞损伤模型,肾气丸以8.64 g/kg剂量灌胃SD大鼠制备含药血浆干预细胞,CCK8法检测细胞增殖抑制率,Western blot检测关键通路蛋白的表达情况。结果 筛选得到肾气丸中58种潜在成分和198个靶点与2型糖尿病关联,成分-靶点网络图显示肾气丸可能通过AKT1、JUN、TNF与PTGS2等关键基因,经由磷脂酰肌醇3激酶(PI3K)/蛋白激酶B(AKT)信号通路、丝裂原活化蛋白激酶(MAPK)信号通路、肿瘤坏死因子(TNF)信号通路、神经配体受体交互作用和低氧诱导因子-1(HIF-1)信号通路等发挥干预2型糖尿病的作用;细胞实验中,肾气丸含药血浆可降低高糖引起的细胞增殖抑制率,上调PI3K、AKT及p-AKT蛋白表达(P<0.01),其激活PI3K/ AKT通路作用效应与通路激活剂740Y-P相当(P>0.05)。结论 肾气丸通过多成分、多靶点的作用效应干预2型糖尿病,可促进细胞线粒体能量转换及细胞信号传导过程。初步证实了肾气丸能够影响PI3K/ AKT通路相关蛋白表达,可为进一步机制研究及临床应用提供参考。

关键词: 肾气丸, 网络药理学, 2型糖尿病, 潜在靶点, 大鼠, 胰岛细胞瘤细胞

Abstract: Objective To explore the active components,core targets and mechanism of Kidney-qi Pill in treating for Type 2 diabetes mellitus (T2DM),based on network pharmacology and preliminarily verify it through in vitro experiment.Methods The TCM database@Taiwan,Chemistry Database and TCMSP database were used to filter the chemical composition and target of Kidney-qi pill.The corresponding gene name of each target was obtained in UniProt database.Then,the target genes of Kidney-qi pill and T2DM were plotted,depending on the relationship between chemical composition and target.The String software was used to construct Kidney-qi pill ingredients-targets network.The DAVID database was used to analyze the pathway enrichment of the interaction gene,and core signaling pathways were performed by using KEGG pathway analysis tool.Insulinoma cell line INS-1 cells in rats were used to establish damage model by using high concentration of glucose.SD rats were administered intragastrically with 8.64 g/kg Kidney-qi pill to prepare medicated plasma,which was then extracted for INS-1 cells experiment.The growth inhibition was tested by using CCK-8 essay.Key signal pathways were validated by using Western blot assay.Results 58 chemical composition of Kidney-qi pill were analyzed.A total of 198 human target genes related to T2DM were obtained.The network analysis indicated that Kidney-qi pill can treat T2DM by regulating AKT1,JUN,TNF and PTGS2 targets,involving phosphatidylinositol 3 kinase (PI3K)/ protein kinase B (AKT) signaling pathway,mitogen-activated protein kinase (MAPK) signaling pathway,tumor necrosis factor (TNF) signaling pathway,Neuroactive ligand-receptor interaction and hypoxia inducible factor-1(HIF-1) signaling pathway.Kidney-qi pill medicated plasma significantly promoted the proliferation of INS-1 cells,and apparently increased the expression of PI3K,AKT and p-AKT (P<0.01).There was no significant difference in Kidney-qi pill medicated plasma group and 740 Y-P group (P>0.05). Conclusion Conclusion Kidney-qi pill contains herbs of the same or different active ingredients,and this prescription can effectively treat T2DM.Based on the network pharmacology,Kidney-qi pill can promote cell signal transduction,regulate mitochondrial function and energy metabolism.The experiments have confirmed that Kidney-qi pill can affect the expression of PI3K/ AKT pathway related proteins.This research can provide theoretical basis for further experiments and clinical application.

Key words: Kidney-qi pill, network pharmacology, type 2 diabetes mellitus, potential target, rats, INS-1 cells

中图分类号: 

  • R285.5