中国畜牧兽医 ›› 2023, Vol. 50 ›› Issue (2): 764-778.doi: 10.16431/j.cnki.1671-7236.2023.02.034

• 基础兽医 • 上一篇    下一篇

基于网络药理学探讨麻黄甘草汤治疗肺水肿的作用机制

夏瑾瑾1, 余捷1, 李亚娜1, 刘佳丽1, 闫普普1, 陈晓兰2, 郭利伟1   

  1. 1. 长江大学, 荆州 434025;
    2. 江苏农牧科技职业学院, 泰州 225300
  • 收稿日期:2022-09-13 出版日期:2023-02-05 发布日期:2023-02-06
  • 通讯作者: 郭利伟 E-mail:guolw@yangtzeu.edu.cn
  • 作者简介:夏瑾瑾,E-mail:2546745817@qq.com
  • 基金资助:
    国家自然科学青年基金资助项目(31602099)

Study on the Mechanism of Mahuang Gancao Decoction in Treating Pulmonary Edema Based on Network Pharmacology

XIA Jinjin1, YU Jie1, LI Yana1, LIU Jiali1, YAN Pupu1, CHEN Xiaolan2, GUO Liwei1   

  1. 1. Yangtze University, Jingzhou 434025, China;
    2. Jiangsu Agri-Animal Husbandry Vocational College, Taizhou 225300, China
  • Received:2022-09-13 Online:2023-02-05 Published:2023-02-06

摘要: 【目的】预测麻黄甘草汤治疗肺水肿的潜在作用靶点及作用机制,并通过动物体内试验加以验证,为深入研究麻黄甘草汤的药理作用提供参考依据。【方法】通过中药系统药理学分析平台(TCMSP)数据库获取麻黄、甘草的活性成分及其作用靶点,利用GeneCards数据库收集肺水肿疾病的作用靶点,将麻黄甘草汤的成分作用靶点和肺水肿疾病靶点取交集,基于STRING数据库构建蛋白相互作用网络(PPI),使用Cytoscape 3.6软件构建成分-靶点网络图。利用DAVID数据库进行GO功能和KEGG通路富集分析。对获得的关键成分和核心靶点通过AutoDock Vina进行分子对接,用以佐证网络药理学预测结果。选用2%氯化铵构建小鼠肺水肿模型,分为空白对照组、氯化铵模型组及麻黄甘草汤治疗组;空白对照组每日上午腹腔注射200 μL生理盐水,氯化铵模型组和麻黄甘草汤治疗组腹腔注射200 μL 2%的氯化铵溶液;氯化铵模型组和空白对照组每晚灌胃200 μL生理盐水,麻黄甘草汤治疗组灌胃200 μL 1 g/mL麻黄甘草汤,连续处理5 d后采用实时荧光定量PCR检测白细胞介素-6(IL6)、诱导型一氧化氮合酶(NOS2)、丝裂原活化蛋白激酶3(MAPK3)和基质金属蛋白酶9(MMP9)基因mRNA表达量变化。【结果】麻黄潜在药物活性成分有23种,甘草潜在药物活性成分有92种,二者药物活性成分作用靶点239个;肺水肿相关作用靶点5 196个;药物、疾病共有靶点213个。其中关键靶点有RAC-α丝氨酸/苏氨酸蛋白激酶(AKT1)、细胞肿瘤抗原p53(TP53)、IL6、血管内皮生长因子A(VEGFA)、雌激素受体1(ESR1)、表皮生长因子受体(EGFR)、MAPK3、过氧化物合酶2(PTGS2)、信号转导和转录激活因子3(STAT3)、MMP9、NOS2等。GO功能富集分析得到954个条目,其中生物过程710个条目,涉及代谢过程、基因转录过程、生物调控过程、对药物刺激过程等;细胞组分92个条目,主要包括细胞膜、细胞质、细胞核、细胞器等;分子功能152个条目,主要有酶的催化、蛋白质的结合、转录调节等。KEGG富集分析170条信号通路,主要涉及肿瘤坏死因子(TNF)、PI3K-Akt、IL17等信号通路;分子对接结果显示,麻黄甘草汤的关键活性成分与疾病核心靶点具有较好的结合能。动物试验实时荧光定量PCR检测结果显示,与空白对照组相比,氯化铵模型组IL6、MAPK3和MMP9基因mRNA表达量显著或极显著上调(P<0.05;P<0.01),NOS2基因mRNA表达量极显著下调(P<0.01);与氯化铵模型组相比,麻黄甘草汤治疗组IL6、MAPK3、MMP9基因mRNA表达量显著或极显著下调(P<0.05;P<0.01),NOS2基因mRNA表达量极显著上调(P<0.01)。【结论】麻黄甘草汤可通过多成分、多靶点、多通路发挥治疗肺水肿的作用,其中的活性成分槲皮素、山奈酚、柚皮素、木犀草素等可能通过作用于IL6、MAPK3、MMP9、NOS2等靶点,进而对TNF、PI3K-AKT、IL17等信号通路产生炎症控制及免疫调节作用。

关键词: 麻黄甘草汤; 肺水肿; 网络药理学; 分子对接

Abstract: 【Objective】 The aim of this study was to predict the potential target and mechanism of Mahuang Gancao decoction in the treatment of pulmonary edema,and to verify it through animal experiments in vivo,so as to provide a reference for further research on the pharmacological effects of Mahuang Gancao decoction.【Method】 The potential active ingredients and related targets of Mahuang Gancao decoction were screened by Traditional Chinese Medicine Systems Pharmacology Database (TCMSP).The related targets of pulmonary edema were obtained from the GeneCards database.And then,the intersection of the active ingredient targets and disease targets was selected.The network diagram of ingredient-target was established with Cytoscape 3.6.GO fuction and KEGG pathway enrichment analysis were carried out in DAVID database.The active ingredients with the highest values and the key core targets were docked and verified by AutoDdock Vina to support the prediction results of network pharmacology.Finally,BALB/c mice were used for experimental verification.2% ammonium chloride was used to construct the pulmonary edema model.Mice were randomly divided into blank group,ammonium chloride model group and Mahuang Gancao decoction treatment group.The blank group was intraperitoneally injected with 200 μL normal saline every morning,and the ammonium chloride model group and the Mahuang Gancao decoction treatment group were intraperitoneally injected with 200 μL 2% ammonium chloride solution,the ammonium chloride model group and the blank group were intragastrically administered with 200 μL normal saline every night,and the Mahuang Gancao decoction treatment group was intragastrically administered with 200 μL 1 g/mL Mahuang Gancao decoction.After 5 days of continuous treatment,the mRNA expression levels of interleukin-6 (IL6),inducible nitric oxide synthase (NOS2),mitogen-activated protein kinase 3 (MAPK3) and matrix metalloproteinase 9 (MMP9) genes were detected by Real-time quantitative PCR.【Result】 There were 23 and 92 potential pharmaceutical active ingredients in ephedra and licorice,respectively,and 239 targets of the two components.5 196 targets of pulmonary edema were identified.There were 213 intersecting targets of drugs and diseases.The core targets were RAC-α serine/threonine protein kinase (AKT1),cell tumor antigen p53 (TP53), IL6,vascular endothelial growth factor A (VEGFA),estrogen receptor 1 (ESR1),epidermal growth factor receptor (EGFR),MAPK3,peroxidase synthase 2 (PTGS2),signal transducer and activator of transcription 3 (STAT3),MMP9,NOS2,etc.954 items were obtained by GO function enrichment analysis,including 710 items of biological process,involving metabolic process,gene transcription process,biological regulation process,drug stimulation process,etc.Cell component 92 items,mainly included cell membrane,cytoplasm,nucleus,organelles,etc.152 items of molecular function mainly included enzyme catalysis,protein binding,transcriptional regulation,etc.The results of the KEGG pathway enrichment analysis were involved in 170 signaling pathways,mainly related to tumor necrosis factor (TNF),PI3K-Akt,IL17 and other signaling pathways.Results of molecular docking showed that the key active ingredients had a good binding ability with the core targets.Real-time PCR results showed that compared with the blank group,the mRNA expression of IL6, MAPK3 and MMP9 genes in ammonium chloride model group were significantly or extremely significantly up-regulated (P<0.05 or P<0.01),and the mRNA expression of NOS2 gene was extremely significantly down-regulated (P<0.01).Compared with the ammonium chloride model group,the mRNA expression of IL6, MAPK3 and MMP9 genes in Mahuang Gancao decoction treatment group were significantly or extremely significantly down-regulated (P<0.05 or P<0.01),and the mRNA expression of NOS2 gene was extremely significantly up-regulated (P<0.01).【Conclusion】 Mahuang Gancao decoction could treat pulmonary edema through multiple components,multiple targets and multiple pathways.Quercetin,kaempferol,naringenin,luteolin and so on,which were active components,might exert inflammatory control and immunomodulatory effects on TNF,PI3K-Akt,IL17 signaling pathways through acting on IL6,MAPK3,MMP9,NOS2 and other targets.

Key words: Mahuang Gancao decoction; pulmonary edema; network pharmacology; molecular docking

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