基于网络药理学和体外实验验证探讨吲哚-3-甲醇治疗肥胖的作用机制

Mechanism of Indole-3-Methanol in Treatment of Obesity Based on Network Pharmacology and in Vitro Experiment

目的:根据分子对接技术与网络药理学初步预测出吲哚-3-甲醇作用的靶点和相关信号通路,再通过体外实验来验证网络药理学与分子对接技术得出的结果。方法:利用SwissTargetPrediction成分靶点预测数据库及查阅文献获得吲哚-3-甲醇的药理作用相关的靶点;利用在线人类孟德尔遗传病数据库(OMIM)、基因数据库(GeneCards)、毒性与基因比较数据库(CTD)取得与肥胖相关的靶点;在通过STRING数据库得到吲哚-3-甲醇-肥胖靶点蛋白质-蛋白质相互作用(PPI)网络图;通过Cytoscape 3.8.2软件进行筛选和分析;利用DAVID 6.8数据库对药物-疾病交集靶点进行基因本体(GO)功能注释和京都基因与基因组百科全书(KEGG)通路富集分析。通过AutoDock vina 1.1.2软件对吲哚-3-甲醇和疾病靶点进行分子对接;最后通过体外实验验证了药物抗肥胖的作用。结果:预测结果得出吲哚-3-甲醇与肥胖共同靶点有80个,包括基质金属蛋白酶(MMP)-9、Janus激酶(JAK)2等;KEGG富集分析结果显示吲哚-3-甲醇治疗肥胖主要作用于肿瘤坏死因子(TNF)信号通路、血管内皮生长因子(VEGR)信号通路、酪氨酸激酶受体2(ErbB2)、VEGF等信号通路;分子对接显示吲哚-3-甲醇与肥胖相关蛋白(FTO)受体表现出良好的对接活性;体外蛋白免疫印迹、噻唑蓝(MTT)和油红O染色结果表明,不同剂量的吲哚-3-甲醇能有效抑制3T3-L1细胞内FTO蛋白的表达(P<0.05)。结论:吲哚-3-甲醇治疗肥胖的作用机制可能与抑制FTO蛋白表达有关,进而与抑制脂肪细胞增殖能力有关。该研究为吲哚-3-甲醇抗肥胖提供了基本实验依据。

Objective:To preliminarily predict the targets and signaling pathways of indole-3-methanol in the treatment of obesity based on molecular docking technology and network pharmacology,and then verify the prediction results by the experiment in vitro.Method:The pharmacological targets of indole-3-methanol were obtained from SwissTargetPrediction and literature review.Obesity-related targets were obtained from Online Mendelian Inheritance in Man(OMIM),GeneCards,and Comparative Toxicogenomics Database(CTD).The protein-protein interaction network of the targets of indole-3-methanol and obesity was built by STRING.Cytoscape 3.8.2 was used for target screening.Gene ontology(GO)functional annotation and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment were performed for the common targets shared by obesity and indole-3-methanol in DAVID 6.8.AutoDock Vina 1.1.2 was employed to perform the molecular docking between indole-3-methanol and disease targets.Finally,the in vitro experiment was carried out to verify the anti-obesity effect of indole-3-methanol.Result:Indole-3-methanol and obesity shared 80common targets,which included matrix metalloproteinase(MMP)-9,Janus kinase(JAK)2,etc.KEGG enrichment predicted that indole-3-methanol mainly acted on tumor necrosis factor(TNF),vascular endothelial growth factor(VEGF),tyrosine kinase receptor 2(ErbB2),and epidermal growth factor receptor(EGFR)signaling pathways in the treatment of obesity.Molecular docking showed that indole-3-methanol had good binding activity with fat mass and obesity-associated protein(FTO).The results of Western blot,MTT assay,and oil-red O staining showed that indole-3-methanol down-regulated the expression of FTO in 3T3-L1 cells(P<0.05).Conclusion:Indole-3-methanol may treat obesity by down-regulating the expression of FTO protein and further inhibiting adipocyte proliferation.This study provides an experimental basis for deciphering the antiobesity mechanism of indole-3-methanol.