良附丸治疗胃溃疡的网络药理学作用机制研究

目的:研究良附丸治疗胃溃疡的作用机制。方法:基于网络药理学,通过检索中药系统药理学技术平台(TCMSP)、Gene-Cards、OMIM和DisGeNET等数据库,筛选良附丸的活性成分、靶点和胃溃疡相关靶点,并利用Venny 2.1软件筛选良附丸活性成分与胃溃疡的共同靶点;通过STRING数据库构建良附丸治疗胃溃疡的蛋白互作网络图,并利用Cytoscape 3.7.2软件对该网络进行拓扑分析;通过DAVID数据库对良附丸的活性成分与胃溃疡的共同靶点进行基因本体(GO)功能和京都基因与基因组百科全书(KEGG)通路富集分析,并构建良附丸活性成分-胃溃疡靶点-KEGG通路网络图。在动物实验验证中,取小鼠随机分为空白组、模型组、雷尼替丁组(阳性对照,39 mg/kg)和良附丸低、中、高剂量组(0.78、1.56、3.12 g/kg),分别灌胃相应药物,每天1次,连续7天。末次给药后,除空白组外,其余各组小鼠灌胃无水乙醇(0.01 mL/g),以复制胃溃疡模型。造模成功后,取小鼠胃组织观察病变情况,并计算胃溃疡指数;检测小鼠胃组织中TP53、c-Jun、p38丝裂原活化蛋白激酶(p38 MAPK)、蛋白激酶B(Akt)、磷酸化Akt(p-Akt)和肿瘤坏死因子α(TNF-α)蛋白的表达水平。结果:共筛选出良附丸中的9个活性成分(如异鼠李素、β-谷甾醇、山柰酚等)和166个共同靶点。良附丸可通过细胞对化学刺激的反应、蛋白质结合、细胞外空间等GO功能,以及癌症信号通路、乙型肝炎、膀胱癌、胰腺癌、TNF信号通路、前列腺癌等KEGG通路发挥治疗胃溃疡作用。动物实验结果显示,与模型组比较,良附丸能显著降低小鼠胃组织病变情况和胃溃疡指数,以及TP53、c-Jun、p38 MAPK、Akt、p-Akt和TNF-α蛋白的表达水平(P<0.05或P<0.01)。结论:筛选出了良附丸治疗胃溃疡的9个活性成分和166个靶点;良附丸可通过抑制MAPK、NF-κB、TNF和PI3K/Akt等多条通路等,抑制炎症因子的表达,从而发挥改善胃溃疡的作用。

Study on Network Pharmacology Mechanism of Liangfu Pills in the Treatment of Gastric Ulcer

OBJECTIVE:To study the mechanism of Liangfu pills in the treatment of gastric ulcer.METHODS:Based on network pharmacology,the active ingredients,targets and gastric ulcer related targets of Liangfu pill were screened by searching TCMSP,GeneCards,OMIM and DisGeNET databases,and the common targets of Liangfu pill and gastric ulcer were screened by Venny 2.1 software.The protein interaction network of Liangfu pill in the treatment of gastric ulcer was constructed by STRING database.Cytoscape 3.7.2 software was used to construct topological analysis of the network.The function of GO and the enrichment of KEGG pathway were analyzed by DAVID database.Active component-gastric ulcer target-KEGG pathway network was constructed for Liangfu pill.In animal experiment,mice were randomly divided into blank group,model group,ranitidine group(positive control,39 mg/kg),Liangfu pill low-,medium-and high-dose groups(0.78,1.56,3.12 g/kg),and were given corresponding drugs intragastrically,once a day,for consecutive 7 days.After last administration,except for blank group,the other groups were given absolute ethanol(0.01 mL/g)intragastrically to induce the gastric ulcer model.After modeling,the pathological changes of gastric tissue were observed and the gastric ulcer index was calculated;the expression of TP53,c-Jun,p38 MAPK,Akt,p-Akt and TNF-αprotein in gastric tissues of mice were detected.RESULTS:A total of 9 active ingredients(i.g.isorhamnetin,β-gsterol,kaempferol)and 166 common targets were screened in Liangfu pills.Liangfu pill can treat gastric ulcer through GO functions such as cell response to chemical stimulation,protein binding and extracellular space,and KEGG pathways such as cancer signaling pathway,hepatitis B,bladder cancer,pancreatic cancer,TNF signaling pathway and prostate.The results of animal experiments showed that compared with model group,Liangfu pills could significantly reduce the pathological changes of gastric tissue and gastric ulcer index,as well as the protein expression of TP53,c-Jun,p38 MAPK,Akt,p-Akt and TNF-α(P<0.05 or P<0.01).CONCLUSIONS:The 9 active components and 166 targets of Liangfu pill were screened out.Liangfu pill can inhibit the expression of inflammatory factors by inhibiting MAPK,NF-κB,TNF and PI3K/Akt signaling pathway,and thus play an important role in improving gastric ulcer.