基于网络药理学和动物实验探究解毒复正方防治肝癌的作用机制

Investigating the Mechanism of JDFZ in Preventing and Treating Hepatocellular Carcinoma Based on Network Pharmacology and Animal Experiments

关键词:  解毒复正方  网络药理  肝癌  EGFR/MAPK

Key words:  jiedu fuzheng formula (JDFZ)  network pharmacology  hepatocellular carcinoma (HCC)  EGFR/MAPK signaling pathway

摘要:

目的 基于网络药理学与动物实验相结合的方法,探讨解毒复正方(JDFZ)抗肝癌的作用机制。方法 通过TCMSP、HERB数据库筛选获得JDFZ潜在作用靶点,整合GeneCards、OMIM及DrugBank数据库获取肝癌相关靶点,筛选两者交集靶点构建蛋白质互作网络(PPI)。将交集靶点导入STRING数据库,去除游离的蛋白点,并将“PPI score>0.9”的靶点信息导入Cytoscape 3.10.1软件运用Network Analyzer功能对导入数据进行拓扑结构分析,分析筛选核心靶点后进行基因本体富集(GO)和基因组百科全书(KEGG)通路分析,并构建“药物-成分-靶点-疾病”网络图(C-T)。进一步构建H22荷瘤小鼠模型进行实验验证。 结果 通过TCMSP、HERB数据库筛选获得JDFZ潜在作用靶点342个(经去重处理),整合GeneCards、OMIM及DrugBank数据库获取肝癌相关靶点18708个。筛选两者交集靶点共314个,构建蛋白质互作网络(PPI)识别出TP53、JUN、SRC、AKT1、MAPK3、EGFR等22个关键核心靶点,槲皮素、木犀草素β-谷甾醇等是JDFZ核心成分；GO功能富集分析显示333个显著条目,包括生物过程(BP)264项、细胞组分(CC)25项、分子功能(MF)44项；KEGG通路富集分析揭示163条相关信号通路,主要涉及MAPK信号通路、细胞凋亡调控、TNF信号通路等。JDFZ可显著抑制肿瘤生长,瘤重显著降低(P<0.05),病理显示肿瘤细胞结构明显破坏；同时其显著降低血清AST、ALT水平及AST/ALT比值(P<0.01,P<0.05)；Western blot检测显示肿瘤组织中p-EGFR/EGFR和p-ERK/ERK蛋白表达水平显著下调(P<0.01)。结论 本研究首次揭示JDFZ可能通过调控EGFR/ERK信号通路轴,抑制肝癌细胞增殖并减轻肝损伤,为JDFZ治疗肝癌提供了药理学依据。

Abstract:

ABSTRACT: OBJECTIVE Based on the method combining network pharmacology and with animal experiments, the mechanism of action of Jiedu Fuzheng Formula (JDFZ) in anti-liver cancer was explored. METHODS Potential therapeutic targets of JDFZ were retrieved from the TCMSP and HERB databases, HCC-related targets were collected from GeneCards, OMIM, and DrugBank. The intersection targets of the two were screened to build a protein interaction network (PPI), the intersection targets were imported into the string database to remove free protein spots, and the target information of "PPI score>0.9" was imported into Cytoscape 3.10.1 software. The network analyzer function was used to analyze the topology structure of the imported data. After the core targets were analyzed and screened, gene ontology enrichment (go) and genome Encyclopedia (KEGG) pathway analysis were carried out, and construct a "Drug Ingredient Target Disease" network diagram (C-T). After the relevant data were exported, the mapping analysis was carried out by using the visualization platform. Further construct an H22 tumor bearing mouse model for experimental validation. RESULTS Potential therapeutic targets of JDFZ were retrieved from the TCMSP and HERB databases, yielding 342 unique targets. HCC-related targets (18,708) were collected from GeneCards, OMIM, and DrugBank. Intersection analysis identified 314 overlapping targets, and protein-protein interaction (PPI) network analysis revealed 22 core targets, including TP53, JUN, SRC, AKT1, MAPK3, and EGFR. Quercetin, luteolin and β-sitosterol are the core components of JDFZ. Gene Ontology (GO) enrichment analysis identified 333 significant terms, encompassing 264 biological processes (BP), 25 cellular components (CC), and 44 molecular functions (MF). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis highlighted 163 pathways, primarily involving the MAPK signaling pathway, apoptosis regulation, and TNF signaling pathway. Experimental validation using an H22 hepatocellular carcinoma xenograft mouse model demonstrated that JDFZ significantly inhibited tumor growth (reduced tumor weight, P < 0.05) and disrupted tumor cell architecture. Additionally, JDFZ markedly decreased serum AST and ALT levels and the AST/ALT ratio (P < 0.01 and P < 0.05, respectively). Western blot analysis revealed significant downregulation of p-EGFR/EGFR and p-ERK/ERK protein expression in tumor tissues (P < 0.01). CONCLUSION This study first demonstrates that JDFZ exerts anti-HCC effects by modulating the EGFR/ERK signaling pathway axis, inhibiting tumor proliferation, and alleviating liver injury, providing a pharmacological foundation for the clinical application of JDFZ in HCC treatment.