| 易振,刘金萍,吴婷,等.基于生物信息学、网络药理学和分子对接探究甘草附子汤治疗强直性脊柱炎的机制[J].安徽医药,2024,28(12):2359-2367. |
| 基于生物信息学、网络药理学和分子对接探究甘草附子汤治疗强直性脊柱炎的机制 |
| Based on bioinformatics, network pharmacology and molecular docking, the mechanism of Gancao Fuzi decoction in the treatment of ankylosing spondylitis were explored |
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| DOI：10.3969/j.issn.1009-6469.2024.12.006 |
| 中文关键词: 脊柱炎，强直性 甘草 附子 生物信息学 白细胞介素 -10 芳基烃受体 免疫相关基因 网络药理学 分子对接 |
| 英文关键词: Spondylitis, ankylosing Iquorice root Prepared common monkshood branched root Bioinformatics Interleukin-10 Aryl hydrocarbon receptor Immune-related genes Network pharmacology Molecular docking |
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| 中文摘要: |
| 目的基于生物信息学、网络药理学和分子对接探究甘草附子汤治疗强直性脊柱炎( AS)的机制。方法从 GEO数据库检索 2023年 7月以前与 AS相关的基因表达谱数据获得 GSE25101、GSE73754基因芯片，在 R语言中对 2个芯片进行整合，利用 limma包筛选差异表达基因( DEGs)同时在 GenegCards、Msigdb、IMMPort 3个数据库中下载免疫相关基因并取交集。基于 DEGs与 3个数据库免疫相关基因的交集，取交集，再筛选出相关免疫 -差异表达基因( IDEG)的相关矩阵；通过构建风险模型判断 IDEG的诊断效果，选择 LASSO回归模型识别 AS预测的特征基因，并绘制校准曲线和决策曲线， Cox比例风险回归模型分析结果使用森林图展现；采用受试者操作特征曲线( ROC曲线)验证风险模型和特征基因的表达；将 DEGs和 IDEG导入 String数据库得到蛋白质 -蛋白质相互作用( PPI)，使用 Cytoscape软件进行分析并进行可视化；利用 clusterProfiler包对 IDEG进行基因本体( GO)和京都基因和基因组数据库( KEGG)富集分析，筛选出特征基因。然后，利用 CIBERSORT反卷积法分析 16个基因与 22种免疫细胞的相关性；基于 TCMSP、Uniprot数据库中搜索甘草附子汤的中药活性成分及其药物靶点，利用 DisGeNET、Gene-Cards数据库检索疾病靶点，取中药靶点及疾病靶点交集后导入 String数据库，使用 Cytoscape软件构建疾病基因与药物共有靶点及靶点蛋白互作网络，最终，使用 Autodock、Pymol进行分子对接并可视化。结果 GSE25101和 GSE73754并集后共得到 190个 DEGs，筛选出 16个 IDEG:CXCL8、HLA-DQA1、MMP9、TGFBR3、S100A12、PRF1、KIR2DL4、CARD11、HLA-C、ADM、ADRB2、 CD81、IL2RB、APOBEC3A、KIR2DL3、CXCR1；在风险模型中，使用 LASSO算法从 16个基因中识别出 9个最具影响力的特征基因系数，校准曲线显示，线图模型预测与理想模型预测几乎相同，并且决策曲线分析或复合遗传模型中的单一预测风险评分优于随机模型中的单一预测风险评分， 9个特征基因总 ROC曲线下面积( AUC)值为 0.91，单个特征基因 AUC值均大于 0.60；将其得到的蛋白 ID导入 PPI得到其 9个潜在靶点: CXCL8、HLA-DQA1、MMP9、TGFBR3、CARD11、HLA-C、ADRB2、CD81、IL2RB。富集分析显示，分子功能(MF2)中的富集最突出的项目是免疫受体活性、细胞因子结合、主要组织相容性复合物( MHC)Ⅱ类蛋白复合物结合等；细胞组分( CC)中的富集最突出的项目是 MHC蛋白复合体、内质网膜管腔侧的整体成分、内质网膜的腔侧等；生物过程( BP)中的富集最突出的项目是受体介导的内吞作用、受体内在化、 G蛋白偶联受体信号通路的负调控、白细胞移动等。免疫浸润结果显示， AS病人的巨噬细胞 M2、静息态树突状细胞、记忆 B细胞、 T细胞 CD4记忆激活、 T细胞滤泡辅助性 T细胞均低于对照组病人； AS病人的中性粒细胞、单核细胞、嗜酸细胞( EOS)、激活的树突状细胞均高于对照组病人；基于甘草附子汤，在 TCMSP中共获得 342个活性成分，两个数据库取交集后得到 43个疾病靶点，与甘草附子汤取交集靶点后得到共同靶点 5个，即酪氨酸蛋白磷酸酶非受体 22型( PTPN22)、嘌呤霉素敏感的氨基肽酶( NPEPPS)、一氧化氮合酶 2(NOS2)、白细胞介素 -10(IL-10)、芳基烃受体( AHR)，将 5个共同靶点导入 Cytoscape得到 2个核心靶点，即 IL-10、AHR，同时活性成分丁香酚与 IL-10、AHR对接结合能分别为 .17.70 kJ/mol、.17.03 kJ/mol，胱氨酸与 IL-10、AHR对接结合能分别为 .11.67 kJ/mol、.13.47 kJ/ mol。结论 IL-10、AHR是参与甘草附子汤调节 AS的关键靶点，同时甘草附子汤通过多成分、多靶点治疗 AS。 |
| 英文摘要: |
| Objective To explore the mechanism of Gancao Fuzi decoction in the treatment of ankylosing spondylitis(AS)on the basis of bioinformatics, network pharmacology and molecular docking.Methods The GSE25101 and GSE73754 gene chips were obtainedfrom GEO database by searching gene expression profiling data related to AS before July 2023, the 2 chips were integrated in R lan-guage, and differentially expressed genes (DEGs) were screened by using the limma package, and meanwhile, immune-related geneswere downloaded from 3 databases, namely GenegCards, Msigdb and IMMPort, and the intersection was taken. Based on the intersec-tion of DEGs and immune-related genes in three databases, the correlation matrix of related immune-differentially expressed genes(IDEG) was screened. The diagnostic effect of IDEG was judged by constructing a risk model. The LASSO regression model was select-ed to identify the characteristic genes predicted by AS, and the calibration curve and decision curve were drawn. The Cox regressionanalysis results were shown by forest map. The receiver operating characteristic (ROC) curve was used to verify the risk model and theexpression of characteristic genes. DEGs and IDEG were imported into String database to obtain protein-protein interaction (PPI),which was analyzed and visualized by Cytoscape software. The clusterProfiler package was used to perform gene ontology (GO) and Kyo-to encyclopedia of genes and genomes (KEGG) enrichment analysis on IDEG to screen out characteristic genes. Then, the correlationbetween 16 genes and 22 immune cells was analyzed by CIBERSORT deconvolution method. Based on TCMSP and Uniprot database,the active components and drug targets of Gancao Fuzi decoction were searched. The disease targets were searched in DisGeNET andGeneCards databases. The intersection of traditional Chinese medicine targets and disease targets was imported into String database.Cytoscape software was used to construct the common targets and target protein interaction network of disease genes and drugs. Finally,Autodock and Pymol were used for molecular docking and visualization.Results A total of 190 DEGs were obtained after combiningGSE25101 and GSE73754, and 16 IDEGs were screened out, namely, CXCL8, HLA-DQA1, MMP9, TGFBR3, S100A12, PRF1, KIR2DL4, CARD11, HLA-C, ADM, ADRB2, CD81, IL2RB, APOBEC3A, KIR2DL3, and CXCR1; in the risk model, the LASSO algo-rithm was used to identify the 9 most influential characteristic gene coefficients from 16 genes. The calibration curve showed that theprediction of the line graph model was almost the same as that of the ideal model, and the single prediction risk score in the decisioncurve analysis or the composite genetic model was better than that in the random model. The total ROC curve AUC value of the 9 char-acteristic genes was 0.91, and the AUC value of the single characteristic gene was greater than 0.60. The obtained protein ID was intro-duced into PPI to obtain nine potential targets, namely, CXCL8, HLA-DQA1, MMP9, TGFBR3, CARD11, HLA-C, ADRB2, CD81, andIL2RB. Enrichment analysis results showed that the most prominent items in molecular function (MF2) were immune receptor activity,cytokine binding, Major Histocompatibility Complex (MHC) class Ⅱ protein complex binding, etc; the most prominent items in the en-richment of cell component (CC) were MHC protein complex, the whole component of the lumen side of the endoplasmic reticulummembrane, the lumen side of the endoplasmic reticulum membrane, etc; the most prominent enrichment items in biological processes(BP) were receptor-mediated endocytosis, receptor internalization, negative regulation of G protein-coupled receptor signaling pathway,leukocyte migration, etc. The results of immune infiltration showed that macrophage M2, resting dendritic cells, memory B cells, CD4memory activation of T cells and T follicular helper T cells in AS patients were less than those in the control group; neutrophils, mono-cytes, eosinophils (EOS) and activated dendritic cells in AS patients were more than those in the control group. Based on Gancao Fuzi decoction, a total of 342 active components were obtained in TCMSP. After the intersection of the two databases, 43 disease targetswere obtained. After their intersection with Gancao Fuzi decoction, five common targets were obtained, namely PTPN22, NPEPPS, NOS2, IL-10 and AHR. Then the five common targets were introduced into Cytoscape to obtain two core targets, namely IL-10 and AHR. Meanwhile, the binding energy of docking of the active component eugenol with IL-10 and AHR was .17.70 kJ/mol and .17.03 kJ/mol, respectively. The binding energy of docking of cystine with IL-10 and AHR was .11.67 kJ/mol and .13.47 kJ/mol, respectively. Conclusion IL-10 and AHR are the key targets involved in the regulation of AS by Gancao Fuzi decoction, while Gancao Fuzi decoc-tion treats AS through multi-component and multi-target. |
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