微量元素与健康研究

目的 以疾病状态转录组为生物学约束，整合网络药理学、蛋白质-蛋白质相互作用(protein-protein interaction, PPI)拓扑、GSE158163儿童全血RNA测序证据及分子对接，识别玄参干预A组链球菌(group A Streptococcus, GAS)相关急性咽炎的疾病相关候选靶点、主要炎症免疫环节及代表性成分-靶点关系。方法 从TCMSP、SymMap、HERB及相关文献整理玄参成分，采用SwissTargetPrediction预测人源靶点；从GeneCards、DisGeNET和OMIM检索急性咽炎相关靶点，经标准化、去重和交集分析后构建成分-靶点-疾病网络及高置信度STRING PPI网络，并进行GO和KEGG富集。GSE158163原始count矩阵含37个样本，经CPM归一化、低表达过滤和Gene Symbol整理后保留15 047个基因；以急性GAS咽炎与无GAS证据无症状对照为主分析，以病毒阴性无症状对照为严格对照，并以GAS携带者比较辅助区分活动性感染与携带状态。探索性差异分析采用log₂(CPM+0.5)转换、Welch t检验和Benjamini-Hochberg校正，以FDR<0.05且|log_2FC|>0.5为阈值；15个优先靶点进一步以limma-voom和DESeq2基于原始count矩阵进行稳健性复核，综合PPI拓扑、疾病表达及生物学相关性进行证据分级，并对6个代表性成分与6个受体进行分子对接。结果 共获得玄参候选成分115个、作用靶点693个，与961个急性咽炎相关靶点交集得到163个候选作用靶点；成分-靶点-疾病网络中108个成分形成有效连接。STRING高置信度PPI网络含58个节点、99条边。主分析检出6 441个差异表达基因，其中上调1 007个、下调5 434个。整合证据确定15个一级优先靶点：ESR1、STAT3、JAK2、KDM1A、LCK、ERBB2、JUN、RARA、APEX1、ABL1、CASP1、IL1B、TLR2、DNMT1和FGFR1;其表达方向在limma-voom与DESeq2复核中保持一致。36组成分-受体组合的Vina结合能为-9.85～-5.38 kcal/mol,其中Acteoside-JAK2、Acteoside-ERBB2和Acteoside-TLR2评分较低，Buergerinin C-CASP1/ERBB2及p-Methoxycinnamic acid-ESR1等亦呈现代表性结合模式。结论 玄参潜在作用主要集中于GAS感染后的宿主炎症免疫反应，涉及TLR2相关病原识别、CASP1/IL-1β炎症效应、JAK2/STAT3相关细胞因子应答以及炎症相关氧化应激和免疫调节。Acteoside、Ferulic acid、p-Methoxycinnamic acid、Buergerinin C等代表性成分对多个疾病相关靶点呈交叉结构覆盖，提示玄参的潜在药效物质基础更符合多种成分共同参与、协同作用于同一疾病网络的特点。

Objective To identify disease-relevant candidate targets, major inflammatory and immune processes, and representative compound-target relationships underlying the potential effects of Scrophulariae Radix on group A Streptococcus(GAS)-associated acute pharyngitis by integrating network pharmacology, protein-protein interaction(PPI) network topology, whole-blood RNA-sequencing data from children in the GSE158163 dataset, and molecular docking, with disease-state transcriptomic profiles serving as biological constraints.Methods The chemical constituents of Scrophulariae Radix were compiled from TCMSP, SymMap, HERB, and relevant literature, and their human protein targets were predicted using SwissTargetPrediction. Acute pharyngitis-related targets were retrieved from GeneCards, DisGeNET, and OMIM. Following target standardization, deduplication, and intersection analysis, a compound-target-disease network and a high-confidence STRING PPI network were constructed, followed by Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analyses. The raw count matrix of GSE158163 comprised 37 samples. After counts-per-million(CPM) normalization, low-expression filtering, and Gene Symbol annotation, 15,047 genes were retained. Acute GAS pharyngitis versus asymptomatic controls without evidence of GAS infection was used as the primary comparison, while virus-negative asymptomatic controls served as a stringent control. Comparisons with GAS carriers were additionally used to help distinguish active infection from the carrier state. Exploratory differential expression analysis was performed using log₂(CPM+0.5) transformation, Welch's t-test, and Benjamini-Hochberg correction, with FDR<0.05 and |log_2FC|>0.5 as the significance thresholds. The robustness of differential expression for 15 prioritized targets was further assessed using limma-voom and DESeq2 based on the raw count matrix. Evidence was graded by integrating PPI topology, disease-associated expression patterns, and biological relevance. Finally, six representative compounds were docked against six selected receptor proteins.Results A total of 115 candidate constituents and 693 potential targets of Scrophulariae Radix were identified. Intersection with 961 acute pharyngitis-related targets yielded 163 candidate therapeutic targets, and 108 constituents formed valid connections in the compound-target-disease network. The high-confidence STRING PPI network comprised 58 nodes and 99 edges. The primary transcriptomic analysis identified 6,441 differentially expressed genes, including 1,007 upregulated and 5,434 downregulated genes. Integrative evidence analysis identified 15 Tier 1 priority targets: ESR1, STAT3, JAK2, KDM1A, LCK, ERBB2, JUN, RARA, APEX1, ABL1, CASP1, IL1B, TLR2, DNMT1, and FGFR1. The direction of differential expression of these targets was consistent in the limma-voom and DESeq2 robustness analyses. AutoDock Vina binding energies for the 36 compound-receptor combinations ranged from-9.85 to-5.38 kcal/mol. Among them, acteoside-JAK2, acteoside-ERBB2, and acteoside-TLR2 exhibited relatively low binding energies, while buergerinin C-CASP1/ERBB2 and p-methoxycinnamic acid-ESR1 also displayed representative binding modes.Conclusion The potential effects of Scrophulariae Radix appear to be primarily associated with modulation of host inflammatory and immune responses following GAS infection, involving TLR2-related pathogen recognition, CASP1/IL-1β-mediated inflammatory responses, JAK2/STAT3-related cytokine signaling, and inflammation-associated oxidative stress and immune regulation. Representative constituents, including acteoside, ferulic acid, p-methoxycinnamic acid, and buergerinin C, showed overlapping structural coverage across multiple disease-relevant targets, suggesting that the potential pharmacologically active basis of Scrophulariae Radix is characterized by the coordinated involvement of multiple constituents acting on a shared disease network.