Integrating Multi-Omics, Network Pharmacology, and Experimental Validation to Unveil the Molecular Mechanisms of Fructus Xanthii in Treating Asthma

Yiqun Dong^1*Zhaosen Fan²Xiaoxiao Li¹Yucheng Ming¹Dong Wang¹Yuanda Song^1*Youpeng Fan^1*

• ¹Qilu Medical University, Zibo City, China
• ²Shandong Benon Biological Technology Co., Ltd. No. 10818 Xiaoqinghe South Road, Tianqiao District, Jinan City, Shandong Province, China., jinan, China

This study employs an integrated approach combining multi-omics analysis, network pharmacology, machine learning, and experimental validation to elucidate the molecular mechanisms of Fructus Xanthii (Chinese name: Cang-Er-Zi) in treating asthma. Asthma-related differentially expressed genes (DEGs) were identified from GEO datasets (GSE63142 and GSE14787), yielding 3,755 DEGs (1,946 upregulated, including TFF1, FKBP5, AKR1B10, CXCL14, CST1, NOS2, CPA3, KRT6A, PHACTR3, TCN1, CLCA1, and CEACAM5; 1,809 downregulated, including LOC100132287, C7orf26, LTF, GPR156, SCGB3A1, SCGB1A1, CSH1, TMEM45A, C20orf46, FHOD3, C3, and LYPD2). Weighted gene co-expression network analysis (WGCNA) on GSE14787 identified the MEblack module (741 genes, e.g., POMP, GUSBL2, RBM39, PSMA6, RFX1, TCEB1, and PSMD6) as highly correlated with asthma pathogenesis (correlation coefficient 0.42). Active ingredient targets of Fructus Xanthii were predicted using TCMSP and SwissTargetPrediction, resulting in 1,317 potential targets, with 100 intersecting asthma DEGs. A "disease-drug-ingredient-target" network (96 nodes, 503 edges) highlighted core targets such as ALB, IL6, TNF, and HSP90AB1. Integration of machine learning algorithms (RF, SVM, XGB) with PPI network analysis identified seven hub targets: HSP90AB1, CCNB1, CASP9, CDK6, NR3C1, ERBB2, and CCK. Molecular docking demonstrated strong binding affinities (<-5 kcal/mol) between carboxyatractyloside and key targets (e.g., HSP90AB1 at -10.09 kcal/mol), with molecular dynamics simulations confirming complex stability (RMSD <3 Å, Rg 2.18–3.26 nm). Immune infiltration profiling via CIBERSORT showed reduced M2 macrophages and memory B cells alongside elevated plasma cells in asthma samples, underscoring Fructus Xanthii's immunomodulatory potential. In vivo experiments in an ovalbumin-induced murine asthma model validated the dose-dependent anti-inflammatory effects of Fructus Xanthii aqueous extract, manifesting as attenuated body weight loss, improved lung histopathology (H&E staining and scoring), decreased BALF cytokine levels (TNF-α, IL-6, IL-1β, IL-5), and suppressed mRNA/protein expression of hub genes (HSP90AB1, CCNB1, CASP9, PI3K, AKT1) via qPCR and Western blot. Collectively, Fructus Xanthii may exerts anti-asthmatic effects by modulating HSP90AB1/IL6/TNF and PI3K-AKT pathways, regulating inflammation, cell cycle, apoptosis, and immune homeostasis, thereby providing empirical support for multi-target traditional Chinese medicine strategies in asthma management.