Network Toxicology and Bioinformatics Reveal Synovial Immune-Inflammatory Mechanisms of DEHP/DBP in Osteoarthritis

Background: Osteoarthritis (OA) is increasingly recognized as an immune-inflammatory disease of the joint, with synovial dysfunction contributing to cartilage degeneration. Phthalate plasticizers such as di(2-ethylhexyl) phthalate (DEHP) and dibutyl phthalate (DBP) are ubiquitous environmental toxicants; however, their potential association with synovial immune perturbation in OA remains unclear. Methods: We integrated four synovial-tissue transcriptome datasets from GEO (GSE1919, GSE82107, GSE55235, GSE55457) and performed batch correction and differential expression analysis, followed by weighted gene co-expression network analysis (WGCNA) to identify phenotype-associated modules. Three machine-learning algorithms (LASSO, SVM-RFE, Random Forest) were used to select robust markers, which were validated in an independent dataset (GSE12021). Immune infiltration was profiled by ssGSEA. DEHP/DBP targets were predicted using cheminformatics resources and intersected with phenotype-related genes. Molecular docking and all-atom molecular dynamics (MD) simulations evaluated the binding modes and stability of DEHP with core protein targets. Results: We identified 1,394 differentially expressed genes and OA-associated WGCNA modules enriched for cytokine signaling and leukocyte activation. Intersecting phthalate-predicted targets with phenotype-related and differentially expressed genes yielded plasticizer-linked OA candidate genes. Convergent machine-learning selection nominated six core genes—ATF3, SLC2A3, PIM1, VEGFA, RGS4 and TSPO—showing consistent dysregulation and diagnostic utility across training and validation cohorts. ssGSEA indicated altered synovial immune-cell landscapes, and several core genes correlated with pro-inflammatory cell subsets. Docking and MD suggested comparatively stable binding of DEHP to TSPO, SLC2A3 and PIM1, supporting a plausible molecular interface between phthalate exposure and synovial immune dysregulation in OA. Conclusions: This integrative network-toxicology study links DEHP/DBP exposure to synovial immune–inflammatory signatures in OA and prioritizes six core genes as potential biomarkers or mechanistic candidates. The computational findings generate testable hypotheses for experimental validation and may inform exposure-mitigation and therapeutic strategies.