Comprehensive analysis of the association between perfluorooctanoic acid exposure and osteosarcoma progression.

Perfluorooctanoic acid (PFOA), a persistent environmental pollutant, represents a chronic environmental stressor, yet its role in osteosarcoma progression remains unclear. Here, we integrated network toxicology, machine learning, immune infiltration analysis, single-cell RNA sequencing, molecular docking, molecular dynamics simulation, and in vitro assays to define the mechanisms linking long-term PFOA exposure to osteosarcoma. We identified 28 shared targets between PFOA-associated proteins and osteosarcoma-related genes, which were mainly enriched in oxidative stress, immune regulation, and metabolic reprogramming pathways. Machine learning prioritized nine candidate hub genes, including PNP, SOD2, MAT1A, RELA, PSAT1, CXCL12, PHGDH, PDK4, and CFD, with PHGDH, PSAT1, PDK4, and MAT1A suggesting a metabolism-related axis relevant to PFOA-associated osteosarcoma alterations. Single-cell and immune analyses showed distinct cell-type-specific expression patterns of these genes and their close associations with the tumor immune microenvironment. Molecular docking and molecular dynamics simulations further suggested potential structural compatibility between PFOA and several prioritized hub proteins. Experimentally, prolonged exposure to non-cytotoxic concentrations of PFOA enhanced osteosarcoma cell proliferation and invasion, whereas high concentrations were cytotoxic. Collectively, these findings suggest that long-term PFOA exposure may promote osteosarcoma progression by coupling environmental stress with metabolic reprogramming and immune dysregulation.