Environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin induces non-obstructive azoospermia: New insights from network toxicology, integrated machine learning, and biomolecular modeling

As industrial pollution intensifies, global male semen quality has been declining at a rate of 2.64 % per year in the 21st century. Among the various types of infertility, non-obstructive azoospermia (NOA) is the most severe and is closely associated with exposure to environmental toxins. The molecular mechanisms by which 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a typical persistent organic pollutant, induces NOA have yet to be systematically elucidated. This study employed the single-sample Gene Set Enrichment Analysis (ssGSEA) method to identify key toxicological pathways and constructed a diagnostic model based on 113 machine learning algorithms. By integrating Weighted Gene Co-expression Network Analysis (WGCNA) and single-cell analysis, we identified hub genes associated with the Sertoli Cell-Only Syndrome (SCOS) subtype. Finally, biomolecular modeling was conducted to validate the binding efficacy of the hub genes with TCDD. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis based on the ssGSEA method indicated that TCDD may disrupt spermatogenesis by activating the Tumor Necrosis Factor (TNF) and Mitogen-Activated Protein Kinase (MAPK) signaling pathways while inhibiting the Vascular Endothelial Growth Factor (VEGF) signaling pathway, ultimately leading to NOA. Through the integration of machine learning techniques, 5 hub genes (AUC > 0.7) induced by TCDD and associated with NOA were identified: Androgen receptor (AR), Chromodomain Helicase DNA-Binding Protein 1 (CHD1), Discoidin Domain Receptor Tyrosine Kinase 2 (DDR2), Retinoic Acid Receptor-Related Orphan Receptor Alpha (RORA), and Glutamate Ionotropic Receptor AMPA Type Subunit 1 (GRIA1). WGCNA and single-cell analysis revealed that AR and DDR2 were specifically expressed in the testicular tissues of NOA patients and were closely associated with SCOS (p < 0.05). Immune infiltration analysis suggested that TCDD induces abnormal infiltration of various immune cells, indicating its close relationship with immune inflammatory responses (p < 0.05). Biomolecular modeling further demonstrated a strong binding affinity between AR and TCDD (∆G = -8.3 kcal·mol⁻¹, Etotal = -37.79 kcal·mol⁻¹), highlighting the critical role of AR in TCDD-induced NOA. This study reveals the potential molecular mechanisms by which TCDD induces NOA, providing new targets for the development of diagnostic and therapeutic strategies.