Multifunctional assays and molecular simulations reveal the protective mechanisms of QNZ in mitigating PFOA-induced cytotoxicity in Nthy-ori3-1 cells.

Perfluorooctanoic acid (PFOA) is a persistent toxic chemical that enters the human body through the food chain and causes harm. In this study, human thyroid follicular epithelial cells (Nthy-ori3-1 cells) were used as a model. PFOA was shown to reduce cell viability through a cell activity assay. Notably, at a concentration of 100 µmol/L, PFOA enhanced the activity of antioxidant enzymes ((Total superoxide dismutase (T-SOD), catalase (CAT), glutathione (GSH), total Antioxidant Capacity (T-AOC)), while the level of intracellular reactive oxygen species (ROS) remained elevated. PFOA triggered oxidative stress by inducing abnormal ROS levels, which subsequently exacerbated the inflammatory response and affected cellular activity. Therefore, pretreatment with 10 µmol/L EVP4593 (QNZ, an NF-κB inhibitor) increased the percentage of normal cells from 74.06 % to 83.74 % at 1 µmol/L PFOA. Molecular docking and molecular dynamics indicated that QNZ competitively inhibited the interaction between PFOA and nuclear factor kappa-B (NF-κB) proteins. Correspondingly, Western blotting and qPCR analyses confirmed that QNZ suppressed the expression of apoptosis-related proteins (Tumor protein p53 (P53), tumor necrosis factor-alpha (TNF-α), Bcl2-associated X (BAX), B-cell lymphoma-2 (BCL-2), cysteine aspartate protease-3 (Caspase-3), NF-κB) and their related genes. Additionally, PFOA prompted cells to release thyroxine (T4) and triiodothyronine (T3). This study not only elucidates the toxic effects of PFOA on cells, but also reveals the defense mechanism by which QNZ effectively alleviates cell toxicity.