Toxicol Mech Methods. 2026 Mar 18:1-16. doi: 10.1080/15376516.2026.2645333. Online ahead of print.
ABSTRACT
Arsenic (As) and cadmium (Cd) frequently co-occur in groundwater and target renal proximal tubules. Although each metal can engage ferroptosis pathways, how co-exposure shapes the integrated transcript-protein-lipid ROS axis remains unclear. Human HK-2 cells were exposed for 24 h to commonly used experimental concentrations of As (10 µM), Cd (10 µM), or As + Cd (5 µM + 5 µM). We profiled (i) a focused ferroptosis/oxidative stress gene panel and visualized Z-score heatmaps; (ii) ferroptosis-related proteins (GPX4, SLC7A11, ACSL4, PTGS2, TFR1, ferritin heavy/light chains (FTH/FTL), LC3, Bax/Bcl-2, cleaved caspase-3) by Western blot with densitometric analysis; and (iii) lipid peroxidation using BODIPY 581/591 C11 imaging (oxidized green/reduced red). Statistical analyses were performed using one-way ANOVA followed by Tukey’s post hoc test. These readouts were interpreted alongside measurements of cell viability, ROS, MDA, and metabolomics profiling. Co-exposure produced the most coherent transcriptional shifts across modules regulating iron handling, glutathione-cysteine metabolism, lipid remodeling, CoQ/mitochondrial function, macroautophagy, and NRF2 signaling. Protein analyses corroborated pathway activation: GPX4 and SLC7A11 decreased, whereas ACSL4, PTGS2, and TFR1 increased. FTH/FTL exhibited a downward trend, and LC3-II levels increased, consistent with impaired peroxide detoxification, enhanced polyunsaturated fatty acid (PUFA)-phospholipid acylation, increased iron import, and autophagy engagement. BODIPY C11 imaging revealed the highest oxidized fraction in the As + Cd group, indicating a maximal lipid-peroxide burden. As and Cd act cooperatively to (1) weaken the GPX4-xCT (cystine/glutamate antiporter; SLC7A11) antioxidant axis, (2) increase iron availability and iron-handling stress, and (3) remodel membrane PUFA pools, collectively tipping renal epithelial cells toward ferroptosis. These multi-layer signatures nominate ferroptosis inhibition and iron handling as potential therapeutic targets for mixed-metal nephrotoxicity.
PMID:41847867 | DOI:10.1080/15376516.2026.2645333
