Distinct concentration-dependent oxidative stress profiles by cadmium in a rat kidney proximal tubule cell line

Abstract

Levels and chemical species of reactive oxygen/nitrogen species (ROS/RNS) determine oxidative eustress and distress. Abundance of uptake pathways and high oxygen consumption for ATP-dependent transport makes the renal proximal tubule particularly susceptible to cadmium (Cd²⁺)-induced oxidative stress by targeting ROS/RNS generation or antioxidant defence mechanisms, such as superoxide dismutase (SOD) or H₂O₂-metabolizing catalase (CAT). Though ROS/RNS are well-evidenced, the role of distinct ROS profiles in Cd²⁺ concentration-dependent toxicity is not clear. In renal cells, Cd²⁺ (10-50 µM) oxidized dihydrorhodamine 123, reaching a maximum at 2-3 h. Increases (up to fourfold) in lipid peroxidation by TBARS assay and H₂O₂ by Amplex Red were evident within 30 min. ROS and loss in cell viability by MTT assay with 50 µM Cd²⁺ could not be fully reversed by SOD mimetics Tempol and MnTBAP nor by SOD1 overexpression, whereas CAT expression and α-tocopherol were effective. SOD and CAT activities were attenuated below controls only with >6 h 50 µM Cd²⁺, yet augmented by up to 1.5- and 1.2-fold, respectively, by 10 µM Cd²⁺. Moreover, 10 µM, but not 25-50 µM Cd²⁺, caused 1.7-fold increase in superoxide anion (O₂^•-), detected by dihydroethidium, paralled by loss in cell viability, that was abolished by Tempol, MnTBAP, α-tocopherol and SOD1 or CAT overexpression. H₂O₂-generating NADPH oxidase 4 (NOX4) was attenuated by ~50% with 10 µM Cd²⁺ at 3 h compared to upregulation by 50 µM Cd²⁺ (~1.4-fold, 30 min), which was sustained for 24 h. In summary, O₂^•- predominates with low-moderate Cd²⁺, driving an adaptive response, whereas oxidative stress by elevated H₂O₂ at high Cd²⁺ triggers cell death signaling pathways.Highlights Different levels of reactive oxygen species are generated, depending on cadmium concentration. Superoxide anion predominates and H₂O₂ is suppressed with low cadmium representing oxidative eustress. High cadmium fosters H₂O₂ by inhibiting catalase and increasing NOX4 leading to oxidative distress. Superoxide dismutase mimetics and overexpression were less effective with high versus low cadmium. Oxidative stress profile could dictate downstream signalling pathways.

Keywords: Catalase; Hydrogen peroxide; Reactive oxygen species; Redox; Superoxide.