Effects of nitrate (NO3−) stress-induced exacerbated cadmium (Cd2+) toxicity on the inflammatory response, oxidative defense, and apoptosis in juvenile Japanese flounder (Paralichthys olivaceus)

Due to the discharge of industrial wastewater, urban domestic sewage, and intensive marine aquaculture tailwater, nitrate (NO₃⁻) pollution has emerged as a significant issue in offshore waters. Nitrate pollution affects aquatic life and may interact with other pollutants, leading to comprehensive toxicity. Cadmium (Cd²⁺) is the most widespread metal contaminant, adversely affecting aquatic life in the coastal waters of China. Despite this, few studies have focused on the synergistic toxicity of NO₃⁻ and Cd²⁺ in marine organisms. This study conducted a 30-day exposure experiment on marine Japanese flounder (Paralichthys olivaceus) to explore the synergistic toxicity of NO₃⁻ and Cd²⁺. Our results demonstrated that the exposure to Cd²⁺ alone induced slight histopathological changes in the liver. However, malformations such as hepatic vacuolar degeneration and sinusoid dilatation were exacerbated under co-exposure. Moreover, co-exposure induced the downregulation of antioxidants and the upregulation of the product malonaldehyde (MDA) from lipid peroxidation, indicating potent oxidative stress in the liver. The increased mRNA expression of IL-8, TNF-α, and IL-1β, along with the decreased expression level of TGF-β, indicated a synergistic inflammatory response in the organisms. Furthermore, the co-exposure led to an abnormal expression of P53, caspase-3, caspase-9, Bcl-2, and Bax, and disturbed the apoptosis in the liver through TUNEL staining analysis. Overall, our results imply that co-exposure synergistically affects inflammation, redox status, and apoptosis in flounders. Therefore, the findings from this study provide valuable perspectives on the ecological risk assessment of marine teleosts co-exposure to NO₃⁻ and Cd²⁺.