Antioxidant activity of endogenously produced nitric oxide against the zinc oxide nanoparticle-induced oxidative stress in primary hepatocytes of air-breathing catfish, Clarias magur

The facultative air-breathing magur catfish (Clarias magur) regularly encounter various environmental challenges including the exposure to nanomaterials discarded as industrial wastes in water bodies. The present investigation aimed at determining the possible ZnO NP-induced oxidative stress and also the antioxidant strategy of nitric oxide (NO), generated endogenously, in primary hepatocytes of magur catfish. Exposure of primary hepatocytes to different concentrations of ZnO NPs (5 and 10 μg/mL) led to a sharp rise of intracellular concentrations of hydrogen peroxide (H₂O₂) and malondialdehyde (MDA) within 6 h, which decreased gradually at later stages. This phenomenon was accompanied by an initial decrease of superoxide dismutase (SOD) and catalase (CAT) activities, the expression of their corresponding genes and the enzyme protein levels, with a subsequent significant increase of all these parameters at later stages. Most interestingly, exposure to ZnO NPs also stimulated the NO production by the primary hepatocytes as a consequence of induction of inducible nitric oxide synthase (iNOS) activity, higher expression of nos2 gene and iNOS protein. Furthermore, when the NO production by the hepatocytes was inhibited by either aminoguanidine (inhibitor for iNOS) or BAY (inhibitor for NFκB) in the presence of ZnO NPs, the intracellular concentrations of H₂O₂ and MDA was significantly elevated. This elevation was accompanied by a subsequent decrease of sod and cat genes expression, thereby suggesting that the inhibition of NO production leads to oxidative stress. Thus, it is believed that the magur catfish uses the strategy of stimulation of endogenous NO production by inducing the nos2 gene and simultaneous NO-mediated induction of sod and cat genes to defend against the NP-induced oxidative stress. It is the first report of such NO-mediated antioxidant strategy in any teleost fish to defend against the NP-induced oxidative stress and corresponding cellular damages.