ZnO Nanoparticles Interacts With Unique Serum Proteins and Induce Stress Signaling During Hepatoxicity ‐ A Proteomics and Molecular Pathways Study

The release of synthesized zinc oxide nanoparticles (ZnO-NPs) into the environment and consequent human exposure risk brought enormous attention in recent years. In recent decades, a range of toxicological effects were highlighted on hepatotoxic risk during ZnO-NPs exposure without precise mechanistic verification. The current study employed proteomic profiling (LC-MS/MS) to unveil the interaction of ZnO-NPs with human serum proteins (protein corona) and its relevant hepatotoxic mechanisms. Further, chicken embryo model and primary hepatocyte cell culture protocols (fluorescence assays and confocal imaging) were used to explore developmental hepatotoxicity and cellular stress responses respectively. Further, mediatory role of redox enzymes by siRNA experiments and synergistic effect of ZnO-NPs on cellular impact of disease risk factors were also explored. The protein corona composition provides plausible mechanistic insights on liver transport and associated hepatotoxic pathways (Reactome and KEGG). Moreover, the detrimental impact on developing embryonic liver, RBC and tissue redox enzymes were also observed. The expression studies (RT-qPCR and western blot) indicate an elevated response of oxidative stress associated genes (HO-1 and NQO1) and cellular stress signaling factors (HSP27, JNK, P53, c-JUN, MAPK). The current study revealed a stimulatory effect of ZnO-NPs on oxidative stress, cellular calcium overload and autophagy. Further, the mediatory effect of NOX2 and XO on ZnO-NPs induced cytotoxicity and superoxide generation was validated. Notably, ZnO-NPs exposure increased the adverse effect of inflammatory and metabolic risk factors in hepatocytes. Overall results highlight novel mechanistic insights on hepatotoxic effect of ZnO-NPs towards a precise assessment on the magnitude of human exposure risk.