Abstract: BACKGROUND: As the primary organ for arsenic metabolism in the body, the liver has become a focal point for research on the mechanisms of arsenic toxicity.
OBJECTIVE: To investigate the role of pregnane X receptor in sodium arsenite-induced oxidative stress and inflammatory injury in human normal hepatocytes.
METHODS: Human normal hepatocyte MIHA cells were exposed to 0 (control), 10, 20, 30 μmol/L sodium arsenite for 48 hours. Changes in cell morphology were observed. Cell viability was measured via the cell counting kit-8 assay. Intracellular reactive oxygen species levels were detected using fluorescent probe staining combined with a microplate reader. Intracellular malondialdehyde content, glutathione reductase activity, and total superoxide dismutase activity were assessed using the thiobarbituric acid method, nicotinamide adenine dinucleotide phosphate method, and WST-8 method, respectively. Levels of interleukin-6, tumor necrosis factor-α, and interleukin-1β in the cell supernatant were quantified via ELISA. mRNA expression of pregnane X receptor and cytochrome P450 3A4 enzyme was analyzed using real-time quantitative PCR. Protein expression levels of pregnane X receptor, cytochrome P450 3A4 enzyme, nuclear factor κB p65, nuclear factor κB p-p65, proliferating nuclear antigen, interleukin-6, interleukin-1β, tumor necrosis factor-α, inhibitor of nuclear factor κB α, cyclooxygenase 2, phosphorylated inhibitor of nuclear factor κBα, nuclear factor erythroid 2-related factor 2, kelch-like ECH-associated protein 1 antibody, phosphorylated nuclear transcription factor erythroid 2-related factor 2 were evaluated via western blot.
RESULTS AND CONCLUSION: Compared with the control group, cells in the sodium arsenite groups at various concentrations showed unclear cell membrane boundaries, reduced cytoplasm, decreased cell fusion rates, and widened intracellular space. Compared with the control group, the intracellular levels of reactive oxygen species and malondialdehyde were significantly increased in all sodium arsenite concentration groups (P < 0.05). The levels of interleukin-6, interleukin-1β, and tumor necrosis factor-α in the cell supernatant were also significantly increased (P < 0.05). The protein expressions of p-nuclear factor κB inhibitor protein α, nuclear factor κB p-p65, nuclear factor κB p65, tumor necrosis factor-α, and interleukin 1β were all increased (P < 0.05), and cell survival rates were all decreased (P < 0.05). The protein expressions of proliferation nuclear antigen, nuclear factor erythroid 2-related factor 2, phosphorylated nuclear factor erythroid 2-related factor 2 protein expression, and total superoxide dismutase activity were all decreased (P < 0.05). The mRNA and protein expressions of progesterone X receptor and cytochrome P450 3A4 enzyme were all decreased (P < 0.05). Compared with the control group, the intracellular glutathione reductase activity was reduced in the 20 and 30 μmol/L sodium arsenite groups (P < 0.05), and the expression of kelch-like ECH-associated protein 1, 
interleukin-6, and cyclooxygenase-2 proteins was increased (P < 0.05). To conclude, sodium arsenite may induce oxidative stress and inflammatory damage in liver cells by down-regulating the expression of pregnane X receptor, inhibiting the Nrf2 antioxidant pathway and activating the nuclear factor-κB inflammatory pathway, while also suppressing the expression of cytochrome P450 3A4 enzyme.

Key words:  sodium arsenite, liver injury, pregnane X receptor, oxidative stress, nuclear factor-κB signaling pathway, inflammatory response, CYP3A4, tissue construction