Role of NOX4 and NF-κB in oxidative stress of HepG2 cells induced by sodium fluoride

Objective To investigate the role of NOX4 and NF-κB in oxidative stress of HepG2 cells induced by sodium fluoride

Methods HepG2 cells were treated with sodium fluoride at concentrations of 0, 10, 20, 40, and 80 mg/L for 24 h, and the intramolecular malondialdehyde (MDA)content was determined by thiobarbituric acid method. The intracellular total superoxide dismutase (SOD) activity was determined by hydroxylamine method. The phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) in cells were detected by western blotting. Furthermore, pretreatment with NOX4 inhibitor apocynin for 30 min, then treated with 40 mg/L sodium fluoride for 24 h. The phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) in cells were detected by Western blotting.

Results Compared with control group, MDA content in 20, 40 and 80 mg/L sodium fluoride treated groups increased significantly (P < 0.01). Compared with control group, the total SOD activity in HepG2 cells decreased significantly after sodium fluoride treatment (P < 0.05). Compared with control group, the phosphorylation level of nuclear factor p65 of groups treated with 40 and 80 mg/L sodium fluoride and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) in groups treated with 20, 40 and 80 mg/L sodium fluoride increased significantly(P < 0.01 or 0.05). Apocynin pretreatment significantly decreased the phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4(NOX4) (P < 0.05 or 0.01).

Conclusions Induction of oxidative stress in HepG2 cells by sodium fluoride may be associated with activation of the NF-κB/NOX4 pathway.