Effects of sodium arsenite on DNA methyl transferases and antagonistic effects of

Background  Exposure to arsenic can cause skin lesions in humans. DNA methylation changescaused by arsenic are one of its pathogenic mechanisms, but how arsenic causes DNAmethylation changes is not yet clear.

Objective  This study aims to investigate the effects of sodium arsenite (NaAsO₂) exposure onDNA methyl transferases (DNMTs) mRNA and protein expression levels of immortalized humankeratinocytes (HaCaT), and to explore the associations between oxidative stress induced byarsenic and the changes in DNMTs mRNA and protein expression levels.

Methods  HaCaT cells were treated with NaAsO₂ (0, 0.625, 1.25, and 2.50 μmol·L^-1, respectively);HaCaT cells were pretreated with antioxidant N-acetylcystein (NAC) 10.0μmol·L^-1 for 4h, and thentreated with 2.50 μmol·L^-1 NaAsO₂ for 72 h. Immunofluorescence method was used to detect thecontent of reactive oxygen species (ROS), biochemical method was used to detect the content ofmalondialdehyde (MDA), real-time fluorescence quantitative PCR was used to detect the DNMT1,DNMT3a, and DNMT3b mRNA expression levels, and Western blotting was used to detect the DNMTs protein expression levels in HaCaTcells.

Results  After 72 h of NaAsO₂ treatment, the ROS and MDA levels in HaCaT cells increased with higher dose (F_trend=441.675, P <0.001;F_trend=22.430, P=0.012); DNMT1 mRNA and protein expression levels in HaCaT cells in the 1.25 and 2.50 μmol·L^-1 groups were higherthan those in the control group (P <0.05). With the increase of NaAsO₂ dose, DNMT1 mRNA and protein expression levels in HaCaT cellsincreased (F_trend=30.280, P=0.001; F_trend=40.421, P <0.001), DNMT3a mRNA and protein expression levels decreased (F_trend=226.283, P <0.001; F_trend=30.848, P=0.001), DNMT3b protein expression level increased (F_trend=15.095, P=0.005), and there was no significant differencein the DNMT3b mRNA expression level between the NaAsO₂ treatment groups and the control group (P > 0.05). After NAC treatment, ROScontent, MDA level, and DNMT1 mRNA and protein expression levels in HaCaT cells decreased significantly compared with the 2.50 μmol·L^-1NaAsO₂ group (P <0.05); DNMT3a mRNA and protein expression levels did not change significantly (P > 0.05); DNMT3b protein expressionlevel decreased significantly (P <0.05), and its mRNA expression level did not change significantly (P> 0.05).

Conclusion  NaAsO₂ can change the mRNA and protein expression levels of DNMTs in HaCaT cells, and these changes may be related tothe oxidative stress induced by arsenic.