The Mechanism Of Sodium Arsenite-induced DNA Damage In INS-1Cells

Introduction: Arsenic （As） is a highly toxic metalloid element which isubiquitous in nature. Chronic arsenic poisoning has become a major public healthproblem facing in the world. There are many wards of chronic arsenic poisoning. Inorder to protect people’s health, World Health Organization （WHO）, European Union,Japan and the United States （U.S.） has revised arsenic standard in drinking water to10μg/L, the new “Drinking Water Standards” of China （July1,2006） has revisedarsenic content standard from50μg/L to10μg/L.The U.S. Centers for Disease Control （CDC） and the International Agency forResearch on Cancer （IARC） assessed Arsenic as the first class of carcinogens. Arsenic（As） has been evaluated for its genotoxic effects. However, the mechanisms of theeffects are not well understood. Studies have shown that inorganic arsenic iscarcinogen for human skin and lung, and the occurrence of liver cancer, bladdercancer, kidney cancer and colorectal cancer is closely related to inorganic arsenic. Inaddition, the findings show that sodium arsenite has genetic toxicity, but there is littleresearch on the mechanism of genotoxicity. Studies have shown that sodium arsenitecan lead to increase of reactive oxygen species （ROS） content in cells to generateincreased and8-hydroxy-deoxy guanosine （8-OHdG） expression increased.In this study, INS-1cells were treated with different concentrations of NaAsO₂,intracellular ROS and glutathione （GSH） level,8-hydroxy-deoxy guanosine （8-OHdG）expression and DNA damage were explored to investigate the possible mechanism ofDNA damage caused by sodium arsenite.Methods: Single cell gel electrophoresis assay （SCGE） was used to evaluate thegenetic toxicity of sodium arsenite in INS-1. To elucidate the oxidative DNA damagemechanism in INS-1cells, we used the2’,7’-dichlorofluorescein diacetate（DCFH-DA） and o-phthalaldehyde （OPT） to monitor the levels of ROS and glutathione （GSH）. In addition,8-OHdG, which is a reliable marker for oxidativeDNA damage, was also measured by immunocytochemistry staining analysis. The datawere statistically analyzed by SPSS11.5software.Results: DNA damage induced by sodium arsenite （2-16μmol/L） wassignificantly increased in a dose-dependent manner in INS-1cells for12h in theSCGE assay. Twelve hours exposure of INS-1cells to different concentrations ofsodium arsenite （2-16μmol/L） results in a significant increase in the8-OHdGformation. Moreover, the formation of intracellular ROS was significantly increasedin NaAsO₂-treated cells exposed to different concentrations （0-16μmol/L） for6hoursand12hours in a dose-dependent manner. Sodium arsenite from8μmol/L to16μmol/L treated with INS-1cells for24hours, the expression levels of intracelluarROS was increased, INS-1cells were exposed to sodium arsenite （2-16μmol/L） for48hours, but no obvious effect was found （data not shown）; The levels of intracelluarGSH observed in INS-1cells was a statistically significant increase when the cellswere exposed to （2-16μmol/L） sodium arsenite for6,12,24and48hours.Conclusion: sodium arsenite caused DNA strand breaks in INS-1which inducedby oxidative stress.