The Study Of Autophagy Cell Death Caused By Sodium Arsenite In Pancreatic β-cells Through ROS Pathway

Objective: Arsenic is a kind of toxic non-metallic compounds which widelydistributed in rocks、soil and water, exposed to high concentration of arsenic hasbecome a major public health problem all around the world. Arsenic toxicity of thepancreas gradually caused widespread concern, epidemiological and experimentalstudies show that arsenic exposure is a risk factor for type II diabetes, the mainmechanism is the cell damage or apoptosis which caused by disfunction of the islet βcell but the exact mechanism is not clear yet. For further investigate the effects ofarsenic on the pancreatic cells injury, This study focus on cell autophagy caused bysodium arsenite in pancreaticβ-cells (INS-1cells),and the effect of the reactive oxygenspecies (ROS) in this process.Methods: Rat islet β cells (INS-1) were selected for the study.3-（4,5–Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) was used to detectthe toxic effects of different concentrations of sodium arsenite on INS-1cells; Detectcell viability using different reagents for the pre-treatment when treated INS-1cellswith the same concentration of sodium arsenite;2’,7’–dihydro-dichloro withinfluorescein (DCFH) was used to detect intracellular ROS levels; Detect theintracellular ROS levels using different reagents for the pre-treatment when treated INS-1cells with the same concertration of sodium arsenite;Variations ofautophagosome were observed after Cyto-ID Green staining by fluorescencemicroscopy; Western blot was used to detect the intracellular protein level ofmicrotubules associated protein light chain3(LC3). SPSS17.0statistical software wasused for statistical analysis.Results:24hours after treated with sodium arsenite（1μmol/L、2μmol/L、4μmol/L）, INS-1cells activity were significantly weakened, and the cell activitydecreasing with the doses of sodium arsenite increasing; DCFH fluorescence valuesincreasing with doses of sodium arsenite increasing; Along with the dose of sodiumarsenite increased autophagosome that fluorescent stained increased; The expression ofLC3-II protein increasing with the doses of sodium arsenite increasing; Afther2hourspretreatment by lysosomal fusion inhibitor chloroquine (C q), the expression of LC3-IIprotein was significantly increased; Afther1hours pretreatment by ROS scavengerNAC, compared with the untreated group, the fluorescent stained autophagosome and theexpression of LC3-II protein was significantly reduced; Afther2hours pretreatment byautophagy inhibitor3-Methyladenine (3-MA),compared with the untreated group,cells activity were significantly strengthen; After2hours pretreatment by autophagyinducer rapamycin, compared with the untreated group, cells activity were significantlyweakened.Conclusion: Sodium arsenite caused autophagic cell death in INS-1cells throughROS pathway.