Inorganic Arsenic Induces Pyroptosis And ? Cells Dysfunction Through Stimulating The IRE1?/TNF-? Pathway And Protective Effect Of Taurine

Objective:Chronic exposure to inorganic arsenic?iAs?is a risk factor for?cells dysfunction.Our previous studies demonstrated that iAs induced endoplasmic reticulum stress and pyroptosis in C57/BL6J mice,and taurine?Tau?decreased the activation of pyroptosis.Inositol-requiring enzyme 1 alpha?IRE1??,one of the three classic ER stress signaling pathways,is a key driver of inflammation.However,the mechanism of IRE1?regulates pyroptosis is still unclear.Therefore,in this study,we investigated the mechanism of As₂O₃-induced?cells dysfunction and the protection of Tau in pancreas of rat offspring.Methods:In vivo,the Wistar mice were used as experimental model.All the pregnant rats and offspring were exposed to As₂O₃?2⁸ mg/kg?and Tau?150 mg/kg?once a day.The pregnant rats were given As₂O₃ and Tau by gavage from GD 6 until pup postnatal day 28?PND 28?.The offspring were given As₂O₃ and Tau by gavage from PND 28 until PND 42.At last,the offspring were euthanized,and the pancreas were collected for the following experiments.The morphology of pancreas was observed by hematoxylin-eosin?HE?staining.The expression of inositol-requiring enzyme 1 alpha?IRE1??,phosphorylated protein p-IRE1?,tumor necrosis factor-??TNF-??,NOD-like receptor family pyrin domain-containing 3?NLRP3?,pro-caspase-1,caspase-1,apoptosis-associated speck-like protein?ASC?,pro-interleukin?IL?-1?,IL-1?and MPO in pancreas were detected by Western blot.In vitro,we used INS-1 cells for the study.The cytotoxicity of As₂O₃ was detected by MTT assay.The expression of p-IRE1?in INS-1 cells treated with 1⁴?M As₂O₃for 24 h was detected by immunofluorescence staining;The expression of TNF-?,NLRP3,pro-caspase-1,caspase-1,ASC,pro-IL-1?,IL-1?was detected by Western blot.INS-1 cells were pretreated with NLRP3 inhibitor?MCC950?,TNF-?inhibitor?Lenalidomide?,IRE1?inhibitor?Irestatin 9389?and Tau before treatment with 4?M As₂O₃ for 24 h.Immunofluorescence staining was used to detect the expression of p-IRE1?;Western blot was used to detect the expression of TNF-?,NLRP3,caspase-1,IL-1?in INS-1 cells.Flow cytometry was used to examine the pyroptotic INS-1 cells.Lactate dehydrogenase?LDH?assay kit was used to measure the LDH release.The Rat INS?Insulin?ELISA Kit was used to measure the insulin release.Results:In vivo,the structures of pancreas in As₂O₃-treated groups were irregularly arranged and not clear,with glands swelling.After Tau treatment,improvements on the morphology of pancreas were observed;The p-IRE1?,TNF-?,NLRP3,caspase-1,the level of IL-1?and MPO were significantly increased in the As₂O₃ treated group at 8 mg/kg and all were significantly decreased in Tau treated group compared with only As₂O₃-treated group.In vitro,INS-1 cells were treated with different concentrations of As₂O₃ for 24 h,and INS-1 cells viability decreased with the increase of As₂O₃ concentration.After treated with 1⁴?M As₂O₃ for 24 h,the expression of p-IRE1?,TNF-?,NLRP3,caspase-1 and IL-1?were increased.After treated with NLRP3 inhibitor?MCC950?,TNF-?inhibitor?Lenalidomide?,IRE1?inhibitor?Irestatin 9389?and Tau the expression of NLRP3 inflammasome related proteins were decreased compared with the As₂O₃-treated group;The results of flow cytometry showed that the percentages of pyroptotic cells was significantly decreased compared with the As₂O₃-treated group;The level of LDH release rate in INS-1 cells were decreased compared with the As₂O₃-treated group;Importantly,the insulin release suppressed by As₂O₃ were promoted.Conclusion:In conclusion,our study demonstrated that As₂O₃ upregulated the level of IRE1?,TNF-?and triggered NLRP3 inflammasome activation,leading to INS-1 cells pyroptosis and dysfunction;however,these effects could be improved by Tau.Tau markedly protected As₂O₃-induced?cells dysfunction by reduced the phosphorylation of IRE1?,secretion of TNF-?and activation of pyroptosis.