Arsenic Impairs Mitophagy Through Pink1 Pathway In Pancreas And The Protective Effect Of Taurine

Objective:Arsenic is a toxic nonmetallic compound,which is widely distributed in rocks,soil and water resources.Chronic arsenic poisoning is a global public health problem.Arsenic exposure is recognized as one of the risk factors of diabetes mellitus,possibly due to the cell apoptosis or injury induced by arsenic exposure in islet?-cell.PINK1 is a protein kinase,mainly located in the outer membrane of mitochondria,is an important protein that mediates mitophagy.In our previous work,it was found that arsenic exposure elevated the total level of intracellular reactive oxygen species.Active oxygen mainly acts on mitochondria,and mitochondrial autophagy can effectively remove damaged mitochondria.However,whether arsenic exposure causes mitophagy dysfunction and whether mitophagy dysfunction caused by arsenic exposure is associated with PINK1 is unclear.Taurine,also known as?-aminoethanesulfonic acid,was first isolated from the bezoar,is a sulfur-containing non-protein amino acids.It has been reported that taurine has the effect of lowering blood sugar and antioxidant,and related studies have shown that taurine can effectively reduce the production of reactive oxygen species.In the previous study,we demonstrated that taurine was able to effectively attenuated the autophagic injury of offspring rat liver and HepG2 cells induced by As₂O₃,but whether it could prevent the mitophagy dysfunction caused by arsenic exposure and whether it played an interventional role via the PINK1 pathway is not clear.In order to further explore the effect of arsenic exposure on rat pancreatic cell mitophagy and the protective effect of taurine,in this study we investigate the potential molecular mechanism of NaAsO₂ and As₂O₃ and the protective mechanism of taurine in rat islet?-cell.Methods:In vivo,Wistar rats were used as experimental subjects.The pregnant rats were exposed to As₂O₃ at a concentration of 2 mg/kg BW⁸ mg/kg BW,and were exposed to 8 mg/kg BW As₂O₃ after pretreatment with taurine at a concentration of 150 mg/kg BW,until the offspring were weaned,and then the offspring rats were exposed to the same dose of As₂O₃ for sexual maturation for 57 days.We used the electronic microscopy to observe the formation of mitochondria in pancreas cells of offspring rats.The expression of PINK1 protein,Bcl-2 protein and Bax protein in the pancreas cells of offspring rats were detected by Western blotting.In vitro,we treated INS-1 cells with the NaAsO₂ concentration of 1?M to 4?M at1h?3h?6h?12h?24h.The morphology of mitochondria in INS-1 cells was observed by electron microscopy.The mitochondrial transmembrane potential???m?was observed by JC-1 staining with fluorescence microscopy.The expression of PPARgprotein in INS-1 cells was evaluated by Western blot.The expression of PINK1protein in INS-1 cells was detected by immunofluorescence assay.Whether apoptosis occurred in INS-1 cells was observed by TUNEL assay.After pretreatment with taurine and PPARgactivator rosiglitazone,the INS-1 cells were treated with NaAsO₂in the same manner as above,we detected the changing of the experimental index measured by the methods mentioned above respectively.Results:In vivo,electron microscopy revealed that swelling and vacuole-like changes of mitochondria were observed in the rat pancreas exposed to different concentrations of As₂O₃?2 mg/kg BW⁸ mg/kg BW?.The degree of swelling was significantly correlated with As₂O₃ concentration,but the mitochondria swelling of taurine protected group was not obvious.The expression of PINK1 protein in the pancreas of the offspring rats decreased as the concentration of As₂O₃ increased,and the expression of PINK1 protein in the taurine protected group was higher than that in the 8mg/kg BW As₂O₃ exposed group.The expression of Bax protein in pancreas of offspring rats increased with the increase of As₂O₃ concentration,while the change of Bcl-2 protein was opposite.Compared with 8 mg/kg BW As₂O₃ exposure group,the expression of Bax protein was decreased and the expression of Bcl-2 protein was increased in taurine protected group.In vitro,electron microscopy revealed that the mitochondria in INS-1 cells were swollen and vacuolated when exposed to 4?M NaAsO₂,but the swelling was not obvious in the taurine protected group.INS-1 cells were treated with NaAsO₂ at a concentration of 1?M⁴?M for 1 h,3 h,6 h,12 h,24 h.INS-1 cells were treated with NaAsO₂ for 6 h,the mitochondrial membrane potentials of INS-1 cells were decreased by fluorescence microscopy,while the mitochondrial membrane potential was relieved after pretreatment with taurine.The expression of PPARgprotein in INS-1 decreased significantly with the increase of NaAsO₂ exposure dose,but in the protective group and the PPARgactivator group,the expression level of the protein was significantly higher than the group of 4?M NaAsO₂.INS-1 cells were treated with NaAsO₂ at a concentration of 1?M⁴?M for 1 h,3 h,6 h,12 h,24 h,the expression of PINK1protein was significantly decreased with the increase of NaAsO₂ exposure dose at 24 h.And in the taurine protected group and PPARgactivator group,the expression level was significantly higher than the group of 4?M NaAsO₂.TUNEL assay showed that after being treated with NaAsO₂ for 24 h,INS-1 cells began to apoptosis.Pretreatment with taurine reduced the incidence of apoptosis.Conclusion:Arsenic can cause apoptosis of rat pancreas cells;Arsenic exposure can down-regulate the expression of PINK1 to inhibit mitochondrial autophagy and cause apoptosis;Taurine can act as a protective agent to alleviate mitochondrial autophagy and apoptosis induced by arsenic exposure.