Study On Sodium Arsenite Inducing The Apoptosis Of PC12 Cells Through GRP78/CHOP Pathways And Protective Roles Of Hydrogen Sulfide

Objective: To explore the mechanisms of injuries caused by sodium arsenite（NaAsO₂）in PC12 cells and the effects of hydrogen sulfide（H2S）on cells exposed to NaAsO₂.Methods: The morphology and cell viability of PC12 cells were observed by inverted microscope and MTT assay.Apoptosis were detected with Hoechst33342/PI or Annexin V-FITC/PI double staining by flow cytometry and fluorescence microscope.Caspase-3 activity were assayed spectrophotometrically.The specific level of H2 S was detected by WSP-1.The protein expression of 3-mercaptopyruvate sulfotransferase（3MST）,apoptosis and endoplasmic reticulum stress-related proteins Bax,Bcl-2,GRP78,CHOP and Caspase-12 were measured by Western blot.Results: 1.After treatment with different concentrations of NaAsO₂（20,40,60,80 or 100μM）for 12,24,36 or 48 h,the cells shrank,cell gap widened and dead cell increased.Cell viability showed a dose and time-dependent decrease（P<0.05）.2.Compared with the control group,accompanied by the increase of the apoptosis rate,the activity of Caspase-3 also increased with the up-regulation of expression of Bax,GRP78,Cleaved caspase-12 and CHOP proteins whereas the expression of Bcl-2 protein was decreased in NaAsO₂ group cells（P <0.05）.3.The expression of 3MST protein and the level of intracellular H2 S were also reduced after treatment with different concentrations of Na AsO₂ for 24 h（P<0.05）.4.Pretreatment with NaHS（H2S donor,100-400 μM）for 30 min could reverse the changes of the cell morphology and the increase of apoptosis and the Caspase-3 activity.Meanwhile,NaHS could also up-regulate Bcl-2 protein and down-regulate the expression of Bax,GRP78,CHOP and Cleaved caspase-12.Conclusion: 1.NaAsO₂ can induce the injury of PC12 cells through GRP78/CHOP signaling pathways of endoplasmic reticulum stress;2.H2 S plays a neuroprotective role by antagonizing GRP78 / CHOP pathway.