Sulfiredoxin-1Protects PC12Cells Against Oxidative Stress Induced By Hydrogen Peroxide

Background: Oxidative stress results in protein oxidation and isimplicated in cerebral disease, such as Parkinson’s disease, Alzheimer’sdisease and ischemic stroke. Sulfiredoxin-1（Srxn1） is an endogenousantioxidant protein that has neuroprotective effects. The mechanisms ofSrxn1in oxidative stress have not been well studied, however.Objective: To explore the protection of Srxn1in PC12cells againstoxidative stress induced by hydrogen peroxide and its potential mechanism.Methods:（1） PC12cells were exposed to different concentrations ofhydrogen peroxide （60uM,100uM,140uM,180uM,200uM,240uM） for24h, and then we screened the optimal concentrations of hydrogenperoxide by MTT and LDH assays. We used the screened concentration ofhydrogen peroxide exposure to PC12cells for different times （3h,6h,12h,24h,48h） following screening the optimal exposure time by means ofMTT and LDH analysis.（2） Building three interfere plasmid vectors ofSrxn1（PLVT574, PLVT575, PLVT576） and one negative plasmid vector （PLVT4）, we transient transfected them to PC12cells by lipofectamine2000. After that we observed cell characteristics and transfectionefficiency under inverted microscope and fluorescence microscope. Todetermine interference efficiency, we used RT-PCR and Western Blotassays.（3） Using MTT and LDH assays to measure cell viability and celldamage, superoxide dismutase （SOD） and reduced glutathione kit to assayintracellular oxidative stress state. RT-PCR and immunoblot analysisfurther measured the relationship between Srxn1level and2-Cys Prdxsactivity.Results:（1） The results of MTT and LDH showed that180uM H₂O₂exposure for24h is the optimal oxidative stress model.（2） RT-PCR andimmunoblot analysis showed that the interference efficiency of PLVT574,PLVT576and PLVT4were not significant, but that of PLVT575wassignificant （P<0.01）, compared with control groups.（3） MTT and LDHshowed that knockdowns of Srxn1attenuated cell viability and promotedcell damage. Superoxide dismutase （SOD） and reduced glutathione kitassays suggested that SOD and GSH were reduced in Srxn1knockdownPC12cells. Furthermore, the decreased Srxn1correlated with a reduction in2-Cys Prdxs activity and2-Cys Prdxs protein levels were increased in theH₂O₂dosed cells, as measured by RT-PCR and immunoblot analysis.Conclusions: These results suggested that Srxn1can protect PC12cellsfrom H₂O₂-induced oxidative stress and are involve in Prdxs activity. Srxn1 play a protective role against oxidative injury and demonstrates potential as atarget for neuroprotective intervention in oxidative stress.