[Effects on survival of shRNA mediated APE/Ref1 gene silencing in rat spiral ganglion cells in oxidative stress].

Objective To investigate the effects of reducing APE/Refl expression in the cultures of rat spiral ganglion cells with oxidative damage induced by H2O2. Methods Primary cultured rat spiral ganglion cells were infected with small interfering RNA to APE/Refl （Apel siRNA） for 72 h,followed by treating with H2O2（0,10,25,50,100 and 300 μmol/L） for 1 h,and then cultured in normal medium for 24 h.Western blot were used to detect the level of APE/Refl protein and phosphorylation of histone protein H2AX in the infected cells.The caspase3 activation was tested by spectrophotometric method.The cell viability was determined by MTT and the apoptosis of spiral ganglion cells was determined by terminal-deoxynucleotidyl transferase mediated nick and labeling （TUNEL）. Results Western blot showed that infection with Ape1siRNA resulted in APE/Ref1 reduced expression in the spiral ganglion cells.Exposing spiral ganglion cultures with reduced expression of APE/Ref1 to H2O2 （50,100,300 μmol/L） for 1 h resulted in increasing in the phosphorylation of histone protein H2AX.The reduction in APE/Refl significantly reduced cell viability in cultures 24 h after 1 h expression to 50-300 μmol/L H2O2.The apoptosis of cells and caspase 3 activity was detected significantly improved. Conclusions The induced of APE/Refl results in significantly decrease in spiral ganglion cells viability in oxidative stress.The repairing function of APE/Refl is necessary for optimal levels of neuronal rat spiral ganglion cells survival. Key words: Spiral ganglion; RNA, small interfering; DNA-（apurinic or apyrimidinic site） lyase; Oxidative stress; Hydrogen peroxide