The Protective Mechanism Of Resveratrol In Oxidative Stress-Induced PC12Cell Injury

Objectives This experiment through the establishment of PC12cell oxidativestress model in vitro, to investigate the mechanism of injured PC12cells by H2O2.To observe the protective effect of resveratrol pretreatment on PC12cells injured bytreating with H2O2, to study its possible mechanism of action, whether through theantioxidant, regulating apoptosis related protein.Methods Treatment with H2O2in PC12cells in this study, PC12cells oxidativestress damage model. The viability of PC12cell induced by H2O2was detected byMTT assay. LDH kit and GSH kit was used to determine the cell damage. Todetermine the optimal protection of resveratrol concentration based on the aboveexperimental results. Cellular signaling factors such as p38MAPK, Bax, Bcl-2andCaspase-3phosphorylation were tested by western blot. To investigate the specificsignal transduction mechanism of neuroprotective effects of resveratrol.Results1MTT assay was used to detect cell activity. Compared with normalcontrol group, cells treated single resveratrol（10,20,40,80,100μmol/L）showedno significant change in cells survival rate（P>0.05）. Cells vitality fell downobviously（55.1±2.9%, P<0.05） about after PC12cells treated with H2O2（300μM）action time4h. Compared with the model group, the survival rate ofcells was increased significantly in different concentration of resveratrol（10,20,40,80and100μmol/L） pretreatment+H2O2group. In addition,40μM was the mosteffective concentration （89.3±3.1%, P<0.05）, indicating that resveratrol couldprotect neurons from oxidative damage.2Colorimetric method analysisdemonstrated, compared with the control group, PC12cells were treated withdifferent concentrations of H2O2for4h, the leakage of lactate dehydrogenase in thesupernatant increased gradually, the activity of GSH in cells decreased graduallywith the increasing of H2O2concentration（P<0.05）. Showed that during injuryPC12cells membrane integrity was damaged and cells antioxidant ability wasdecreased. However, compared with the model group, the leakage of LDH in theculture supernatant were reduced significantly while the levels of intracellularglutathione were increased significantly in different concentrations of resveratrol pretreatment group. In addition, it was the lowest（239.2±14.1U/L, P<0.05）and itwas the highest （33.3±1.3nmol/mg prot, P<0.05）in40μmol/L resveratrol group.Indicating that resveratrol could protect the integrity of the cell membrane, improvethe antioxidant ability of cells to achieve the neuroprotective effects.3Westernblotting analysis demonstrated, comparing to those in the control cells, the levels ofphosphorylation of p38MAP kinase, Bax and cleaved caspase-3increasedsignificantly（246.2±18.3%,248.2±14.1%,220.1±18.1%, P<0.05）while thelevels of Bcl-2decreased significantly（64.2±5.6%, P<0.05）at4h after the H2O2induced. The Bcl-2/Bax ratio decreased. Compared with the model group, the levelsof phosphorylation of p38MAP kinase, Bax and cleaved caspase-3decreasedsignificantly（129.2±7.3%,149.6±10.8%,167.9±19.0%, P<0.05）while the levelsof Bcl-2increased significantly（89.6±6.9%, P<0.05）. The Bcl-2/Bax ratioincreased. Indicating that resveratrol suppresses apoptosis and oxidative damageinduced by H2O2via up-regulation of bcl-2and down-regulation of bax, Caspase-3and p38MAPK pathway, increased the Bcl-2/Bax ratio. By using the p38MAPKinhibitor SB203580, the results are similar to those of Res further proof thatresveratrol could play a neuroprotective effect by inhibiting p38MAPK signalpathway.Conclusions1P38MAPK signalig pathway mediated the PC12cells injured byH2O2.2Resveratrol pretreatment has protective effects on PC12cells by thepotential mechanism of inhibiting the activation of p38MAPK signaling pathway,regulation of bcl-2and bax, improve the ratio of Bcl-2/Bax further blockage ofCaspase-3.