The Effect And Mechanism Of Anthocyanins On Hydrogen Peroxide Induced Apoptosis Of Airway Epithelial Cells

ObjectivesIn this study,bronchial epithelial cells were treated with different concentrations of anthocyanin and an oxidative stress model was established by hydrogen peroxide to investigate the effect and possible mechanism of anthocyanin on hydrogen peroxide induced apoptosis.Methods:Hydrogen peroxide was treated on bronchial epithelial cells to induce oxidative stress model.1.MTT method was used to detect the effect of anthocyanin on hydrogen peroxide induced the activity inhibition in 16HBE cell;2.DCFH-DA probe was loaded into epithelial cells and to detect the effect of anthocyanin on intracellular reactive oxygen species by flow cytometry;3.Colorimetry was used to detect the changes of MDA level,SOD and GPx activity in 16HBE cells;4.Apoptosis was observed with hoechst33342 staining by the fluorescence microscope;5.JC-1 probe was loaded into the cell to detect the changes of mitochondrial membrane potential;6.The levels of anti-apoptotic protein bcl-2,pro-apoptotic protein bax,PTEN,and phosphorylation of AMPK and AKT were detected by Western Blot.Results:1.MTT results showed that anthocyanin at concentrations of 125 ?g/ml and 250?g/ml could inhibit hydrogen peroxide induced cytotoxicity(P<0.05);2.Anthocyanin could significantly inhibit the increase of intracellular ROS and MDA and the decrease of SOD and GPx activity induced by hydrogen peroxide(P<0.05),which is consistent with the antioxidant NAC.3.The results of hoechst 33342 staining showed that anthocyanin could signifi-cantly inhibit apoptosis induced by hydrogen peroxide;4.The results of mitochondrial membrane potential assay kit with JC-1 showed that anthocyanin could significantly inhibit the mitochondrial membrane potential re-duction induced by oxidative stress;5.WB results showed that in the hydrogen peroxide treatment group,the phos-phorylation level of AMPK and AKT was significantly increased,while the expression of PTEN was increased(P<0.05),which same effect as AMPK inhibitor Compound C.After anthocyanin treatment,AMPK and AKT phosphorylation level was significantly reduced,PTEN expression was decreased(P<0.05).6.WB results showed that anthocyanin or Compound C could significantly increase the ex-pression of bcl-2 and decrease the expression of bax in hydroxide-treated cells(P<0.05).Adding LY294002 in combination with anthocyanin or Compound C could significantly inhibit the ex-pression of bcl-2 and increase the expression of bax(P<0.05).Conclusion:Anthocyanin can inhibit hydrogen peroxide induced oxidative stress and apopto-sis of 16HBE cells,and its mechanism may be to regulate the PTEN/AKT signaling pathway by inhibiting AMPK activation induced by oxidative stress.