Interventional Effect Of Lutein Against Oxidative Stress Induced By H₂O₂on M（u|¨）ller Cell And Its Mechanism

Background Oxidative stress is one main cause of the age-related macular degeneration.Lutein has an important role for prevent age-related macular degeneration, but its antioxidant mechanism remains unclear.Objective The present study was to investigate the effect of lutein on oxidative stress of Muller cells on Nrf2/ARE signaling pathway.Methods Muller cells at Logarithmimic growth phase were incubated in the96well plate overnight. Draw the growth curve of Muller cell by different concentrations of hydrogen peroxide, get median lethal dose of hydrogen peroxide(160umol/L).The cells were divided into the blank group and the control group treated with160μmol/LH2O2. Muller cells were treated with different concentrations:lutein at12.5,25,50mg/L and hydrogen peroxide (H2O2) at160μmol/L in10%serum for24hours.The effect of different concentrations of lutein and hydrogen peroxide on Muller cells were evaluated by MTT assay. Different concentrations of reactive oxygen species(ROS) was detected by Flow cytometry(FCM);The expressions of Nrf2mRNA-. HO-1and Caspase-3mRNA in Muller cells were detected by quantitative real-time PCR(qRT-PCR); The expressions of Nrf2protein、HO-1protein and Caspase-3protein in Miiller cells were detected by Western blot.Results The A570value of Miiller cells showed a significant difference among the different groups (F=43.89,P<0.05).Compared with the control group,As70values were significantly higher in the experimental groups (P<0.05).FCM and CLSM analysis showed a significant difference among the different groups (P<0.05),Compared with the control group,ROS were significantly lower in the experimental groups (P<0.05),ROS was positively correlated with lutein.qRT-PCR examination revealed Nrf2mRNA and HO-1mRNA of the experimental groups were significantly higher compared with the control group (P<0.05),the higher concentrationer of lutein,the more Nrf2mRNA and HO-1mRNA (P<0.05), Caspase-3mRNA of the experimental groups were significantly lower compared with the control group (P<0.05),the higher concentration of lutein,the less Caspase-3mRNA (P<0.05); Western blot examination revealed Nrf2protein of the experimental groups in nuclear were significantly higher compared with the control group (P<0.05),the higher concentration of lutein,the more Nrf2protein in nuclear (P<0.05).There were no significant between Nrf2in cytoplasmic in different groups (P>0.05); HO-1protein of the experimental groups in nuclear were significantly higher compared with the control group (P<0.05), Caspase-3protein of the experimental groups in nuclear were significantly lower compared with the control group (P<0.05), HO-1protein and Nrf2protein in nuclear was positively correlated with lutein, Caspase-3protein was negatively correlated with lutein.Conclusions Lutein can induce the expression of antioxidant enzymes by the way of inducing the expression and nuclear translocation of Nrf2,consequently to reduce the oxidative stress(ROS) status,it decreased the expression of caspase-3and inhibition the apoptosis.