The mechanism of resveratrol regulating iron death through nuclear factor erythroid 2-related factor 2/NAD(P)H:quinone oxidoreductase 1 signaling pathway
Objective To investigate the potential mechanism of resveratrol regulating iron death in HT22 cells of mouse hippocampal neurons by quantitative polymerase chain reaction(qPCR)and Western blotting.Differences between groups were determined by ANOVA and Tukey's post hoc tests.Methods HT22 cell ferroptosis model was induced by Erastin.The groups of control,Erastin,40 and 80 µmol/L RES,and Erastin+RES+ML385 were set up.Cell viability was tested by cell counting kit-8(CCK-8)assay,apoptosis was detected by flow cytometry and terminal-deoxynucleotidyl transferase media-ted nick end labeling(TUNEL),intracellular iron and ROS determined by kits,and Nrf2/NQO1 detected by quantitative polymerase chain reaction(qPCR)and Western blotting.Differences between groups were determined by ANOVA and Tukey's post hoc tests.Results The cell viability of 0.6 µmol/L Erastin group was significantly lower than that of 40μmol/L RES group(51.00±2.90 vs.68.12±2.24,t=2.902,P＜0.05).The apoptosis rate of HT22 cells treated with 80μmol/L RES was significantly lower than that of Erastin group(6.12±0.93 vs.18.21±2.05,t=3.858,P＜0.05).The contents of intra cellular active iron and ROS in 80 µmol/L RES group were significantly lower than those in Erastin group(active iron:43.14±3.13 vs.65.11±4.07,t=3.821,P＜0.05;ROS:8.04±0.83 vs.25.02±1.57,t=5.852,P＜0.05).The mRNA and protein expressions of Nrf2,NQO1 and HO-1 after RES pre-treatment were significantly higher than those in Erastin group(NQO1 mRNA:0.32±0.04 vs.1.01±0.07,t=3.928,P＜0.05;Nrf2 mRNA:0.25±0.03 vs.1.04±0.06,t=3.785,P＜0.05;HO-1 mR-NA:0.26±0.05 vs.1.02±0.08,t=3.957,P＜0.05;NQO1 protein:0.96±0.04 vs.0.45±0.07,t=3.792,P＜0.05;Nrf2 protein:0.91±0.07 vs.0.32±0.05,t=3.957,P＜0.05;HO-1 protein:1.21±0.04 vs.0.25±0.03,t=4.535,P＜0.05),while ML385 reversed the protective effect of RES,resulting in significantly lower expression of NQO1,Nrf2 and HO-1 protein in RES+Erastin+ML385 group than in RES+Erastin group(NQO1:0.67±0.05 vs.0.92±0.08,t=4.276,P＜0.05;Nrf2:0.71±0.05 vs.0.94±0.07,t=4.121,P＜0.05;HO-1:0.64±0.04 vs.1.16±0.04,t=3.978,P＜0.05).Conclusion Resveratrol may inhibit Erastin-induced iron death in HT22 cells by activating Nrf2/NQO1 pathway.
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