Ginger Compound -Shogaol and Its Cysteine-conjugated Metabolite (M2) Activate Nrf2 in Colon Epithelial Cells in Vitro and in Vivo. Chem Res Toxicol. 2014 Aug 22, Chen H, Fu J, Chen H, Hu Y, Soroka DN, Prigge J, Schmidt EE, Yan F, Major MB, Chen XL, Sang S.
In this study, we identified Nrf2 as a molecular target of -shogaol (6S), a bioactive compound isolated from ginger, in colon epithelial cells in vitro and in vivo. Following 6S treatment of HCT-116 cells, the intracellular GSH/GSSG ratio was initially diminished but then elevated above the basal level. Intracellular reactive oxygen species (ROS) correlated inversely with the GSH/GSSG ratio. Further analysis using gene microarray showed that 6S up-regulated the expression of Nrf2 target genes (AKR1B10, FTL, GGTLA4 and HMOX1) in HCT-116 cells. Western blotting confirmed up-regulation, phosphorylation and nuclear translocation of Nrf2 protein followed by Keap1 decrease and up-regulation of Nrf2 target genes (AKR1B10, FTL, GGTLA4, HMOX1, and MT1) and glutathione synthesis genes (GCLC, GCLM). Pretreatment of cells with a specific inhibitor of p38 (SB202190), PI3K (LY294002), or MEK1 (PD098059) attenuated these effects of 6S. Using ultrahigh performance liquid chromatography-tandem mass spectrometry, we found that 6S modified multiple cysteine residues of Keap1 protein. In vivo 6S treatment induced Nrf2 nuclear translocation and significantly up-regulated the expression of MT1, HMOX1 and GCLC in the colon of wild-type (WT) mice, but not Nrf2-/- mice. Similar to 6S, a cysteine-conjugated metabolite of 6S (M2), which was previously found to be a carrier of 6S in vitro and in vivo, also activated Nrf2. Our data demonstrated that 6S and its cysteine-conjugated metabolite M2 activate Nrf2 in colon epithelial cells in vitro and in vivo through Keap1-dependent and -independent manners.
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