Phase Ⅱ enzymes activity induced by curcumin accompanying translocation of Nrf2 and activation of ARE binding activity

摘要

Aim To investigate the effect of curcumin on induction of glutathione S-transferases (GST), NADP(H):quinone oxidoreductase (NQO) and explore their possible molecular mechanism. Methods The activity of GST, NQO and cellular reduced glutathione (GSH) content were measured by spectrophotometrical methods. Cellular changes in the distribution of NF-E2 related factor 2 (Nrf2) was detected by western blotting analysis. Nrf2-AREs (antioxidant-responsive elements) binding activity was examined by electrophoretic mobility shift assay (EMSA). Results Curcumin caused a significant increase in GST and NQO activity, with a maximal value being observed at 20, 25 μmol/L, respectively. At concentration higher than 30 μmol/L, curcumin appeared to be less effective in stimulating GST and NQO activity. Treatment of HT-29 with curcumin caused a modest increase in the cellular GSH content at 3 h (P<0.05). All concentrations of curcumin continued to increase GSH content during the next 9 h. No significant change in the GSH content was observed at either 1 or 2 h after curcumin exposure. Upon treatment of cells with 20 μmol/L curcumin, Nrf2 protein accumulated rapidly (3 h) in the nucleus. This data suggested that curcumin mediate an increase of Nrf2 in the nuclear, a common component of ARE binding complexes. The retarded Nrf2-AREs complexes progressively formed upon treatment with curcumin at the concentration of 20 μmol/L for indicated time. Conclusion These results demonstrated that induction of GST and NQO activity by curcumin may be mediated by translocation of transcription factor Nrf2 from cytoplasm to nuclear and increased binding activity of Nrf2-ARE complexes.It is possible that this effect efficiently protects cells from oxidative stress and should be evaluated as a new therapeutic approach in oxidative stress-mediated damage.