AIM To explore the effect of interleukin(IL)-22 on in vitro model of alcoholic liver fibrosis hepatic stellate cells(HSCs), and whether this is related to regulation of Nrf2-keap1-ARE.METHODS HSC-T6 cells were incubat...AIM To explore the effect of interleukin(IL)-22 on in vitro model of alcoholic liver fibrosis hepatic stellate cells(HSCs), and whether this is related to regulation of Nrf2-keap1-ARE.METHODS HSC-T6 cells were incubated with 25, 50, 100, 200 and 400 μmol/L acetaldehyde. After 24 and 48 h, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay was used to detect proliferation of HSCs to choose the best concentration and action time. We used the optimal concentration of acetaldehyde(200 μmol/L) to stimulate HSCs for 24 h, and treated the cells with a final concentration of 10, 20 or 50 ng/m L IL-22. The cell proliferation rate was detected by MTT assay. The cell cycle was analyzed by flow cytometry. The expression of nuclear factor-related factor(Nrf)2 and α-smooth muscle antigen was detected by western blotting and immunocytochemistry. The levels of malondialdehyde(MDA) and glutathione(GSH) were measured by spectrophotometry. RESULTS In the MTT assay, when HSCs were incubated with acetaldehyde, activity and proliferation were higher than in the control group, and were most obvious after 48 h treatment with 200 μmol/L acetaldehyde. The number of cells in G0/G1 phases was decreased and the number in S phase was increased in comparison with the control group. When treated with different concentrations of IL-22, HSC-T6 cell activity and proliferation rate were markedly decreased in a dosedependent manner, and cell cycle progression was arrested from G1 to S phase. Western blotting and immunocytochemistry demonstrated that expression of Nrf2 total protein was not significantly affected. Expression of Nrf2 nuclear protein was low in thecontrol group, increased slightly in the model group(or acetaldehyde-stimulated group), and increased more obviously in the IL-22 intervention groups. The levels of MDA and GSH in the model group were significantly enhanced in comparison with those in the control group. In cells treated with IL-22, the MDA level was attenuated but the GSH level was further increased. These changes were dose-dependent. CONCLUSION IL-22 inhibits acetaldehyde-induced HSC activation and proliferation, which may be related to nuclear translocation of Nrf2 and increased activity of the antioxidant axis Nrf2-keap1-ARE.展开更多
文摘AIM To explore the effect of interleukin(IL)-22 on in vitro model of alcoholic liver fibrosis hepatic stellate cells(HSCs), and whether this is related to regulation of Nrf2-keap1-ARE.METHODS HSC-T6 cells were incubated with 25, 50, 100, 200 and 400 μmol/L acetaldehyde. After 24 and 48 h, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT) assay was used to detect proliferation of HSCs to choose the best concentration and action time. We used the optimal concentration of acetaldehyde(200 μmol/L) to stimulate HSCs for 24 h, and treated the cells with a final concentration of 10, 20 or 50 ng/m L IL-22. The cell proliferation rate was detected by MTT assay. The cell cycle was analyzed by flow cytometry. The expression of nuclear factor-related factor(Nrf)2 and α-smooth muscle antigen was detected by western blotting and immunocytochemistry. The levels of malondialdehyde(MDA) and glutathione(GSH) were measured by spectrophotometry. RESULTS In the MTT assay, when HSCs were incubated with acetaldehyde, activity and proliferation were higher than in the control group, and were most obvious after 48 h treatment with 200 μmol/L acetaldehyde. The number of cells in G0/G1 phases was decreased and the number in S phase was increased in comparison with the control group. When treated with different concentrations of IL-22, HSC-T6 cell activity and proliferation rate were markedly decreased in a dosedependent manner, and cell cycle progression was arrested from G1 to S phase. Western blotting and immunocytochemistry demonstrated that expression of Nrf2 total protein was not significantly affected. Expression of Nrf2 nuclear protein was low in thecontrol group, increased slightly in the model group(or acetaldehyde-stimulated group), and increased more obviously in the IL-22 intervention groups. The levels of MDA and GSH in the model group were significantly enhanced in comparison with those in the control group. In cells treated with IL-22, the MDA level was attenuated but the GSH level was further increased. These changes were dose-dependent. CONCLUSION IL-22 inhibits acetaldehyde-induced HSC activation and proliferation, which may be related to nuclear translocation of Nrf2 and increased activity of the antioxidant axis Nrf2-keap1-ARE.
