This study examined the effect of copper ions on the proliferation of hepatic stellate cells （HSCs） and the role of oxidative stress in this process in order to gain insight into the mechanism of he- patic fibrosis ...This study examined the effect of copper ions on the proliferation of hepatic stellate cells （HSCs） and the role of oxidative stress in this process in order to gain insight into the mechanism of he- patic fibrosis in Wilson＇s disease. LX-2 cells, a cell line of human HSCs, were cultured in vitro and treated with different agents including copper sulfate, N-acetyl cysteine （NAC） and buthionine sulfoxi- mine （BSO） for different time. The proliferation of LX-2 cells was measured by non-radioactive cell proliferation assay. Real-time PCR and Westem blotting were used to detect the mRNA and protein ex- pression of platelet-derived growth factor receptor 13 subunit （PDGFI3R）, ELISA to determine the level of glutathione （GSH） and oxidized glutathione （GSSG）, dichlorofluorescein assay to measure the level of reactive oxygen species （ROS）, and lipid hydroperoxide assay to quantify the level of lipid peroxide （LPO）. The results showed that copper sulfate over a certain concentration range could promote the pro- liferation of LX-2 cells in a time- and dose-dependent manner. The effect was most manifest when LX-2 cells were treated with copper sulfate at a concentration of 100 ~tmol/L for 24 h. Additionally, copper sulfate could dose-dependently increase the levels of ROS and LPO, and decrease the ratio of GSH/GSSG in LX-2 cells. The copper-induced increase in mRNA and protein expression of PDGF^R was significantly inhibited in LX-2 cells pre-treated with NAC, a precursor of GSH, and this phenome- non could be reversed by the intervention of BSO, an inhibitor of NAC. It was concluded that copper ions may directly stimulate the proliferation of HSCs via oxidative stress. Anti-oxidative stress therapies may help suppress the copper-induced activation and proliferation of HSCs.展开更多
文摘This study examined the effect of copper ions on the proliferation of hepatic stellate cells （HSCs） and the role of oxidative stress in this process in order to gain insight into the mechanism of he- patic fibrosis in Wilson＇s disease. LX-2 cells, a cell line of human HSCs, were cultured in vitro and treated with different agents including copper sulfate, N-acetyl cysteine （NAC） and buthionine sulfoxi- mine （BSO） for different time. The proliferation of LX-2 cells was measured by non-radioactive cell proliferation assay. Real-time PCR and Westem blotting were used to detect the mRNA and protein ex- pression of platelet-derived growth factor receptor 13 subunit （PDGFI3R）, ELISA to determine the level of glutathione （GSH） and oxidized glutathione （GSSG）, dichlorofluorescein assay to measure the level of reactive oxygen species （ROS）, and lipid hydroperoxide assay to quantify the level of lipid peroxide （LPO）. The results showed that copper sulfate over a certain concentration range could promote the pro- liferation of LX-2 cells in a time- and dose-dependent manner. The effect was most manifest when LX-2 cells were treated with copper sulfate at a concentration of 100 ~tmol/L for 24 h. Additionally, copper sulfate could dose-dependently increase the levels of ROS and LPO, and decrease the ratio of GSH/GSSG in LX-2 cells. The copper-induced increase in mRNA and protein expression of PDGF^R was significantly inhibited in LX-2 cells pre-treated with NAC, a precursor of GSH, and this phenome- non could be reversed by the intervention of BSO, an inhibitor of NAC. It was concluded that copper ions may directly stimulate the proliferation of HSCs via oxidative stress. Anti-oxidative stress therapies may help suppress the copper-induced activation and proliferation of HSCs.
