论文部分内容阅读
目的评估8-羟基喹啉酮(CuQ)对HepG2细胞的DNA损伤作用并阐明其可能的作用机制。方法 CuQ 0～4μmol.L-1处理HepG2细胞不同时间后,通过单细胞凝胶电泳实验检测细胞DNA损伤;分光光度法测定过氧化氢酶活性;苯二醛法测定细胞内谷胱甘肽(GSH)水平;硫代巴比妥酸反应物(TBARS)法检测细胞内脂质过氧化水平;Western印迹法检测NF-κB p65的变化;免疫组化方法检测细胞内8-羟基脱氧鸟苷(8-OHdG)的表达水平。结果 HepG2细胞与CuQ 0.5～4μmol.L-1作用1 h后,DNA的迁移距离明显增加(P<0.05),提示CuQ可引起DNA链断裂。CuQ能够造成细胞内GSH水平以及过氧化氢酶活性的降低。随着CuQ剂量的增加及染毒时间的延长,NF-κB由细胞浆逐渐转移至细胞核。CuQ还可以引起细胞内TBARS水平增高及8-OHdG表达水平的增强。采用GSH合成特异抑制剂DL-甲硫氨酸磺酰亚胺(BSO)预处理细胞,可明显增强CuQ对HepG2细胞DNA的损伤(P<0.05)。结论 CuQ可造成HepG2细胞氧化性DNA损伤,其作用机制与氧化应激及NF-κB p65在细胞核蓄积增高有关。
Objective To evaluate the DNA damage effect of 8-hydroxyquinolinone (CuQ) on HepG2 cells and elucidate its possible mechanism. Methods After HepG2 cells were treated with CuQ 0 ~ 4μmol.L-1 for different time, DNA damage was detected by single cell gel electrophoresis. The activity of catalase was determined by spectrophotometry. The content of glutathione GSH) were detected by enzyme-linked immunosorbent assay (ELISA). The levels of lipid peroxidation in cells were detected by TBARS assay. The changes of NF-κB p65 were detected by Western blotting and the levels of 8-hydroxydeoxyguanosine 8-OHdG) expression levels. Results The migration distance of DNA in HepG2 cells treated with CuQ 0.5 ~ 4μmol.L-1 for 1 h increased significantly (P <0.05), suggesting that CuQ can cause DNA strand breaks. CuQ can cause a decrease in intracellular GSH levels and catalase activity. With the increase of CuQ dose and exposure time, NF-κB gradually transferred from the cytoplasm to the nucleus. CuQ can also cause increased levels of intracellular TBARS and 8-OHdG expression levels. Pretreatment of cells with GSH-specific inhibitor DL-methionine sulfonylimide (BSO) significantly enhanced the DNA damage of HepG2 cells induced by CuQ (P <0.05). Conclusion CuQ can cause oxidative DNA damage in HepG2 cells, and its mechanism is related to oxidative stress and the increase of nuclear accumulation of NF-κB p65.