Cai Xiaoqiang, Pan Nina, Zou Guolin
State Key Laboratory of Virology, Department of Biotechnology, College of Life Sciences, Wuhan University, Wuhan, 430072, China.
Biometals. 2007 Feb;20(1):1-11. doi: 10.1007/s10534-006-9008-0. Epub 2006 May 9.
The mechanism of cytotoxicity on liver carcinoma Bel-7402 cells induced by copper-1,10-phenanthroline, Cu(OP)2, has been studied. Cell viability and apoptotic rate were examined in cells treated with Cu(OP)2 or Cu2+ alone. It was found that the apoptosis induced by Cu(OP)2 could not be induced by Cu2+ or OP alone in our experimental conditions. Total copper content in cells was measured by atomic absorption spectrophotometry, and the abnormal elevation of intracellular copper transported by lipophilic OP ligand may play the role of initial factor in the apoptosis, which caused subsequent redox state changes in cells. Intracellular levels of reactive oxygen species (ROS) were detected by fluorescent probe 2',7'-dichlorofluorescein diacetate (DCFH-DA). Reduced (GSH) and total glutathione (GSSG + GSH) were determined by High-performance liquid chromatography (HPLC) after derivatization, and the ratios of GSH/GSSG were subsequently calculated. The overproduction of ROS and the decreased GSH/GSSG ratio were observed in cells which represented the occurrence of oxidative stress in the apoptosis. Oxidative DNA damage was also found in cells treated with Cu(OP)2 in the early stage of the apoptosis, and it suggests that the activation of DNA repair system may be involved in the pathway of the apoptosis induced by Cu(OP)2.
研究了1,10 - 菲啰啉铜（Cu(OP)₂）对肝癌Bel - 7402细胞的细胞毒性作用机制。分别检测了单独用Cu(OP)₂或Cu²⁺处理的细胞的活力和凋亡率。结果发现，在我们的实验条件下，单独的Cu²⁺或OP不能诱导出Cu(OP)₂所诱导的细胞凋亡。用原子吸收分光光度法测定细胞中的总铜含量，亲脂性OP配体转运的细胞内铜异常升高可能在细胞凋亡中起初始因子的作用，进而导致细胞随后的氧化还原状态变化。用荧光探针2',7'-二氯二氢荧光素二乙酸酯（DCFH - DA）检测细胞内活性氧（ROS）水平。衍生化后用高效液相色谱法（HPLC）测定还原型谷胱甘肽（GSH）和总谷胱甘肽（GSSG + GSH），随后计算GSH/GSSG比值。在细胞中观察到ROS的过量产生和GSH/GSSG比值的降低，这表明细胞凋亡过程中发生了氧化应激。在用Cu(OP)₂处理的细胞凋亡早期还发现了氧化性DNA损伤，这表明DNA修复系统的激活可能参与了Cu(OP)₂诱导的细胞凋亡途径。
