Timur Mujgan, Akbas S Halide, Ozben Tomris
Department of Biochemistry, Faculty of Medicine, Akdeniz University, Antalya, Turkey.
Acta Biochim Pol. 2005;52(4):897-902. Epub 2005 Nov 7.
Topotecan, a semisynthetic water-soluble derivative of camptothecin exerts its cytotoxic effect by inhibiting topoisomerase I and causes double-strand DNA breaks which inhibit DNA function and ultimately lead to cell death. In previous studies it was shown that camptothecin causes ROS formation. The aim of this study was to investigate if Topotecan like camptotecin causes oxidative stress in MCF-7 human breast cancer cell line. Determining the oxidant effect of Topotecan may elucidate a possible alternative mechanism for its cytotoxicity.
MCF-7 cells were cultured and exposed to Topotecan for 24 h at 37 degrees C. The viability of the cells (% of control) was measured using the colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Lipid peroxidation (TBARS), protein oxidation (carbonyl content), sulfhydryl, glutathione (GSH) levels, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities were determined in MCF-7 cells with and without Topotecan incubation.
We found the IC(50) concentration of Topotecan as 0.218 microM in MCF-7 cells. This concentration of Topotecan was used in the incubations of the cells. Our data indicated increased oxidative status, as revealed by increased lipid peroxidation and protein oxidation, and decreased GSH and sulfhydryl levels in MCF-7 cells exposed to Topotecan compared to control cells. In contrast, there was a slight increase in SOD and a significant increase in GPx and catalase activity in MCF-7 cells incubated with Topotecan compared to the control.
These results support our hypothesis that Topotecan increases oxidative stress in MCF-7 cells.
拓扑替康是喜树碱的半合成水溶性衍生物，通过抑制拓扑异构酶I发挥细胞毒性作用，导致双链DNA断裂，从而抑制DNA功能并最终导致细胞死亡。在先前的研究中表明，喜树碱会导致活性氧的形成。本研究的目的是调查拓扑替康是否像喜树碱一样在MCF-7人乳腺癌细胞系中引起氧化应激。确定拓扑替康的氧化作用可能会阐明其细胞毒性的一种可能的替代机制。
培养MCF-7细胞，并在37℃下将其暴露于拓扑替康24小时。使用比色法3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四氮唑(MTT) 测定法测量细胞活力（相对于对照的百分比）。在有或没有拓扑替康孵育的MCF-7细胞中测定脂质过氧化（硫代巴比妥酸反应物）、蛋白质氧化（羰基含量）、巯基、谷胱甘肽(GSH) 水平、超氧化物歧化酶(SOD)、过氧化氢酶(CAT) 和谷胱甘肽过氧化物酶(GPx) 的活性。
我们发现MCF-7细胞中拓扑替康的IC(50) 浓度为0.218 microM。该浓度的拓扑替康用于细胞孵育。我们的数据表明，与对照细胞相比，暴露于拓扑替康的MCF-7细胞中脂质过氧化和蛋白质氧化增加，GSH和巯基水平降低，氧化状态增加。相反，与对照相比，用拓扑替康孵育的MCF-7细胞中SOD略有增加，GPx和过氧化氢酶活性显著增加。
这些结果支持我们的假设，即拓扑替康会增加MCF-7细胞中的氧化应激。
