Yu Zhanjiang, Yang Xiaoda, Wang Kui
Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing 100083, PR China.
Toxicology. 2006 Jun 1;223(1-2):1-8. doi: 10.1016/j.tox.2006.02.021. Epub 2006 Apr 3.
The aim of this work is to define the relationship between heat shock protein (HSP) and reactive oxygen species (ROS) in the cells exposed to different concentrations of metal ions, and to evaluate a new method for tracing the dynamic levels of cellular reactive oxygen species using a HSE-SEAP reporter gene. The expression of heat shock protein was measured using a secreted alkaline phosphatase (SEAP) reporter gene transformed into HeLa cell strain, the levels of superoxide anion (O(2)(-)) and hydrogen peroxide (H(2)O(2)) were determined by NBT reduction assay and DCFH staining flow cytometry (FCM), respectively. The experimental results demonstrated that the expression of heat shock protein induced by metal ions was linearly related to the cellular superoxide anion level before cytotoxic effects were observed, but not related to the cellular hydrogen peroxide level. The experimental results suggested that metal ions might induce heat shock protein by elevating cellular superoxide anion level, and thus the expression of heat shock protein indicated by the HSE-SEAP reporter gene can be an effective model for monitoring the dynamic level of superoxide anion and early metal-induced oxidative stress/cytotoxicity.
本研究旨在确定暴露于不同浓度金属离子的细胞中热休克蛋白（HSP）与活性氧（ROS）之间的关系，并评估一种使用HSE-SEAP报告基因追踪细胞活性氧动态水平的新方法。通过将分泌型碱性磷酸酶（SEAP）报告基因转入HeLa细胞系来检测热休克蛋白的表达，分别采用NBT还原法和DCFH染色流式细胞术（FCM）测定超氧阴离子（O(2)(-)）和过氧化氢（H(2)O(2)）的水平。实验结果表明，在观察到细胞毒性效应之前，金属离子诱导的热休克蛋白表达与细胞超氧阴离子水平呈线性相关，但与细胞过氧化氢水平无关。实验结果提示，金属离子可能通过提高细胞超氧阴离子水平来诱导热休克蛋白，因此，由HSE-SEAP报告基因指示的热休克蛋白表达可作为监测超氧阴离子动态水平和早期金属诱导的氧化应激/细胞毒性的有效模型。
