Environmental Chemistry and Technology Program, University of Wisconsin-Madison, 660 North Park Street, Madison, Wisconsin 53706, USA.
Environ Toxicol. 2013 Sep;28(9):516-23. doi: 10.1002/tox.20743. Epub 2011 Jul 22.
The toxicity of metals to organisms is, in-part, related to the formation of reactive oxygen species (ROS) in cells and subsequent oxidative stress. ROS are by-products of normal respiration and photosynthesis processes in organisms, but environmental factors, like metal exposure, can stimulate excess production. Metals involved in several different mechanisms such as Haber-Weiss cycling and Fenton-type reactions can produce ROS. Some metals, such as Cd, may contribute to oxidative stress indirectly by depleting cellular antioxidants. We investigated the measurement of ROS as a sensitive biomarker of metal toxicity (that could possibly be implemented in a biotic ligand model for algae) and we compared ROS induction in response to several contrasting transition metals (Cu, V, Ni, Zn, and Cd). We also compared the ROS response to glutathione and growth toxicity endpoints measured in a previous study. The cell-permeable dye, 2'7'dichlorodihydrofluorescein diacetate, was used as a probe to detect formation of ROS in Chlamydomonas reinhardtii cells. Metal-exposed cells were incubated with the fluorescent dye in a 96-well plate and monitored over 5.5 h. A dose-response of ROS formation was observed with Cu exposure in the range of 20-500 nM. Cu produced more ROS compared with either Zn or Cd (both nonredox active metals). The redox-active metal V produced increased ROS with increased concentration. The measurement of ROS may be a useful indicator of Cu toxicity, but the signal to noise ratio was better for the glutathione endpoint assay.
金属对生物体的毒性部分与细胞中活性氧（ROS）的形成以及随后的氧化应激有关。ROS 是生物体正常呼吸和光合作用过程的副产品，但环境因素，如金属暴露，会刺激过量产生。参与几种不同机制的金属，如 Haber-Weiss 循环和 Fenton 型反应，可以产生 ROS。一些金属，如 Cd，可以通过耗尽细胞内抗氧化剂间接导致氧化应激。我们研究了 ROS 作为金属毒性敏感生物标志物的测量（可能在藻类的生物配体模型中实施），并比较了几种对比过渡金属（Cu、V、Ni、Zn 和 Cd）对 ROS 诱导的反应。我们还比较了与先前研究中测量的谷胱甘肽和生长毒性终点相关的 ROS 反应。细胞通透性染料 2'7'-二氯二氢荧光素二乙酸酯被用作探针，以检测莱茵衣藻细胞中 ROS 的形成。将暴露于金属的细胞与荧光染料一起在 96 孔板中孵育，并在 5.5 小时内进行监测。在 20-500 nM 的 Cu 暴露范围内观察到 ROS 形成的剂量反应。与非氧化还原活性金属 Zn 或 Cd 相比，Cu 产生了更多的 ROS。随着浓度的增加，氧化还原活性金属 V 产生的 ROS 增加。ROS 的测量可能是 Cu 毒性的有用指标，但谷胱甘肽终点测定的信号噪声比更好。
