Lage O M, Sansonetty F, O'Connor J E, Parente A M
Departamento de Botânica da Faculdade de Ciências and Centro Interdisciplinar de Investigação Marinha e Ambiental, Universidade do Porto, Porto, Portugal.
Cytometry. 2001 Jul 1;44(3):226-35. doi: 10.1002/1097-0320(20010701)44:3<226::aid-cyto1115>3.0.co;2-9.
Copper(II) is a heavy metal whose levels have increased in some marine ecosystems to polluting levels. Dinoflagellates, an important phytoplankton group, are at the base of aquatic food chains and bioaccumulation of copper by these microorganisms can result in complex ecosystem alterations, so we investigated how copper disturbs those cells.
Cytotoxic effects of sublethal and lethal copper concentrations ranging from 4.2 nM (control condition) to 3.13 microM estimated labile copper were studied in batch cultures of Amphidinium carterae. Cell morphology, motility, autofluorescence, and fluorescein diacetate (FDA)-dependent fluorescence generation were evaluated by flow cytometry (FCM) and microscopy.
Exposure of A. carterae to toxic levels of copper impaired cell mobility, delayed cell proliferation, led to increased green autofluorescence, and at 3.13 microM labile copper also induced encystment and death. Chlorophyll fluorescence, however, was not affected. Kinetic FCM assay of FDA-dependent fluorescence generation showed a dose-dependent enhancement of fluorescein fluorescence immediately after copper addition and in cultures with sustained exposure to this toxicant.
Our data suggest that copper toxicity occurs quickly at the membrane level in relation to oxidative stress generation. Based on fluorescence kinetic studies, the Na(+)/H(+) antiporter seemed to be affected by copper, thereby affecting intracellular pH.
铜（II）是一种重金属，其在一些海洋生态系统中的含量已上升至污染水平。甲藻是重要的浮游植物类群，处于水生食物链的底层，这些微生物对铜的生物累积会导致复杂的生态系统变化，因此我们研究了铜如何干扰这些细胞。
在卡特亚得藻的分批培养中，研究了从4.2 nM（对照条件）到3.13 μM的亚致死和致死铜浓度（估计为不稳定铜）的细胞毒性作用。通过流式细胞术（FCM）和显微镜评估细胞形态、运动性、自发荧光以及依赖于荧光素二乙酸酯（FDA）的荧光产生。
将卡特亚得藻暴露于有毒水平的铜会损害细胞运动性、延迟细胞增殖、导致绿色自发荧光增加，并且在3.13 μM不稳定铜时还会诱导形成包囊和死亡。然而，叶绿素荧光不受影响。对依赖于FDA的荧光产生的动力学FCM分析表明，在添加铜后立即以及在持续暴露于这种毒物的培养物中，荧光素荧光呈剂量依赖性增强。
我们的数据表明，铜毒性在膜水平上与氧化应激的产生相关，迅速发生。基于荧光动力学研究，Na(+)/H(+)反向转运蛋白似乎受到铜的影响，从而影响细胞内pH值。
