Glutathione modulation influences methyl mercury induced neurotoxicity in primary cell cultures of neurons and astrocytes
Glutathione modulation influences methyl mercury induced neurotoxicity in primary cell cultures of neurons and astrocytes
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DOI：
10.1016/j.neuro.2006.01.010
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发表时间：
2006-07-01
期刊：
影响因子：
3.4
通讯作者：
Syversen, Tore
中科院分区：
文献类型：
--
作者：
Kaur, Parvinder;Aschner, Michael;Syversen, Tore
甲基汞（MeHg）具有高度神经毒性，可导致多种神经退行性疾病。在这项研究中，我们研究了谷胱甘肽（GSH）和活性氧（ROS）在甲基汞诱导的神经毒性的作用，使用小脑神经元和星形胶质细胞的原代细胞培养。为了评价GSH对甲基汞诱导的细胞毒性的影响，使用荧光指示剂二氯二氢荧光素二乙酸酯的氯甲基衍生物（CMH（2）DCFDA）和一氯二亚胺（MCB）测量ROS和GSH。用C-14-放射性标记的甲基汞测量细胞相关的甲基汞。MTT法检测线粒体脱氢酶活性。MTT时间轴法研究甲基汞暴露浓度和暴露时间对细胞增殖的影响。用N-乙酰半胱氨酸（NAC）或马来酸二乙酯（DEM）预处理细胞12 h，以改变细胞内GSH含量。用5 μ M MeHg处理30分钟导致活性氧显着增加（p < 0.05）和还原型谷胱甘肽含量降低（p < 0.001）。消耗细胞内GSH的DEM进一步增加了甲基汞诱导的ROS在两种细胞培养物中的产生。相反，NAC的补充增加细胞内GSH，并提供对甲基汞诱导的氧化应激在两种细胞培养物的保护。MTT研究也证实了NAC补充剂在减弱甲基汞诱导的细胞毒性方面的功效。DEM处理后细胞内MeHg含量显著增加（p < 0.02）。总之，GSH的耗竭增加了甲基汞的积累，并增强了甲基汞诱导的氧化应激，相反，补充GSH前体保护免受甲基汞暴露在体外。(c)2006年爱思唯尔公司All rights reserved.
Methyl mercury (MeHg) is highly neurotoxic and may lead to numerous neurodegenerative disorders. In this study, we investigated the role of glutathione (GSH) and reactive oxygen species (ROS) in MeHg-induced neurotoxicity, using primary cell cultures of cerebellar neurons and astrocytes. To evaluate the effect of GSH on MeHg-induced cytotoxicity, ROS and GSH were measured using the fluorescent indicators chloro methyl derivative of di-chloro di-hydro fluorescein diacetate (CMH(2)DCFDA) and monochlorobimane (MCB). Cell-associated MeHg was measured with C-14-radiolabeled MeHg. Mitochondrial dehydrogenase activity was detected by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide]. MTT timeline study was also performed to evaluate the effects of both the concentration and duration of MeHg exposure. The intracellular GSH content was modified by pretreatment with N-acetyl cysteine (NAC) or di-ethyl maleate (DEM) for 12 h. Treatment with 5 mu M MeHg for 30 min led to significant (p < 0.05) increase in ROS and reduction (p < 0.001) in GSH content. Depletion of intracellular GSH by DEM further increased the generation of MeHg-induced ROS in both cell cultures. Conversely, NAC supplementation increased intracellular GSH and provided protection against MeHg-induced oxidative stress in both cell cultures. MTT studies also confirmed the efficacy of NAC supplementation in attenuating MeHg-induced cytotoxicity. The cell-associated MeHg was significantly (p < 0.02) increased after DEM treatment. In summary, depletion of GSH increases MeHg accumulation and enhances MeHg-induced oxidative stress, and conversely, supplementation with GSH precursor protects against MeHg exposure in vitro. (c) 2006 Elsevier Inc. All rights reserved.