Direct speciation analysis of arsenic in sub-cellular compartments using micro-X-ray absorption spectroscopy
Direct speciation analysis of arsenic in sub-cellular compartments using micro-X-ray absorption spectroscopy
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DOI：
10.1016/j.envres.2009.09.006
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发表时间：
2010-07-01
影响因子：
8.3
通讯作者：
Ortega, Richard
中科院分区：
文献类型：
--
作者：
Bacquart, Thomas;Deves, Guillaume;Ortega, Richard
在亚细胞水平上鉴定砷的化学形态是理解砷毒理学和抗肿瘤药理学机制的关键。当使用微束进行X射线吸收近边结构(MU-XANES)时，能够在亚细胞空间中直接进行砷的形态分析，避免了细胞分级和其他可能改变化学形态的制备步骤。这种方法结合了通过共聚焦显微镜或荧光显微镜跟踪单个细胞中的细胞器与通过Mu-XANES对化学物种进行局部分析。在这里，我们报告了用基于Kirkpatrick-Baez X射线聚焦光学的Mu-XANES实验装置所获得的结果，该装置能够在微米空间分辨率(15 x 4.0µm(2))下保持高的入射辐射通量(>；10(11)ph/S)。这一原始的实验装置实现了亚细胞细胞器中砷的直接形态分析，检测下限为10(-15)g。MU-XANES分析表明，As(OH)3是As_2O_3作用下培养的癌细胞胞浆、细胞核和线粒体网络中砷的主要存在形式。另一方面，在暴露于As(OH)(3)的HepG2细胞中观察到As(III)物种的优势，在某些情况下，在HepG2细胞的核结构中氧化成五价形式。对核内混合氧化还原状态的观察表明，细胞群体中存在个体间的差异，这只能通过直接的亚细胞形态分析来证明(C)2009 Elsevier Inc.保留所有权利
Identification of arsenic chemical species at a sub-cellular level is a key to understanding the mechanisms involved in arsenic toxicology and antitumor pharmacology When performed with a microbeam, X-ray absorption near-edge structure (mu-XANES) enables the direct speciation analysis of arsenic in sub-cellular compartments avoiding cell fractionation and other preparation steps that might modify the chemical species. This methodology couples tracking of cellular organelles in a single cell by confocal or epifluorescence microscopy with local analysis of chemical species by mu-XANES. Here we report the results obtained with a mu-XANES experimental setup based on Kirkpatrick-Baez X-ray focusing optics that maintains high flux of incoming radiation (> 10(11) ph/s) at micrometric spatial resolution (15 x 4.0 mu m(2)) This original experimental setup enabled the direct speciation analysis of arsenic in sub-cellular organelles with a 10(-15)g detection limit. mu-XANES shows that inorganic arsenite, As(OH)3, is the main form of arsenic in the cytosol, nucleus, and mitochondrial network of cultured cancer cells exposed to As2O3. On the other hand, a predominance of As(III) species is observed in HepG2 cells exposed to As(OH)(3) with, in some cases, oxidation to a pentavalent form in nuclear structures of HepG2 cells The observation of intra-nuclear mixed redox states suggests an inter-individual variability in a cell population that can only be evidenced with direct sub-cellular speciation analysis (C) 2009 Elsevier Inc All rights reserved