Reactive Oxygen Species Produced by Mutated Mitochondrial Respiratory Chains of Entire Cells Monitored Using Modified Microelectrodes,ChemElectroChem

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Reactive Oxygen Species Produced by Mutated Mitochondrial Respiratory Chains of Entire Cells Monitored Using Modified Microelectrodes
ChemElectroChem ( IF 3.5 ) Pub Date : 2019-01-22 , DOI: 10.1002/celc.201801424
Marco Malferrari ₁ , Anna Ghelli ₂ , Francesco Roggiani ₁ , Giovanni Valenti ₁ , Francesco Paolucci ₁ , Michela Rugolo ₂ , Stefania Rapino ₁

Affiliation

Genetic alterations affecting subunits of the mitochondrial respiratory chain complexes often impair their catalytic activities and result in enhanced production of reactive oxygen species (ROS). An electrochemical setup was employed to quantify mitochondrial ROS production in plasma membrane‐permeabilized cellular models of two genetic diseases: the Δcytb cell line bearing a microdeletion in the mitochondrial MT‐CYB gene causing a severe encephalomyopathy and the RJ206 cell line, harbouring a pathogenic mutation associated with Leber's hereditary optic neuropathy. The responses of black platinum modified microelectrodes to the most common cellular redox buffers, namely, NADH and glutathione, as well as substrates deriving from the oxidative metabolism of glucose, were investigated; a relatively high sensitivity, although lower than that for ROS, was shown for NADH. Time‐resolved amperometric measurements of ROS production upon respiratory chain activation at high NADH/NAD⁺ ratio revealed a 50 % and 100 % increase of ROS in cells bearing defective complex I and complex III, respectively, as compared to wild type cells.

中文翻译：

修饰的微电极监测整个细胞的线粒体呼吸链突变产生的活性氧

影响线粒体呼吸链复合物亚基的遗传改变通常会损害其催化活性，并导致活性氧（ROS）产生增加。在两种遗传疾病的质膜透化细胞模型中，采用电化学装置来定量线粒体ROS的产生：Δcytb细胞系在线粒体MT-CYB中具有微缺失该基因引起严重的脑脊髓病和RJ206细胞系，具有与Leber遗传性视神经病变相关的致病突变。研究了黑色铂修饰的微电极对最常见的细胞氧化还原缓冲液（NADH和谷胱甘肽）以及源自葡萄糖氧化代谢的底物的响应。对于NADH，显示出相对较高的灵敏度，尽管低于对ROS的灵敏度。与野生型细胞相比，在高NADH / NAD ⁺比率下呼吸链激活后时间产生的ROS的时间分辨安培测量显示，带有缺陷复合物I和复合物III的细胞中ROS分别增加了50％和100％。

更新日期：2019-01-22

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