Electron Transport Chain-dependent and -independent Mechanisms of Mitochondrial H2O2 Emission during Long-chain Fatty Acid Oxidation
Electron Transport Chain-dependent and -independent Mechanisms of Mitochondrial H2O2 Emission during Long-chain Fatty Acid Oxidation
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DOI：
10.1074/jbc.m109.026203
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发表时间：
2010-02-19
影响因子：
4.8
通讯作者：
Harper, Mary-Ellen
中科院分区：
文献类型：
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作者：
Seifert, Erin L.;Estey, Carmen;Harper, Mary-Ellen
骨骼肌中的氧化应激是各种病理生理状态的标志，这些病理生理状态也以增加对长链脂肪酸（LCFA）底物的依赖为特征，例如胰岛素抵抗和运动。然而，鲜为人知的是LCFA诱导的活性氧（ROS）在完整的线粒体负担的机制基础，阐明这种机制基础是本研究的目标。具体目标是确定LCFA催化剂与ROS产生相关的程度，并获得相关ROS产生的机制见解。由于LCFA催化剂的中间体和副产物可能干扰抗氧化机制，我们预测LCFA催化剂过程中ROS的形成反映了一个复杂的过程，涉及多个ROS产生位点以及线粒体功能的改变。因此，我们利用了几种互补的方法来探索潜在的机制。使用骨骼肌线粒体，我们的研究结果表明，即使是低供应的LCFA与ROS的形成超过所产生的NADH连接的底物。此外，ROS的产生是明显的跨膜电位的生理范围内，是相对不敏感的膜电位的变化。拓扑结构和膜电位以及使用抑制剂的测定揭示了复合物III和电子转移黄素蛋白（ETF）和ETF-氧化还原酶，作为ROS产生的可能位点。最后，ROS的产生对LCFA分解代谢中间体的基质水平敏感，表明LCFA分解代谢中间体的线粒体输出可以在确定ROS水平中发挥作用。
Oxidative stress in skeletal muscle is a hallmark of various pathophysiologic states that also feature increased reliance on long-chain fatty acid (LCFA) substrate, such as insulin resistance and exercise. However, little is known about the mechanistic basis of the LCFA-induced reactive oxygen species (ROS) burden in intact mitochondria, and elucidation of this mechanistic basis was the goal of this study. Specific aims were to determine the extent to which LCFA catabolism is associated with ROS production and to gain mechanistic insights into the associated ROS production. Because intermediates and by-products of LCFA catabolism may interfere with antioxidant mechanisms, we predicted that ROS formation during LCFA catabolism reflects a complex process involving multiple sites of ROS production as well as modified mitochondrial function. Thus, we utilized several complementary approaches to probe the underlying mechanism(s). Using skeletal muscle mitochondria, our findings indicate that even a low supply of LCFA is associated with ROS formation in excess of that generated by NADH-linked substrates. Moreover, ROS production was evident across the physiologic range of membrane potential and was relatively insensitive to membrane potential changes. Determinations of topology and membrane potential as well as use of inhibitors revealed complex III and the electron transfer flavoprotein (ETF) and ETF-oxidoreductase, as likely sites of ROS production. Finally, ROS production was sensitive to matrix levels of LCFA catabolic intermediates, indicating that mitochondrial export of LCFA catabolic intermediates can play a role in determining ROS levels.