The role of oxidative stress, impaired glycolysis and mitochondrial respiratory redox failure in the cytotoxic effects of 6-hydroxydopamine in vitro
The role of oxidative stress, impaired glycolysis and mitochondrial respiratory redox failure in the cytotoxic effects of 6-hydroxydopamine in vitro
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DOI：
10.1016/j.brainres.2003.12.034
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发表时间：
2004-04-09
期刊：
影响因子：
2.9
通讯作者：
Soliman, KFA
中科院分区：
文献类型：
--
作者：
Mazzio, EA;Reams, RR;Soliman, KFA
神经毒素 6-羟基多巴胺 (6-OHDA) 与帕金森病的神经退行性过程有关。目前的研究旨在阐明 6-OHDA 对神经母细胞瘤 (N-2A) 细胞能量代谢的毒理学影响。 6-OHDA 的毒性相当于厌氧/需氧细胞功能的完全崩溃，这与其他线粒体毒素（例如 MPP+）不同，其目标是有氧代谢的特定丧失。 6-OHDA 的毒性与线粒体氧 (02) 消耗 (MOC)、糖酵解活性、ATP、H+ 离子梯度、膜电位和自氧化产物过氧化氢 (H2O2) 积累的损失平行。用非酶化学计量清除剂（例如羧酸、谷胱甘肽和过氧化氢酶）去除 H2O2 产生了部分保护。用丙酮酸或过氧化氢酶快速去除H2O2仅恢复无氧糖酵解，但没有逆转MOC的损失，表明线粒体损伤与H2O2无关。 6-OHDA 产生的 H2O2 通过脂质过氧化和乳酸脱氢酶抑制导致无氧糖酵解的丧失。 6-OHDA 将氧化细胞色素 c (CYT-C-OX) 维持在还原形式 (CYT-C-RED) 的能力似乎在线粒体损伤中发挥作用。 6-OHDA 对 CYT-C 的还原作用广泛，在几分钟内发生，先于 H2O2 形成，并且不受过氧化氢酶或超氧化物歧化酶的影响。在相似的浓度下，6-OHDA很容易将铁[Fe(III)]的价态改变为Fe(H)，理论上这也将维持还原形式的CYT-C。在分离的线粒体中，6-OHDA 对复合物 1 的影响可以忽略不计，通过维持底物 CYT-C 处于还原状态，抑制复合物 11 并干扰复合物 III。 6-OHDA 导致分离的细胞色素氧化酶（复合物 IV）活性短暂而有效的激增，并由于 6-OHDA 将 CYT-C-OX 回收为 CYT-C-RED 而迅速恢复。典型的线粒体毒素如 MPP+、叠氮化物和抗霉素似乎会抑制 ETC 酶的催化活性。相反，6-OHDA 改变细胞色素的氧化还原，导致底物可用性的损失和氧化还原事件的阻碍。通过将过氧化氢酶与 CYT-C（马心）相结合，实现了针对 6-OHDA 毒性的完全细胞保护并恢复了 MOC。总之，CYT-C 降低特性是儿茶酚胺神经递质所独有的，并且可能在多巴胺能神经元对线粒体损伤的选择性脆弱性中发挥重要作用。 (C) 2004 Elsevier B.V. 保留所有权利。
The neurotoxin, 6-hydroxydopamine (6-OHDA) has been implicated in the neurodegenerative process of Parkinson's disease. The current study was designed to elucidate the toxicological effects of 6-OHDA on energy metabolism in neuroblastoma (N-2A) cells. The toxicity of 6-OHDA corresponds to the total collapse of anaerobic/aerobic cell function, unlike other mitochondrial toxins such as MPP+ that target specific loss of aerobic metabolism. The toxicity of 6-OHDA paralleled the loss of mitochondrial oxygen (02) consumption (MOC), glycolytic activity, ATP, H+ ion gradients, membrane potential and accumulation of the autoxidative product, hydrogen peroxide (H2O2). Removing H2O2 with nonenzymatic stoichiometric scavengers, such as carboxylic acids, glutathione and catalase yielded partial protection. The rapid removal of H2O2 with pyruvate or catalase restored only anaerobic glycolysis, but did not reverse the loss of MOC, indicating mitochondrial impairment is independent of H2O2. The H2O2 generated by 6-OHDA contributed toward the loss of anaerobic glycolysis through lipid peroxidation and lactic acid dehydrogenase inhibition. The ability of 6-OHDA to maintain oxidized cytochrome c (CYT-C-OX) in its reduced form (CYT-C-RED), appears to play a role in mitochondrial impairment. The reduction of CYT-C by 6-OHDA, was extensive, occurred within minutes, preceded formation of H2O2 and was unaffected by catalase or superoxide dismutase. At similar concentrations, 6-OHDA readily altered the valence state of iron [Fe(Ill)] to Fe(H), which would also theoretically sustain CYT-C in its reduced form. In isolated mitochondria, 6-OHDA had negligible effects on complex 1, inhibited complex 11 and interfered with complex III by maintaining the substrate, CYT-C in a reduced state. 6-OHDA caused a transient and potent surge in isolated cytochrome oxidase (complex IV) activity, with rapid recovery as a result of 6-OHDA recycling CYT-C-OX to CYT-C-RED. Typical mitochondrial toxins such as MPP+, azide and antimycin appeared to inhibit the catalytic activity of ETC enzymes. In contrast, 6-OHDA alters the redox of the cytochromes, resulting in loss of substrate availability and obstruction of oxidation-reduction events. Complete cytoprotection against 6-OHDA toxicity and restored MOC was achieved by combining catalase with CYT-C (horse heart). In summary, CYT-C reducing properties are unique to catecholamine neurotransmitters, and may play a significant role in selective vulnerability of dopaminergic neurons to mitochondrial insults. (C) 2004 Elsevier B.V. All rights reserved.