Rescue from galactose-induced death of Leigh Syndrome patient cells by pyruvate and NAD().
Rescue from galactose-induced death of Leigh Syndrome patient cells by pyruvate and NAD().
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DOI：
10.1038/s41419-018-1179-4
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发表时间：
2018-11-14
影响因子：
9
通讯作者：
Koopman WJH
中科院分区：
文献类型：
--
作者：
Iannetti EF;Smeitink JAM;Willems PHGM;Beyrath J;Koopman WJH
线粒体复合物 I (CI) 缺陷的细胞模型显示糖酵解激活，以补偿线粒体 ATP 产生的损失。这种适应可以掩盖细胞生理学中其他相关的缺陷引起的畸变。在这里，我们研究了患有孤立性 CI 缺陷的 Leigh 综合征 (LS) 患者的原代皮肤成纤维细胞的活力、线粒体形态功能、ROS 水平和 ATP 稳态。这些细胞系的核 DNA (nDNA) 编码的 CI 基因（NDUFS7、NDUFS8、NDUFV1）中存在突变，为了防止糖酵解上调，在不含丙酮酸的培养基中培养，其中葡萄糖被半乳糖取代。优化细胞培养方案后，LS 成纤维细胞在半乳糖培养基中死亡，而对照细胞则没有。丙酮酸、苹果酸、草酰乙酸、α-酮戊二酸、天冬氨酸和外源 NAD+ (eNAD) 可剂量依赖性地抑制 LS 细胞死亡，但乳酸、琥珀酸、α-酮丁酸和尿苷则不受抑制。丙酮酸和 eNAD 在不同程度上增加了半乳糖处理的 LS 细胞中的细胞 NAD+ 含量，共孵育研究表明丙酮酸诱导的拯救主要不是由 NAD+ 介导的。从功能上讲，在 LS 细胞中，葡萄糖-半乳糖替代增加了线粒体碎片和质量，使线粒体膜电位 (Δψ) 去极化，增加了 H2DCFDA 氧化 ROS 水平，增加了线粒体 ATP 生成，并减少了细胞 ATP 总含量。这些畸变被丙酮酸和 eNAD 不同程度地拯救，支持了这些化合物通过不同机制拯救半乳糖诱导的 LS 细胞死亡的结论。这些发现建立了基于细胞的干预测试策略，并增强了我们对 CI 缺陷病理生理学的理解。
Cell models of mitochondrial complex I (CI) deficiency display activation of glycolysis to compensate for the loss in mitochondrial ATP production. This adaptation can mask other relevant deficiency-induced aberrations in cell physiology. Here we investigated the viability, mitochondrial morphofunction, ROS levels and ATP homeostasis of primary skin fibroblasts from Leigh Syndrome (LS) patients with isolated CI deficiency. These cell lines harbored mutations in nuclear DNA (nDNA)-encoded CI genes (NDUFS7, NDUFS8, NDUFV1) and, to prevent glycolysis upregulation, were cultured in a pyruvate-free medium in which glucose was replaced by galactose. Following optimization of the cell culture protocol, LS fibroblasts died in the galactose medium, whereas control cells did not. LS cell death was dose-dependently inhibited by pyruvate, malate, oxaloacetate, α-ketoglutarate, aspartate, and exogenous NAD+ (eNAD), but not by lactate, succinate, α-ketobutyrate, and uridine. Pyruvate and eNAD increased the cellular NAD+ content in galactose-treated LS cells to a different extent and co-incubation studies revealed that pyruvate-induced rescue was not primarily mediated by NAD+. Functionally, in LS cells glucose-by-galactose replacement increased mitochondrial fragmentation and mass, depolarized the mitochondrial membrane potential (Δψ), increased H2DCFDA-oxidizing ROS levels, increased mitochondrial ATP generation, and reduced the total cellular ATP content. These aberrations were differentially rescued by pyruvate and eNAD, supporting the conclusion that these compounds rescue galactose-induced LS cell death via different mechanisms. These findings establish a cell-based strategy for intervention testing and enhance our understanding of CI deficiency pathophysiology.
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影响因子：
5.5
作者：
Koopman, Werner J. H.;Verkaart, Sjoerd;Willems, Peter H. G. M.
通讯作者：
Willems, Peter H. G. M.
DOI：
10.1152/ajprenal.2000.278.5.f758
发表时间：
2000-05-01
影响因子：
4.2
作者：
Bergin, E;Levine, JS;Lieberthal, W
通讯作者：
Lieberthal, W
影响因子：
4.8
作者：
Christensen, Caspar Elo;Karlsson, Magnus;Lerche, Mathilde H.
通讯作者：
Lerche, Mathilde H.
DOI：
10.1038/nrneurol.2013.129
发表时间：
2013-08
期刊：
Nature reviews. Neurology
影响因子：
--
作者：
Pfeffer G;Horvath R;Klopstock T;Mootha VK;Suomalainen A;Koene S;Hirano M;Zeviani M;Bindoff LA;Yu-Wai-Man P;Hanna M;Carelli V;McFarland R;Majamaa K;Turnbull DM;Smeitink J;Chinnery PF
通讯作者：
Chinnery PF
影响因子：
14.8
作者：
Iannetti, Eligio F.;Smeitink, Jan A. M.;Koopman, Werner J. H.
通讯作者：
Koopman, Werner J. H.