Menzies Fiona M, Cookson Mark R, Taylor Robert W, Turnbull Douglass M, Chrzanowska-Lightowlers Zofia M A, Dong Lichun, Figlewicz Denise A, Shaw Pamela J
Academic Neurology Unit, University of Sheffield, UK.
Brain. 2002 Jul;125(Pt 7):1522-33. doi: 10.1093/brain/awf167.
The molecular mechanisms by which mutations in the gene for Cu/Zn superoxide dismutase (SOD1) lead to the selective death of motor neurones in familial amyotrophic lateral sclerosis (FALS) remain incompletely understood. Previous evidence has indicated that mitochondrial abnormalities may develop during motor neurone injury, but several important questions remain unanswered. We have developed a cell culture model of FALS in which a motor neurone cell line (NSC34) has been stably transfected to express normal or mutant human SOD1 at levels approximating to those seen in the human disease. The aims of the study were to: (i) investigate whether morphological mitochondrial abnormalities occur at expression levels of mutant SOD1 close to physiological levels; and (ii) determine whether the presence of mutant SOD1 causes abnormalities of mitochondrial respiratory chain function and changes in cellular bioenergetic parameters in motor neuronal cells. Using this cellular model, we demonstrate that the presence of mutant SOD1 results in the development of abnormally swollen and pale staining mitochondria. These morphological changes are accompanied by biochemical abnormalities with specific decreases in the activities of complexes II and IV of the mitochondrial electron transfer chain. These same complexes are inhibited when control NSC34 cells are subjected to oxidative stress induced by serum withdrawal. The decrease in respiratory chain complex activity in the presence of mutant SOD1 was not accompanied by decreased expression of representative proteins present in these complexes. Motor neuronal cells expressing mutant SOD1 showed increased cell death when exposed to oxidative stress by serum withdrawal, whereas the presence of normal human SOD1 exerted a protective effect. Under basal, unstressed culture conditions, no change in the ATP : ADP ratio was observed in the presence of mutant SOD1. However, the mitochondrial changes associated with the presence of mutant SOD1 clearly had adverse cellular bioenergetic consequences as shown by increased cell death in the presence of pharmacological inhibition of the glycolytic pathway. We conclude that one important mechanism by which mutant SOD1 causes motor neurone injury involves inhibition of specific components of the mitochondrial electron transfer chain. Therapeutic measures aimed at protecting mitochondrial respiratory chain function may be useful in SOD1 related familial and possibly other forms of amyotrophic lateral sclerosis.
铜/锌超氧化物歧化酶（SOD1）基因突变导致家族性肌萎缩侧索硬化症（FALS）运动神经元选择性死亡的分子机制仍未完全明确。先前的证据表明，运动神经元损伤期间可能会出现线粒体异常，但几个重要问题仍未得到解答。我们建立了一种FALS细胞培养模型，其中运动神经元细胞系（NSC34）已被稳定转染，以表达接近人类疾病中所见水平的正常或突变型人类SOD1。本研究的目的是：（i）研究在接近生理水平的突变型SOD1表达水平下是否会出现形态学线粒体异常；（ii）确定突变型SOD1的存在是否会导致运动神经元细胞中线粒体呼吸链功能异常以及细胞生物能量参数的变化。使用该细胞模型，我们证明突变型SOD1的存在会导致线粒体异常肿胀和染色变淡。这些形态学变化伴随着生化异常，线粒体电子传递链复合物II和IV的活性出现特定下降。当对照NSC34细胞受到血清剥夺诱导的氧化应激时，这些相同的复合物也会受到抑制。在存在突变型SOD1的情况下，呼吸链复合物活性的下降并未伴随着这些复合物中代表性蛋白质表达的降低。表达突变型SOD1的运动神经元细胞在受到血清剥夺引起的氧化应激时，细胞死亡增加，而正常人类SOD1的存在则发挥了保护作用。在基础的、无应激的培养条件下，在存在突变型SOD1的情况下未观察到ATP:ADP比值的变化。然而，与突变型SOD1存在相关的线粒体变化显然对细胞生物能量产生了不利影响，这表现为在糖酵解途径受到药理学抑制时细胞死亡增加。我们得出结论，突变型SOD1导致运动神经元损伤的一个重要机制涉及抑制线粒体电子传递链的特定成分。旨在保护线粒体呼吸链功能的治疗措施可能对SOD1相关的家族性以及可能其他形式的肌萎缩侧索硬化症有用。
