Sheu K F, Calingasan N Y, Lindsay J G, Gibson G E
Department of Neurology and Neuroscience, Burke Medical Research Institute, Cornell University Medical College, White Plains, New York 10605, USA.
J Neurochem. 1998 Mar;70(3):1143-50. doi: 10.1046/j.1471-4159.1998.70031143.x.
Mitochondrial dysfunction is a common feature of many neurodegenerative disorders. The metabolic encephalopathy caused by thiamine deficiency (TD) is a classic example in which an impairment of cerebral oxidative metabolism leads to selective cell death. In experimental TD in rodents, a reduction in the activity of the thiamine diphosphate-dependent, mitochondrial enzyme alpha-ketoglutarate dehydrogenase complex (KGDHC) occurs before the onset of pathologic lesions and is among the earliest biochemical deficits found. To understand the molecular basis and the significance of the deficiency of KGDHC in TD-induced brain damage, the enzyme activity and protein levels of KGDHC were analyzed. The effect of TD on the subregional/cellular distribution of KGDHC and the anatomic relation of KGDHC with selective cell death were also tested by immunocytochemistry. Consistent with several previous studies, TD dramatically reduced KGDHC activity in both anatomically damaged (thalamus and inferior colliculus) and spared (cerebral cortex) regions. Immunocytochemistry revealed no apparent correlation of regional KGDHC immunoreactivity or its response to TD with affected regions in TD. The basis of the enzymatic and immunocytochemical behavior of KGDHC was further assessed by quantitative immunoblots, using antibodies specific for each of the three KGDHC components. Despite the marked decrease of KGDHC activity in TD, no reduction of any of the three KGDHC protein levels was found. Thus, TD impairs the efficacy of the KGDHC catalytic machinery, whereas the concentration of protein molecules persists. The generalized decline of KGDHC activity with no apparent anatomic selectivity is consistent with the notion that the compromised mitochondrial oxidation sensitizes the brain cells to various other insults that precipitate the cell death. The current TD model provides a relevant experimental system to understand the molecular basis of many neurodegenerative conditions in which mitochondrial dysfunction and KGDHC deficiency are prominent features.
线粒体功能障碍是许多神经退行性疾病的共同特征。硫胺素缺乏（TD）引起的代谢性脑病就是一个典型例子，其中脑氧化代谢受损导致选择性细胞死亡。在啮齿动物的实验性TD中，硫胺素二磷酸依赖性线粒体酶α-酮戊二酸脱氢酶复合体（KGDHC）的活性在病理损伤出现之前就已降低，并且是最早发现的生化缺陷之一。为了了解TD诱导的脑损伤中KGDHC缺乏的分子基础和意义，对KGDHC的酶活性和蛋白质水平进行了分析。还通过免疫细胞化学检测了TD对KGDHC亚区域/细胞分布的影响以及KGDHC与选择性细胞死亡的解剖学关系。与先前的几项研究一致，TD显著降低了解剖学上受损区域（丘脑和下丘）以及未受损区域（大脑皮层）的KGDHC活性。免疫细胞化学显示，区域KGDHC免疫反应性或其对TD的反应与TD中的受影响区域没有明显相关性。使用针对KGDHC三个组分各自的特异性抗体，通过定量免疫印迹进一步评估了KGDHC的酶促和免疫细胞化学行为基础。尽管TD中KGDHC活性显著降低，但未发现三种KGDHC蛋白质水平中的任何一种降低。因此，TD损害了KGDHC催化机制的功效，而蛋白质分子的浓度持续存在。KGDHC活性普遍下降且无明显解剖学选择性，这与线粒体氧化受损使脑细胞对各种其他促使细胞死亡的损伤敏感的观点一致。当前的TD模型提供了一个相关的实验系统，以了解许多神经退行性疾病的分子基础，其中线粒体功能障碍和KGDHC缺乏是突出特征。
