Oxidative stress is recognised as central in a range of neurological diseases including Amyotrophic lateral sclerosis (ALS), a disease characterised by fast progressing death of motor neurons in the brain and spinal cord. Cellular pathology includes cytosolic protein aggregates in motor neurons and glia of which potentially cytotoxic hyper-phosphorylated fragments of the Transactive response DNA Binding Protein 43 kDa (TDP-43) constitute a major component. This is closely associated with an additional loss of nuclear TDP-43 expression indicating a “loss of function” mechanism, accelerating motor neuron (MN) loss. Furthermore, mutations in TDP-43 cause familial ALS and ALS-like disease in animal models. In this study, we investigated the role of glutathione (GSH) in modulating oxidative stress responses in TDP-43 pathology in motor neuron NSC-34 cells. Results demonstrate that depletion of GSH produces pathology similar to that of mutant TDP-43, including occurrence of cytosolic aggregates, TDP-43 phosphorylation and nuclear clearing of endogenous TDP-43. We also demonstrate that introduction of mutant TDP-43^A315T and silencing of endogenous TDP-43, but not overexpression of wild-type TDP-43, result in similar pathology, including depletion of intracellular GSH, possibly resulting from a decreased expression of a regulatory subunit of ɣ-glutamylcysteine ligase (GCLM), a rate limiting enzyme in GSH synthesis. Importantly, treatment of mutant cells with GSH monoethyl ester (GSHe) that directly increases intracellular GSH and bypasses the need for GSH synthesis, protected against mutant-induced TDP-43 pathology, including reducing aggregate formation, nuclear clearance, reactive oxygen species (ROS) production and cell death. Our data strongly suggest that oxidative stress is central to TDP-43 pathology and may result from a loss of function affecting GSH synthesis and that treatments directly aimed at restoring cellular GSH content may be beneficial in preventing cell death in TDP-43-mediated ALS.

氧化应激被认为是一系列神经系统疾病的中心，包括肌萎缩性侧索硬化症（ALS），该疾病的特征是大脑和脊髓中运动神经元快速死亡。细胞病理学包括运动神经元和神经胶质中的胞质蛋白聚集体，其潜在的细胞毒性超磷酸化片段是Transactive Response DNA结合蛋白43 kDa（TDP-43）的主要成分。这与核TDP-43表达的额外丧失密切相关，表明TDP-43的核表达“功能丧失”机制，加速了运动神经元（MN）的丧失。此外，在动物模型中，TDP-43的突变会导致家族性ALS和类似ALS的疾病。在这项研究中，我们研究了谷胱甘肽（GSH）在调节运动神经元NSC-34细胞TDP-43病理中的氧化应激反应中的作用。结果表明，GSH耗竭会产生与突变TDP-43类似的病理，包括胞质聚集体的发生，TDP-43磷酸化和内源性TDP-43的核清除。我们还证明了突变型TDP-43的引入^A315T内源性TDP-43的沉默和沉默，而不是野生型TDP-43的过度表达，导致类似的病理，包括细胞内GSH的耗竭，可能是由γ-谷氨酰半胱氨酸连接酶（GCLM）的调节亚基表达降低所致。 GSH合成中的限速酶。重要的是，用GSH单乙酯（GSHe）处理突变细胞可直接增加细胞内GSH，而无需合成GSH，可保护其免受TDP-43突变致病，包括减少聚集体形成，核清除率，活性氧（ROS）生产和细胞死亡。