Using mitochondrial Ca2+ uptake as a therapeutic target for ALS

使用线粒体 Ca2 摄取作为 ALS 的治疗靶点

• 批准号: 10659923
• 负责人: Lan Wei-LaPierre
• 金额: $ 48万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10659923
• 关键词: ALS pathology; ALS patients; Adult; Alleles; Amyotrophic Lateral Sclerosis; Animals; Attenuated; Axon; Bioenergetics; Brain; Buffers; C9ORF72; Cell Death; Cell Membrane Permeability; Cuprozinc Superoxide Dismutase; DNA-Binding Proteins; Defect; Degenerative Disorder; Disease; Disease Progression; Dissection; Distal; Dominant-Negative Mutation; Electron Microscopy; Event; Exercise; Generations; Genes; Genetic; Genetic Models; Homeostasis; Immunohistochemistry; In Situ; Longevity; Measurement; Membrane Potentials; Mitochondria; Molecular; Motor; Motor Neurons; Mus; Muscle; Muscle Contraction; Muscle Fibers; Muscle denervation procedure; Muscle function; Mutation; Neurodegenerative Disorders; Neuromuscular Junction; Onset of illness; Outcome; Paralysed; Pathogenesis; Pathologic; Pathology; Patients; Performance; Phenotype; Property; Reporting; Research; Respiratory Chain; Respiratory Failure; Role; Signal Transduction; Skeletal Muscle; Soleus Muscle; Spinal Cord; Structure; Symptoms; System; Testing; Therapeutic Effect; Tissues; Transgenic Mice; Treatment Efficacy; Western Blotting; amyotrophic lateral sclerosis therapy; attenuation; axonopathy; behavioral study; effective therapy; extensor digitorum; improved; in vivo; mitochondrial dysfunction; mitochondrial membrane; mouse model; muscular structure; neuron loss; neuronal survival; new therapeutic target; postsynaptic; pre-clinical; preservation; presynaptic; prevent; therapeutic target; uptake

Amyotrophic lateral sclerosis (ALS) is a fatal, adult-onset neurodegenerative disease characterized by progressive motor neuron (MN) loss, muscle denervation, and eventually, paralysis. Currently, no effective treatments are available to stop or reverse ALS disease progression and the precise molecular mechanisms underlie ALS pathogenesis remain elusive. Prior studies revealed decreased mitochondrial respiratory chain activity, altered mitochondrial ultrastructure, and mitochondrial dysfunction in both MN and skeletal muscle (SM) in ALS patients and mouse models. The first sign of ALS pathology occurs at the neuromuscular junction (NMJ), where presynaptic MN axons connect with postsynaptic SM end plates. To date, whether signals resulting in the initial NMJ damage are from MN or SM remain unclear. In this project, we aim to determine the tissue-specific causative role of mitochondrial Ca2+ uptake in SM and MN in disease onset and progression, and the therapeutic efficacy of reducing mitochondrial Ca2+ uptake on NMJ and SM function in ALS mice. We hypothesize that mitochondrial Ca2+ mishandling in both SM and MN actively contribute to ALS disease pathogenesis and that attenuating mitochondrial Ca2+ uptake mitigates mitochondrial damage and preserves NMJ/muscle function. To test this hypothesis, we will use transgenic mice with inducible, SM or MN-specific expression of a dominant negative form of the mitochondrial Ca2+ uniporter to specifically and selectively reduce mitochondrial Ca2+ uptake in SM and MN in hSOD1G93A mice and C9-500 (C9orf72) mice, two mouse models associated with the most prevalent genetic causes for ALS. The central hypothesis will be tested in two Specific Aims. Aim 1 will determine the role of mitochondrial Ca2+ uptake in SM or MN in survival, motor function, NMJ function and in vivo muscle performance in hSOD1G93A and C9-500 mice. Aim 2 will assess the impact of tissue-specific inhibition of mitochondrial Ca2+ uptake in SM or MN on NMJ and muscle structure, MN survival, muscle intrinsic contractile properties, mitochondrial structure and mitochondrial bioenergetics in SM of hSOD1G93A and C9-500 mice. This project will: 1) provide a systematic, longitudinal characterization of SM and NMJ function from a cellular level to whole animal level at different stages of disease progression in hSOD1G93A and C9-500 mice; 2) determine the degrees to which defects in mitochondrial Ca2+ uptake in SM or MN contribute to altered NMJ structure/function, disease onset and progression in hSOD1G93A and C9-500 mice; 3) provide the first detailed dissection on the relative role of mitochondrial Ca2+ uptake in SM and MN in ALS phenotype using the same genetic models and determine the origin of the signals that result in NMJ destruction (from SM or MN or both); 4) provide mechanistic evidence for whether mitochondrial Ca2+ mishandling is a trigger or a target for disease progression in ALS mice, regardless of the causing mutations (mitochondrial related or non-mitochondrial related); and most importantly, 5) test the validity of a potential new therapeutic target (mitochondrial Ca2+ uptake, or the mitochondrial Ca2+ uniporter) for the treatment of ALS.

肌萎缩侧索硬化症 (ALS) 是一种致命的、成人发病的神经退行性疾病，其特征是 进行性运动神经元 (MN) 丧失、肌肉失神经支配，最终导致瘫痪。目前，尚无有效 可用于阻止或逆转 ALS 疾病进展的治疗方法以及精确的分子机制 ALS 发病机制的背后仍然难以捉摸。先前的研究表明线粒体呼吸链减少 MN 和骨骼肌 (SM) 的活性、线粒体超微结构改变以及线粒体功能障碍 在 ALS 患者和小鼠模型中。 ALS 病理学的第一个迹象发生在神经肌肉接头 (NMJ)， 突触前 MN 轴突与突触后 SM 终板连接。迄今为止，信号是否导致 最初的 NMJ 损伤是来自 MN 还是 SM，目前尚不清楚。在这个项目中，我们的目标是确定组织特异性 SM 和 MN 中线粒体 Ca2+ 摄取在疾病发生和进展中的因果作用以及治疗 减少线粒体 Ca2+ 摄取对 ALS 小鼠 NMJ 和 SM 功能的功效。我们假设 SM 和 MN 中的线粒体 Ca2+ 处理不当积极促进 ALS 疾病的发病机制，并且 减弱线粒体 Ca2+ 摄取可减轻线粒体损伤并保留 NMJ/肌肉功能。到 检验这一假设，我们将使用具有可诱导的、SM 或 MN 特异性表达的显性基因的转基因小鼠。 线粒体 Ca2+ 单向转运蛋白的阴性形式可特异性、选择性地减少线粒体 Ca2+ 摄取 hSOD1G93A 小鼠和 C9-500 (C9orf72) 小鼠的 SM 和 MN 中，这两种小鼠模型与最 ALS 的常见遗传原因。中心假设将在两个具体目标中进行检验。目标1将决定 SM 或 MN 中线粒体 Ca2+ 摄取对生存、运动功能、NMJ 功能和体内肌肉的作用 hSOD1G93A 和 C9-500 小鼠中的表现。目标 2 将评估组织特异性抑制的影响 SM 或 MN 中线粒体 Ca2+ 摄取对 NMJ 和肌肉结构、MN 存活、肌肉内在收缩力的影响 hSOD1G93A 和 C9-500 小鼠 SM 的特性、线粒体结构和线粒体生物能学。这 项目将：1）从细胞水平提供 SM 和 NMJ 功能的系统、纵向表征 hSOD1G93A 和 C9-500 小鼠疾病进展不同阶段的整体动物水平； 2）确定 SM 或 MN 中线粒体 Ca2+ 摄取缺陷导致 NMJ 结构/功能改变的程度， hSOD1G93A 和 C9-500 小鼠的疾病发作和进展； 3）提供第一个详细的剖析 使用相同的遗传模型和 MN 中线粒体 Ca2+ 摄取在 ALS 表型中的相对作用 确定导致 NMJ 破坏的信号来源（来自 SM 或 MN 或两者）； 4）提供机制 线粒体 Ca2+ 处理不当是否是 ALS 小鼠疾病进展的触发因素或目标的证据， 无论引起突变（线粒体相关或非线粒体相关）；最重要的是， 5) 测试潜在新治疗靶点（线粒体 Ca2+ 摄取，或线粒体 Ca2+）的有效性 uniporter）用于治疗 ALS。