A Spontaneous Mouse Model of Motor Neuron Disease

描述（由申请方提供）：本提案的目的是表征运动神经元疾病的新自发小鼠模型并鉴定致病突变。运动神经元疾病，如肌萎缩侧索硬化症（ALS）和脊髓性肌萎缩症（SMA）在人群中很常见，其严重程度对患者来说是毁灭性的。这些疾病的分子基础仍然不清楚，尽管在动物模型和人类中鉴定了少数与遗传性运动神经元疾病相关的基因。这些疾病的动物模型对于致病机制的研究至关重要，可以为寻找治疗靶点提供信息，并为测试疗法提供临床前模型。许多现有的运动神经元疾病的动物模型被大量工程化以复制疾病表型，代价是精确地复制人类疾病的遗传学。例如，人类中SOD 1的显性突变导致家族性ALS，而小鼠模型需要人类蛋白质突变形式的过表达以产生类似的表型。相比之下，由于其相关表型而鉴定出的小鼠自发突变根本不是工程化的，因此避免了转基因模型的许多警告。例如，考克斯博士（该提案的主要研究者）鉴定的Ighmbp 2中的自发小鼠突变导致小鼠运动神经元疾病，这直接导致了人类脊髓性肌萎缩伴呼吸窘迫的遗传基础的鉴定。最近，我们在小鼠中发现了一种新的自发隐性突变，导致运动神经元疾病表型。该模型表现出明显的、进行性的神经肌肉功能丧失，特别是在后肢，到5-6月龄时几乎完全瘫痪。脊髓根和神经肌肉接头的检查表明运动神经元缺陷，脊髓腹角运动神经元细胞体的检查显示病理学包括富含曙红的、泛素阳性的包涵体。该模型的病理和表型与之前描述的小鼠模型不同，但准确地概括了人类运动神经元疾病的许多特征。因此，我们提出了两个目标来研究这种新的运动神经元疾病模型。在目标1中，我们将使用综合分析的粗大运动性能，电生理学特征，组织学，免疫细胞化学和电子显微镜检查的表型，以了解疾病的进展和病理运动神经元。这些信息将使我们能够准确地将小鼠表型与相关的人类疾病相关联。在目标2中，我们将使用定位克隆技术来鉴定致病突变。识别潜在的遗传缺陷对于理解运动神经元损失的基础至关重要。如果这种突变发生在已知的运动神经元疾病基因中，它很可能代表相关人类疾病的上级动物模型。如果突变发生在一个新的基因中，它将为导致运动神经元疾病的分子途径提供额外的机制见解，并为相关的人类疾病提供新的候选基因。 公共卫生相关性：肌萎缩侧索硬化症和脊髓性肌萎缩症等运动神经元疾病的严重程度是毁灭性的，但人们对潜在的病理机制知之甚少，而且目前的治疗方法效果有限。我们已经确定了一个新的，可遗传的运动神经元疾病的小鼠模型，并建议克隆基因负责。这项工作将增加我们对运动神经元疾病的理解，因此有助于寻找治疗靶点。此外，它要么将为现有的人类疾病提供上级动物模型，要么更有可能在患者中鉴定家族性运动神经元疾病的新候选基因。

DESCRIPTION (provided by applicant): The goal of this proposal is to characterize a new spontaneous mouse model of motor neuron disease and identify the causative mutation. Motor neuron diseases such as Amyotrophic Lateral Sclerosis (ALS) and Spinal Muscular Atrophies (SMA) are common in the population and devastating in their severity for the patient. The molecular basis for these diseases is still unclear despite the identification of a handful of genes associated with heritable motor neuron diseases in both animal models and humans. Animal models of these diseases are critical for research on the pathogenic mechanisms to inform the search for therapeutic targets and to provide preclinical models for testing therapies. Many existing animal models of motor neuron diseases are heavily engineered to reproduce the disease phenotype at the expense of accurately reproducing the genetics of the human disease. For example, dominant mutations in SOD1 in humans cause a familial form of ALS, whereas mouse models require overexpression of mutant forms of the human protein to develop a similar phenotype. In contrast, spontaneous mutations in mice identified because of their related phenotypes are not engineered at all and therefore avoid many caveats of transgenic models. For example, spontaneous mouse mutations in Ighmbp2 identified by Dr. Cox, a PI on this proposal, cause a motor neuron disease in mice that directly led to the identification of the genetic basis of Spinal Muscular Atrophy with Respiratory Distress in humans. Recently, we have identified a novel spontaneous recessive mutation in mice that causes a motor neuron disease phenotype. This model displays overt, progressive loss of neuromuscular function, particularly in the hind limbs, with almost complete paralysis by 5-6 months of age. Examination of spinal roots and neuromuscular junctions indicate a motor neuron defect, and examination of motor neuron cell bodies in the spinal cord ventral horn reveals pathology including eosin-rich, ubiquitin-positive inclusions. The pathology and phenotype of this model differ from previously described mouse models, but accurately recapitulates many of the hallmarks of human motor neuron diseases. We therefore propose two aims to study this new model of motor neuron disease. In Aim 1, we will examine the phenotype using an integrated analysis of gross motor performance, electrophysiological characterization, and histology, immunocytochemistry, and electron microscopy to understand the disease progression and pathology in the motor neurons. This information will allow us to accurately correlate the mouse phenotype with related human diseases. In Aim 2, we will use positional cloning techniques to identify the causative mutation. Identifying the underlying genetic defect is critical for understanding the basis of the motor neuron loss. If this mutation is in a known motor neuron disease gene, it is likely to represent a superior animal model of the associated human disease. If the mutation is in a novel gene, it will provide additional mechanistic insights into the molecular pathways that lead to motor neuron disease and also provide a new candidate gene for related human diseases. PUBLIC HEALTH RELEVANCE: Motor neuron diseases such as Amyotrophic Lateral Sclerosis and Spinal Muscular Atrophy are devastating in their severity, but the underlying pathological mechanisms are poorly understood and current therapies are of limited effectiveness. We have identified a new, heritable mouse model of motor neuron disease and propose to clone the gene responsible. This work will add to our understanding of motor neuron diseases, and therefore aid the search for therapeutic targets. In addition, it will either provide a superior animal model for an existing human disease, or more likely, identify a new candidate gene for familial motor neuron diseases in patients.