NEURODEGENERATION AND NEUROPROTECTION IN MND

该提案使用体外和体内方法来探索 神经退行性和神经保护机制是积极的运动 神经元疾病（MND）。有两种主要的疾病， 靶向：家族性肌萎缩侧索硬化症（FALS）相关 Cu-Zn超氧化物歧化酶1型（SOD 1）（FALS）（1-3）突变和 浪费的（wst/wst）小鼠，其与调节基因中的缺失相关， PCNA（增殖细胞核抗原）基因区域（4）。之一 这个项目的假设是，有一个类似的途径， 这两种疾病过程中的神经元死亡。 该项目的一个主要目标是阐明细胞死亡的机制， 并确定药物和治疗方法， 关于MND特异性SOD 1相关的FALS和浪费的 老鼠.在第一个具体目标中，我们将研究 突变的SOD 1和鉴定从突变的SOD 1- 诱导细胞死亡。我们将在大肠杆菌中表达野生型和突变型SOD 1基因。 在培养物和中枢神经系统（CNS）中的原代神经元， 重组复制缺陷型腺病毒（AdV）作为载体，和 开始确定这种表达对自由基的影响， 积累、神经元功能和活力以及SOD 1的聚集。我们 还将在酵母中表达野生型和突变型SOD 1基因，以阐明 酶的突变或变化是否导致聚集， 不溶性我们将测试各种代理人的能力，以拯救在 体外培养细胞，由突变体SOD 1诱导凋亡，然后测试 这些药剂是否会影响FALS转基因小鼠的疾病（在 第三个具体目标）。在第二个具体目标中，我们将研究 在被浪费的老鼠中神经元死亡的原因。我们将研究神经元 来测试这些细胞是否有降低的水平， 发生自发性细胞凋亡（体外和体内），或 更容易受到应激条件如氧化应激的影响。一 更好地了解ALS中MN死亡的发病机制， 废鼠可能阐明散发性ALS中细胞死亡的机制 和其他神经退行性过程，也可能导致 确定MND的有效治疗方法。

This proposal uses in vitro and in vivo approaches to explore neurodegenerative and neuroprotective mechanisms that are active in motor neuron diseases (MNDs). There are two main disease conditions that are targeted: familial amyotrophic lateral sclerosis (FALS) associated with mutations in CU-Zn superoxide dismutase type 1 (SOD1)(FALS) (1-3) and the wasted (wst/wst) mouse which is associated with a deletion in a regulatory region of the PCNA (proliferating cell nuclear antigen) gene (4). One of the hypothesis in this project is the there is a similar pathway with respect to neuronal death in both of these disease processes. A major goal of this project is to clarify the mechanisms of cell death, and to identify agents and therapeutic approaches that are important in rescue with respect to MND-specifically SOD1 linked FALS and the wasted mouse. In the first specific aim, we will investigate the "toxicity" of mutant SOD1s and identify agents that rescue neurons from mutant SOD1- induced cell death. We will express wild type and mutant SOD1 genes in primary neurons in culture and in the central nervous system (CNS) using recombinant replication-defective adenoviruses (AdVs) as vectors, and begun to determine the effect of this expression on free radical accumulation, neuronal function and viability, and aggregation of SOD1. We will also express wild type and mutant SOD1 genes in yeast to clarify whether a mutation of change in the enzyme leads to aggregation and insolubility. We will test varied agents for their ability to rescue in vitro cultured cells from mutant SOD1-induced apoptosis, and then test whether these agents will affect disease in the FALS transgenic mouse (in the third specific aim). In the second specific aim we will investigate the reasons for neuronal death in the wasted mice. We will study neurons from the wasted mouse to test whether these cells have a decreased level of PCNA message, undergo spontaneous apoptosis (in vitro and in vivo), or are more susceptible to stress conditions such as oxidative stress. A better understanding of the pathogenesis of MN death in FALS and the wasted mouse may clarify the mechanism(s) of cell death in sporadic ALS and other neurodegenerative processes and also potentially lead to the identification of effective treatments for MNDs.