Modeling the progression of SOD1-linked motor neuron disease

模拟 SOD1 相关运动神经元疾病的进展

基本信息

批准号：
10375086
负责人：
DAVID R BORCHELT
金额：
$ 42.52万
依托单位：
UNIVERSITY OF FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-12-15 至 2026-11-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10375086
关键词：
ALS patients Age Amyotrophic Lateral Sclerosis Anatomy Animals Anti-Inflammatory Agents Astrocytes Attention Award Biological Biological Models Biology Characteristics Data Disease Disease Progression Exhibits Funding Goals Hand Human In Vitro Individual Inflammation Inflammatory Inherited Injections Laboratories Life Limb structure Link Literature Microglia Modeling Molecular Conformation Motor Neuron Disease Mus Mutation Nerve Degeneration Neuraxis Neurodegenerative Disorders Neurons Newborn Infant Paralysed Pathogenesis Pathologic Pathology Pathway interactions Patients Phenotype Population Preparation Prions Process Property Protein Conformation Proteins Reaction Recombinant adeno-associated virus (rAAV)Recombinants Research Resistance Role Route Seeds Signal Transduction Site Source Spinal Spinal Cord Tissues Transgenic Mice Variant Work cytokine design disease-causing mutation early onset encryption familial amyotrophic lateral sclerosis foot illness length improved insight misfolded protein mutant neuroinflammation novel prion-like sciatic nerve selective expression success superoxide dismutase 1 transmission process vector young adult

项目摘要

In the past 5 years, it has become clear that the protein pathology of many human neurodegenerative diseases exhibits characteristics of prions, including transmissibility, strain variation, and the ability to spread from a focal site of introduction. Amyotrophic lateral sclerosis (ALS) stands out as an example where the hallmarks of prion-like spreading is evident as weakness spreads along anatomically connected pathways. In familial ALS caused by mutations in superoxide dismutase 1 (SOD1-ALS), patients inheriting the A4V variant of SOD1 weakness spreads rapidly (average survival <1.5 years after the onset), whereas in patients inheriting the G37R variant weakness spreads slowly (average survival ~17 years). In the initial funding period of this award, our laboratory has uncovered evidence that this defining feature of SOD1-ALS may be explained by prion-like characteristics of mutant SOD1. Transgenic mice that express low levels of ALS mutant SOD1 develop disease late in life if at all. We have shown that paralysis can be accelerated in these mice by injecting spinal cord homogenates prepared from paralyzed mutant SOD1 transgenic mice or from human patients. We have also shown that we can inject these homogenates into the sciatic nerve of vulnerable mice to initiate a disease process that closely mimics the unilateral spread of weakness from one limb to another limb that is seen in humans. We have also successfully used purified recombinant SOD1 fibrilized in vitro to seed early onset paralysis in host mice, proving that SOD1 is capable of acting like a prion. Because we can propagate disease- causing conformations of SOD1 to naïve SOD1 “host proteins”, from hereafter we will refer to the misfolded conformation associated with disease-causing mutant SOD1 as an ALS prion. Building on the success of our initial work, we now propose four Specific Aims that are designed to improve our understanding of the biological role of prion-like spread in the pathogenesis of SOD1-ALS. In Aim 1, we seek to investigate whether the disease-causing mutations encrypt unique strain-like characteristics in misfolded SOD1 that influences the rate of prion-like spread in animal to animal transmission studies. In Aim 2, we seek to determine how the route of transmission and age of the host recipient mouse influence the propagation of SOD1-ALS prions. In our third and fourth Aims, using our novel model system we will turn our attention towards determining the extrinsic factors that determine how misfolded protein conformations may spread in the CNS and whether inflammatory signaling may influence such spread. In Aim 3, we will use newly generated loxp G85R-SOD1:YFP mice to determine the contribution of astrocytes in propagating SOD1-ALS prions. In Aim 4, we will use adeno- associated vectors to express pro- and anti-inflammatory cytokines as a means to assess the role of activated astrocytes and microglia in the propagation and spreading of SOD1-ALS prions. Our over-arching goal is to determine the contribution of intrinsic strain-like attributes in SOD1 and extrinsic non-cell autonomous processes to the prion-like propagation properties of disease-causing SOD1 conformations.

在过去的5年里，人们已经清楚许多人类神经退行性疾病的蛋白质病理学表现出朊病毒的特征，包括传播性、应变变异以及从病毒中传播的能力肌萎缩侧索硬化症 (ALS) 是一个突出的例子，其特征是在家族性肌萎缩侧索硬化症中，随着肌无力沿着解剖学上相连的通路传播，类似朊病毒的传播是显而易见的。由超氧化物歧化酶 1 (SOD1-ALS) 突变引起，遗传 SOD1 A4V 变体的患者无力迅速蔓延（发病后平均生存<1.5年），而在继承了无力的患者中 G37R 变体的弱点蔓延缓慢（平均生存期约为 17 年）。我们的实验室发现了证据表明 SOD1-ALS 的这一定义特征可以用朊病毒样来解释表达低水平 ALS 突变体 SOD1 的转基因小鼠的特征。如果有的话，我们已经证明，通过脊髓注射可以加速这些小鼠的瘫痪。我们有从瘫痪突变型 SOD1 转基因小鼠或人类患者制备的脐带匀浆。还表明我们可以将这些匀浆注射到脆弱小鼠的坐骨神经中以引发疾病过程密切模仿单侧无力从一个肢体传播到另一个肢体，如我们还成功地在体外使用纯化的重组 SOD1 纤维化来播种早期发病。使宿主小鼠瘫痪，证明 SOD1 能够像朊病毒一样发挥作用，因为我们可以传播疾病—— 导致 SOD1 构象为幼稚 SOD1“宿主蛋白”，从下文中我们将提到错误折叠与作为 ALS 朊病毒的致病突变体 SOD1 相关的构象以我们的成功为基础。在最初的工作中，我们现在提出了四个具体目标，旨在提高我们对朊病毒样传播在 SOD1-ALS 发病机制中的生物学作用在目标 1 中，我们试图研究是否存在这种作用。致病突变加密了错误折叠的 SOD1 中独特的菌株样特征，从而影响了在目标 2 中，我们试图确定动物间传播研究中的朊病毒样传播率。在我们的第三个研究中，宿主小鼠的传播和年龄影响 SOD1-ALS 朊病毒的传播。第四个目标，使用我们新颖的模型系统，我们将把注意力转向确定外在因素决定错误折叠的蛋白质构象如何在中枢神经系统中传播以及炎症是否存在的因素在目标 3 中，我们将使用新生成的 loxp G85R-SOD1:YFP 小鼠来进行信号传导。确定星形胶质细胞在传播 SOD1-ALS 朊病毒中的贡献在目标 4 中，我们将使用腺病毒。相关载体表达促炎和抗炎细胞因子，作为评估激活的作用的手段我们的首要目标是星形胶质细胞和小胶质细胞在 SOD1-ALS 朊病毒的繁殖和传播中的作用。确定 SOD1 和外在非细胞自主性中内在应变样属性的贡献过程导致致病 SOD1 构象的类朊病毒传播特性。