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Rescuing Niemann-Pick C Disease: Pathways of Liver and Brain Degeneration

拯救尼曼匹克 C 病：肝脏和大脑退化的途径

基本信息

批准号：
8525469
负责人：
Suzanne R Pfeffer
金额：
$ 30.89万
依托单位：
STANFORD UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-15 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8525469
关键词：
Address Affect Alzheimer&apos s Disease Animals Astrocytes Autophagocytosis Axon Binding Proteins Brain Brain Injuries Brain imaging Bypass Categories Cell Death Cell physiology Cells Cerebellum Cessation of life Childhood Cholesterol Cognitive Defect Degenerative Disorder Degradation Pathway Dichloromethylene Diphosphonate Disease Disease Pathway Disease Progression Dyes Employee Strikes Engineering Evaluation Failure Fibroblasts Foundations Genes Genetic Health Hepatic Hepatocyte Hepatomegaly Histocompatibility Testing ITGAM gene Immune Inflammation Inflammatory Intracellular Transport Label Lead Learning Life Liposomes Liver Location Locomotion Lysosomes Measures Microglia Microscope Monitor Movement Mus Mutation Nerve Degeneration Neurodegenerative Disorders Neuroglia Neurons Opitz trigonocephaly syndrome Organelles Pathology Pathway interactions Patients Process Production Property Proteins Quantum Dots Regulation Role Sirolimus Staging Sterols Structure of thyroid parafollicular cell Supraoptic Vertical Ophthalmoplegia System Technology Testing Tetanus Helper Peptide Therapeutic Therapeutic Intervention Time Tissues Toxin Transgenes Transmembrane Domain Virus Virus Diseases Work cell injury cell type effective therapy experience falls functional restoration gene therapy human TLR7 protein improved in vivo insight liver function liver injury macrophage mouse model mutant neuron component novel strategies prevent relating to nervous system research study trafficking two-photon

项目摘要

DESCRIPTION (provided by applicant): Niemann Pick type C (NPC) disease is a fatal pediatric disorder. The disease is due to mutations in either of two genes, NPC1, which encodes a 13 transmembrane domain sterol-binding protein, and NPC2, which encodes a soluble sterol-binding protein. Loss of either gene causes aberrant organelle trafficking and accumulation of free cholesterol within lysosomes. NPC patients suffer from hepatomegaly and progressive cognitive and locomotion losses, with massive Purkinje neuron (PN) death in the cerebellum. There is no effective treatment for NPC. Specific Aim 1: Determine why Purkinje neurons die in NPC disease and at what stages the disease can be arrested or reversed. We will provide a functional tagged- Npc1 protein to specific classes of cells, eg neurons, astrocyte, or liver cells, in the Npc1-/- background, to define how each cell type contributes to disease. Our transgenes are engineered with Tet-technology to allow cell-specific and temporal regulation of tagged Npc1 production. Specific Aim 2: Learn how NPC disease affects intracellular trafficking in Purkinje neurons. Loss of Npc1 causes striking intracellular trafficking defects in fibroblasts. To learn how PNs are affected, we will culture them and track the trafficking of NGF and other molecules labeled with quantum dots. We will use a portable two-photon microscope in combination with organelle dyes to characterize vesicular movements in living astrocytes and PNs of wild-type and Npc1-/- mice. In living brains of our newly engineered mice we will analyze movements of Npc1-positive organelles in PNs, other neurons, and astrocytes to determine what changes may cause cell death. Specific Aim 3: Determine whether inflammation protects from, or causes, NPC cell death. We will produce functional Npc1 in neurons, hepatocytes, and inflammatory cells, in otherwise Npc-/- mice, and see which of these is most effective in preventing inflammation. We will use mouse mutations that reduce inflammation in combination with Npc1-/-, and see whether PN survival and liver pathology are improved or worsened. We will ablate hepatic macrophages with chlodronate-filled liposomes and assess the role of macrophages in NPC liver damage. Specific Aim 4: Discover whether autophagy protects from, or causes, NPC cell death. We will cross NPC mice with mutants that cannot trigger autophagy: Toll like receptor-7 and beclin-1 deficient mice. Conversely we will test mice that have over-expressing Beclin1 in neurons and consequent have heightened autophagy, or enhance autophagy with Beclin1 virus infections or rapamycin treatments. Lysosome storage disorders like NPC encompass nearly 60 different conditions, most of which damage liver and/or brain function. Some, including NPC, have similarities to Alzheimer disease. We have used a novel approach to engineer mice that will allow us to learn how different cell types and processes contribute to disease. Learning the roles of inflammation and autophagy in NPC neurodegeneration has direct implications for therapeutic interventions that will arrest or reverse disease progression.

描述（由申请人提供）：C 型尼曼皮克病 (NPC) 是一种致命的儿科疾病。该疾病是由于两个基因中的一个突变所致：NPC1（编码 13 个跨膜结构域的甾醇结合蛋白）和 NPC2（编码可溶性甾醇结合蛋白）。任一基因的丢失都会导致细胞器运输异常和溶酶体中游离胆固醇的积累。鼻咽癌患者患有肝肿大、进行性认知和运动丧失，以及小脑中大量浦肯野神经元 (PN) 死亡。鼻咽癌没有有效的治疗方法。具体目标 1：确定浦肯野神经元在鼻咽癌疾病中死亡的原因以及疾病在什么阶段可以被阻止或逆转。我们将在 Npc1-/- 背景下向特定类别的细胞（例如神经元、星形胶质细胞或肝细胞）提供功能性标记 Npc1 蛋白，以定义每种细胞类型如何导致疾病。我们的转基因采用 Tet 技术进行设计，可对标记的 Npc1 生产进行细胞特异性和时间调控。具体目标 2：了解鼻咽癌疾病如何影响浦肯野神经元的细胞内运输。 Npc1 的缺失会导致成纤维细胞中显着的细胞内运输缺陷。为了了解 PN 如何受到影响，我们将培养它们并追踪 NGF 和其他用量子点标记的分子的运输。我们将使用便携式双光子显微镜与细胞器染料相结合来表征活星形胶质细胞中的囊泡运动以及野生型和 Npc1-/- 小鼠的 PN。在我们新改造的小鼠的活体大脑中，我们将分析 PN、其他神经元和星形胶质细胞中 Npc1 阳性细胞器的运动，以确定哪些变化可能导致细胞死亡。具体目标 3：确定炎症是否可以防止或导致 NPC 细胞死亡。我们将在 Npc-/- 小鼠的神经元、肝细胞和炎症细胞中产生功能性 Npc1，并看看其中哪一种在预防炎症方面最有效。我们将使用减少炎症的小鼠突变与 Npc1-/- 相结合，看看 PN 存活率和肝脏病理学是否得到改善或恶化。我们将用充满氯膦酸盐的脂质体消融肝巨噬细胞，并评估巨噬细胞在 NPC 肝损伤中的作用。具体目标 4：发现自噬是否可以防止或导致 NPC 细胞死亡。我们将 NPC 小鼠与不能触发自噬的突变体杂交：Toll 样受体 7 和 beclin-1 缺陷小鼠。相反，我们将测试神经元中过度表达 Beclin1 并因此增强自噬的小鼠，或者通过 Beclin1 病毒感染或雷帕霉素治疗增强自噬。鼻咽癌等溶酶体贮积症涵盖近 60 种不同的病症，其中大多数会损害肝脏和/或大脑功能。包括鼻咽癌在内的一些疾病与阿尔茨海默病有相似之处。我们使用了一种新的方法来改造小鼠，这将使我们能够了解不同的细胞类型和过程如何导致疾病。了解炎症和自噬在鼻咽癌神经变性中的作用对于阻止或逆转疾病进展的治疗干预具有直接影响。