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）：Niemann Pick C型（NPC）疾病是一种致命的小儿疾病。该疾病是由于两个基因NPC1中的任何一个突变引起的，该基因编码13个跨膜结构域固醇结合蛋白和NPC2，该突变编码了编码可溶性固醇结合蛋白的NPC2。丧失基因会导致异常细胞器的运输和溶酶体内游离胆固醇的积累。 NPC患者患有肝肿大，进行性认知和运动损失，小脑中大规模的Purkinje神经元（PN）死亡。 NPC没有有效的治疗方法。具体目标1：确定为什么Purkinje神经元在NPC疾病中死亡，以及在哪些阶段可以被捕或逆转。我们将向NPC1 - / - 背景中的特定细胞（例如神经元，星形胶质细胞或肝细胞）提供功能性标记的NPC1蛋白，以定义每种细胞类型如何对疾病贡献。我们的转基因采用TET技术设计，以允许对标记的NPC1产生的细胞特异性和时间调节。具体目的2：了解NPC疾病如何影响浦肯野神经元的细胞内贩运。 NPC1的丧失会导致成纤维细胞中的细胞内运输缺陷。为了了解PN的影响，我们将培养它们，并跟踪NGF和其他用量子点标记的分子的运输。我们将使用便携式的两光子显微镜与细胞器染料结合使用，以表征野生型星形胶质细胞和野生型和NPC1 - / - 小鼠的囊泡运动。在新工程小鼠的活体大脑中，我们将分析PNS，其他神经元和星形胶质细胞中NPC1阳性细胞器的运动，以确定什么变化可能导致细胞死亡。特定目标3：确定炎症是免受NPC细胞死亡的影响还是原因。我们将在神经元，肝细胞和炎症细胞中产生功能性NPC1，否则将在NPC - / - 小鼠中产生功能性NPC1，并查看其中哪种最有效地预防炎症。我们将使用与NPC1 - / - 结合使用的小鼠突变，并查看PN存活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。特定目的4：发现自噬是保护或原因是NPC细胞死亡。我们将用无法触发自噬的突变体穿越NPC小鼠：像受体7和Beclin-1缺陷小鼠一样。相反，我们将测试在神经元中过度表达beclin1并因此增强自噬或通过Beclin1病毒感染或雷帕霉素治疗增强自噬的小鼠。 NPC等溶酶体储存障碍包括近60种不同的疾病，其中大多数会损害肝脏和/或脑功能。有些（包括NPC）与阿尔茨海默氏病具有相似之处。我们已经使用了一种新颖的方法来设计小鼠，这将使我们能够学习不同的细胞类型和过程如何促进疾病。学习炎症和自噬在NPC神经变性中的作用直接影响治疗干预措施，这些干预措施将阻止或逆转疾病的进展。