Pathogenesis and motor neuron degeneration of a novel disease associated with a P158A mutation in NAMPT

与 NAMPT P158A 突变相关的新型疾病的发病机制和运动神经元变性

基本信息

批准号：
10444087
负责人：
Shinghua Ding
金额：
$ 49.92万
依托单位：
UNIVERSITY OF MISSOURI-COLUMBIA
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-15 至 2027-01-01
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10444087
关键词：
13 year old Aging Anabolism Animals Atrophic Axon Biochemical Reaction Bioenergetics Biological Assay Biological Process Brain Cell Death Cell division Cells Cessation of life Citric Acid Cycle Clinical Code DNA Repair Data Dendritic Spines Development Diagnosis Disease Disease Progression Electron Transport Electrophysiology (science)Energy Metabolism Enzymes Essential Genes Fibroblasts Foot Deformities Functional disorder Gene Expression Profiling Genes Genetic Diseases Glycolysis Goals Hand deformities Hereditary Motor and Sensory Neuropathies Human Image In Vitro Inborn Errors of Metabolism Inherited Ischemic Stroke Lead Link Lower Extremity Mammalian Cell Mammals Metabolic Metabolic Pathway Metabolic dysfunction Metabolism Mitochondria Molecular Motor Motor Cortex Motor Neurons Mus Muscle Muscle Fibers Muscle Weakness Mutant Strains Mice Mutation Nerve Degeneration Neuroglia Neuromuscular Junction Neurons Neuropathy Niacinamide Nicotinamide Mononucleotide Nicotinamide adenine dinucleotide Oxidative Phosphorylation Oxidative Stress Paralysed Pathogenesis Pathologic Pathway interactions Patients Pentosephosphate Pathway Phenotype Play Preparation Production Proteins Recombinants Reporting Respiration Role Sensory Siblings Signal Transduction Single Nucleotide Polymorphism Skin Stable Isotope Labeling Stress Structure Symptoms Synapses Technology Testing Time Tissues Transferase Variant Western Blotting Wheelchairs Wild Type Mouse axonal degeneration base cofactor fluorescence imaging gene function human disease in vivo induced pluripotent stem cell metabolomics mitochondrial dysfunction motor impairment motor neuron degeneration mouse model muscle degeneration mutant mutant mouse model nervous system disorder neuromuscular function neuron loss nicotinamide phosphoribosyltransferase novel patch clamp protective effect response synaptic function tool two-photon

项目摘要

Project Summary Nicotinamide adenine dinucleotide (NAD+) is a cofactor required for glycolysis, the tricarboxylic acid cycle (TCA) and enzymatic reaction in electron transport chain (ETC). In mammalian cells, NAD+ salvage pathway, where nicotinamide phosphoribosyltransferase (NAMPT) is the rate-limiting enzyme, is the predominant pathway for NAD+ biosynthesis. Although the dysregulation of NAD+ in aging and neurodegerative diseases has been reported, genetic diseases caused by NAMPT variants have not been clinically recognized and understood. Here we identified the first case of an inherited neurological disease caused by a homozygous single nucleotide polymorphism (SNP), i.e., a P158A mutation in the coding region of NAMPT gene. The major clinical features of patients include impaired motor coordination, muscle weakness, atrophy of lower extremities, positive Babiński sign. The patients were diagnosed as hereditary motor and sensory neuropathy involving axonal degeneration and neuromuscular junction (NMJ) dysfunction. Using skin-derived patient fibroblasts (p-FBs), our preliminary studies found that P158A mutation causes reduced bioenergetics, mitochondrial dysfunction, and decreased enzymatic activity of NAMPT for NAD+ biosynthesis compared with healthy control fibroblasts (c-FBs). The results indicate the pathological conditions related to the patients is initially resulted from bioenergetic stress and ultimately from neuronal and muscular degeneration. Thus, our project goal is to understand the pathogenesis and the mechanism of neuronal and muscular degeneration of this new disease. To achieve our goal, we generated many molecular tools including P158A-NAMPT mutant mice, c- & p-FBs-derived induced pluripotent stem cells (c- & p-iPSCs including isogenic and patient like p-iPSCs), and iPSC-induced motor neurons (c- & p- iMNs). We propose three Specific Aims. Aim 1 will test the hypothesis P158A mutation in NAMPT causes mitochondrial and synaptic dysfunction of p-iMNs. Using iMNs, we will study the effect of P158A mutation on cellular bioenergetics, glycolytic metabolism and mitochondrial respiration. We will also conduct combined metabolomic and transcriptional profiling to determine the molecular base of metabolic changes caused by P158A mutation. Aim 2 will test the hypothesis that P158A mutation in NAMPT causes MN degeneration. Using the mutant mice, we will study disease progression, upper and lower MN degeneration. We will use electrophysiological and two-photon (2-P) imaging to study the effect of P158A mutation on sensory response and cytosolic and mitochondrial Ca2+ signaling. Aim 3 will test the hypothesis that P158A mutation in NAMPT causes NMJ abnormalities and muscle degeneration. We will assess structural and functional abnormalities of NMJs and muscle contractile response of semitendinosus muscles isolated from the symptomatic mutant mice. A human disease caused by NAMPT mutation has not been reported so far. Our application represents a first in-depth study on the pathogenesis and mechanism of motor neuron and muscle degeneration of a new neurological disease caused by a mutation in NAMPT gene.

项目概要烟酰胺腺嘌呤二核苷酸 (NAD+) 是糖酵解、三羧酸循环 (TCA) 所需的辅助因子和电子传递链 (ETC) 中的酶反应，在哺乳动物细胞中，NAD+ 补救途径，其中烟酰胺磷酸核糖转移酶 (NAMPT) 是限速酶，是 NAD+ 生物合成虽然在衰老和神经退行性疾病中 NAD+ 失调已被证实。据报道，NAMPT 变异引起的遗传性疾病尚未得到临床认识和了解。我们发现了第一例由纯合单核苷酸引起的遗传性神经系统疾病多态性（SNP），即NAMPT基因编码区的P158A突变是该病的主要临床特征。患者包括运动协调受损、肌肉无力、下肢萎缩、巴宾斯基阳性患者被诊断为遗传性运动和感觉神经病，涉及轴突变性。和神经肌肉接头（NMJ）功能障碍，我们的初步研究是使用皮肤来源的患者成纤维细胞（p-FB）。研究发现 P158A 突变会导致生物能降低、线粒体功能障碍和与健康对照成纤维细胞 (c-FB) 相比，NAMPT 的 NAD+ 生物合成酶活性。结果表明，与患者相关的病理状况最初是由生物能量应激和最终来自神经和肌肉退化。因此，我们的项目目标是了解发病机制。以及这种新疾病的神经和肌肉变性的机制为了实现我们的目标，我们。许多生成的分子工具，包括 P158A-NAMPT 突变小鼠、c-和 p-FBs 衍生的诱导多能性干细胞（c- 和 p-iPSC，包括同基因和患者样 p-iPSC），以及 iPSC 诱导的运动神经元（c- 和 p- iMNs）。我们提出三个具体目标 1 将检验 P158A 突变导致 NAMPT 的假设。 p-iMN 的线粒体和突触功能障碍，我们将研究 P158A 突变的影响。我们还将对细胞生物能量学、糖酵解代谢和线粒体呼吸进行联合研究。代谢组学和转录分析以确定由以下原因引起的代谢变化的分子基础 P158A 突变。目标 2 将检验 NAMPT 中 P158A 突变导致 MN 变性的假设。我们将使用突变小鼠来研究疾病进展、上、下 MN 变性。电生理和双光子 (2-P) 成像研究 P158A 突变对感觉反应的影响目标 3 将检验 NAMPT 中 P158A 突变的假设。导致 NMJ 异常和肌肉退化。我们将评估结构和功能异常。从症状突变体中分离出的 NMJ 和半腱肌的肌肉收缩反应迄今为止，我们的应用还没有报道由 NAMPT 突变引起的人类疾病。首次深入研究运动神经元和肌肉变性的发病机制和机制 NAMPT基因突变引起的神经系统疾病。