Identifying quantitative trait loci that regulate enterovirus D68 pathogenesis using the Collaborative Cross

使用协作交叉识别调节肠道病毒 D68 发病机制的数量性状位点

基本信息

批准号：
10113539
负责人：
VINCENT R RACANIELLO
金额：
$ 20.25万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-24 至 2022-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10113539
关键词：
6 year old Admixture Adult Aerosols Air Animal Model Animals Antibodies Antiviral Therapy Apoptosis Asia Asthma Australia Biology Body Weight decreased Brain Bronchiolitis CASP3 gene California Candidate Disease Gene Cells Cerebrospinal Fluid Child Chronic Obstructive Airway Disease Complex Cystic Fibrosis Cytoplasm DNA Fragmentation Data Development Diagnosis Disease Disease Outbreaks Enterovirus Enterovirus 68 Enterovirus Infections Epidemiology Epithelial Cells Etiology Europe Family Gastrointestinal tract structure Generations Genes Genetic Genetic Enhancement Genetic Polymorphism Genetic Predisposition to Disease Genetic Variation Genotype Goals Human Immunofluorescence Immunologic In Situ Nick-End Labeling Inbred Mouse Indirect Immunofluorescence Individual Infection Infection prevention Inherited Knockout Mice Knowledge Lead Liquid substance Lower respiratory tract structure Lung Lung diseases Maps Measures Medical Modeling Monitor Morbidity - disease rate Mus Neuraxis Neurologic Neurologic Dysfunctions Neurons Oral Outcome Paralysed Pathogenesis Pathology Patients Phenotype Plaque Assay Pneumonia Population Predisposition Production Quantitative Trait Loci Randomized Refractory Research Respiration Disorders Respiratory System Rhinovirus Severity of illness Site Slice Spinal Cord Stains Susceptibility Gene System Time Tissues Tropism United States Vaccine Therapy Variant Viral Pathogenesis Viral Proteins Virus Virus Activation Virus Diseases Virus Replication Viviparous-1 protein Work acute flaccid myelitis animal model development bronchial epithelium cell type chromosomal location cytochrome c design disease phenotype emerging pathogen human disease human pathogen in vivo Model infectious disease model insight member mortality mouse model nervous system disorder neurological pathology novel therapeutics pandemic disease respiratory respiratory pathogen seropositive socioeconomics transmission process vaccine evaluation virology

项目摘要

Project Summary Enteroviruses are among the most prevalent human pathogens. Traditionally, these viruses were associated with infection of cells within the alimentary tract, oral-fecal transmission between hosts and the development of rare neurological complications including acute flaccid myelitis (AFM). Yet, numerous members of this virus family including human rhinoviruses and enterovirus D-68 (EV-D68) are respiratory pathogens, replicating within the epithelial cells of the upper and lower respiratory tracts, and transmitted between hosts by aerosols In 2014 it was recognized that EV-D68 infection can also result in AFM. With each subsequent biennial outbreak of EV-D68 infection, the number of children infected with EV-D68 has been rising as well as the number of confirmed cases of AFM. Despite strong epidemiologically evidence that the virus is an etiologic agent of AFM, this association has been questioned due the inability to isolate infectious virus from the cerebrospinal fluid of children with EV-D68 induced paralysis. Together, these observations attest to the socioeconomic impact of EV-D68 infection as well as the unmet medical need for better an understanding of EV-D68 biology and the development of novel therapeutics. Notwithstanding the development of multiple animal models of EV-D68 infection, no system replicates the spectrum of pathologies associated with EV-D68; consequently, these models are insufficient for the development and testing of vaccines and antiviral therapies to treat or prevent infection or the development of AFM. We hypothesize that the limited genetic diversity of inbred mice used in many in vivo models of infectious disease limits the establishment of an animal model of EV-D68 biology. The enhanced genetic diversity of mice of the collaborative cross (CC) emulates that of the human population as well as the subtleties in associated phenotypes that are missing in null mice. The goal of the proposed research is to establish a mouse model of EV-D68 pathology and identify naturally-occurring genetic polymorphisms in loci whose products quantitatively regulate susceptibility to viral disease. To identify CC mice susceptible to EV-D68 infection, air-liquid interface cultures of the bronchial epithelium and organotypic brain slice cultures will be generated from 8- and 28-day old mice of randomized genetic backgrounds and infected with three isolates of EV-D68, and the production of infectious virus will be assessed by plaque assay. We will also identify the cell types that are infected by, and visualize tissue damage resulting from, virus infection by H&E staining and indirect immunofluorescence using antibodies generated against the viral protein VP1, and proteins specific to respiratory and neuronal cell types. The mice identified in Aim 1 in which EV-D68 replicated with the highest and lowest efficiency will be crossed in Aim 2. Analysis of the F1 generation will determine the mode by which EV-D68 susceptibility is inherited, while that of the F2 generation will describe the quantitative trait loci (QTL) that modulate EV-D68 infection. Use of CC mice will promote the development of an animal model that reflects the spectrum of pathologies associated with EV-D68 infection.

项目概要肠道病毒是最流行的人类病原体之一。传统上，这些病毒与随着消化道内细胞的感染、宿主之间的口腔-粪便传播以及感染的发展罕见的神经系统并发症，包括急性弛缓性脊髓炎（AFM）。然而，这种病毒的许多成员包括人鼻病毒和肠道病毒 D-68 (EV-D68) 在内的家族是呼吸道病原体，可复制存在于上呼吸道和下呼吸道的上皮细胞内，并通过气溶胶在宿主之间传播 2014 年，人们认识到 EV-D68 感染也可导致 AFM。随后的每一个双年展随着EV-D68感染的爆发，感染EV-D68的儿童人数不断上升， AFM 确诊病例数。尽管有强有力的流行病学证据表明该病毒是一种病原体 AFM 代理人，该协会因无法从 AFM 中分离出传染性病毒而受到质疑 EV-D68引起的瘫痪儿童的脑脊液。这些观察结果共同证明了 EV-D68 感染的社会经济影响以及未满足的医疗需求，以更好地了解 EV-D68 生物学和新型疗法的开发。尽管发展了多种 EV-D68 感染的动物模型，没有系统能够复制与 EV-D68 相关的病理谱系；因此，这些模型不足以开发和测试疫苗和抗病毒疗法治疗或预防感染或 AFM 的发展。我们假设有限的遗传多样性用于许多传染病体内模型的近交小鼠限制了传染病动物模型的建立 EV-D68 生物学。协作杂交（CC）小鼠遗传多样性的增强与普通小鼠的遗传多样性相似。人类群体以及无效小鼠中缺失的相关表型的微妙之处。目标是拟议的研究是建立 EV-D68 病理学小鼠模型并识别自然发生的位点的遗传多态性，其产物定量调节对病毒性疾病的易感性。识别 CC小鼠易受EV-D68感染，支气管上皮气液界面培养物和器官型脑切片培养物将从随机遗传的 8 日龄和 28 日龄小鼠中产生背景并感染了三种 EV-D68 分离株，并将评估传染性病毒的产生通过噬菌斑测定。我们还将识别被感染的细胞类型，并可视化由此产生的组织损伤通过 H&E 染色和使用针对病毒产生的抗体进行间接免疫荧光来检测病毒感染病毒蛋白 VP1 以及呼吸细胞和神经细胞类型特有的蛋白质。目标 1 中鉴定的小鼠其中 EV-D68 复制的效率最高和最低的将在目标 2 中交叉。 F1 分析一代将决定EV-D68易感性的遗传方式，而F2代将决定EV-D68易感性的遗传方式将描述调节 EV-D68 感染的数量性状基因座 (QTL)。 CC小鼠的使用将促进开发反映与 EV-D68 感染相关的病理谱系的动物模型。