Francisella - ATII interactions in respiratory tularemia

弗朗西斯菌 - ATII 在呼吸道兔热病中的相互作用

• 批准号: 8499239
• 负责人: THOMAS C. ZAHRT
• 金额: $ 21.57万
• 依托单位: MEDICAL COLLEGE OF WISCONSIN
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8499239
• 关键词: A549; Acute; Address; Alveolar; Alveolar Cell; Alveolar Macrophages; Attenuated Live Virus Vaccine; Bacteria; Biological Models; Breathing; Categories; Cell Line; Cells; Complement; Defect; Disease; Distant; Epithelial Cells; Epithelium; Francisella; Francisella tularensis; Generations; Genes; Hematogenous Spread; Human; Immune response; In Vitro; Infection; Inflammatory; Knowledge; Lead; Libraries; Lung; Lung Inflammation; Mediating; Modeling; Mononuclear; Morbidity - disease rate; Mus; Pathogenesis; Phenotype; Pneumonia; Proteins; Pulmonary alveolar structure; Reporting; Respiratory System; Respiratory tract structure; Site; Staging; Surface; Technology; Testing; Tularemia; Virulent; aerosolized; alveolar epithelium; cell type; combat; comparative; cytotoxicity; in vivo Model; insight; interstitial; macrophage; member; mortality; mutant; new therapeutic target; novel; novel therapeutics; pathogen; pneumocyte; prevent; programs; research study; respiratory; response; screening; therapeutic vaccine; trafficking; transcriptome sequencing; transmission process; vaccine candidate

DESCRIPTION (provided by applicant : ABS Francisella tularensis (Ft) is a Gram-negative facultative intracellular pathogen and the causative agent of tularemia, a disease associated with high morbidity and mortality in humans. Pneumonic tularemia, caused by inhalation of aerosolized Ft or hematogenous spread of Ft from distant infectious foci to the lung, is the most lethal form of the disease, resulting in up to 50% mortality without treatment. Characterized by profound disruption of the alveolar epithelium and profuse alveolar and interstitial mononuclear infiltrates, Ft pneumonia is an acute inflammatory infection often resulting in lung damage. Alveolar macrophages are the primary cell type targeted by Ft following infection, serving as both a site of bacterial replication and vehicle for subsequent dissemination. However, Ft encounters other cell types following inhalation infection, including lung epithelial cells. Recently, Ft interaction with alveolar type II and type I (ATII and ATI) pneumocytes was reported during early stages of murine pneumonic tularemia. The significance of these interactions in the lifecycle of Ft remains unclear, and no studies to date have addressed the importance of Ft interactions with human alveolar pneumocytes. Preliminary studies described in this proposal indicate that Ft infects, replicates, and persists in the human ATII-like cell lin A549 without inducing host cell cytopathic effects even at high multiplicities of infection, a phenotype that contrasts that seen following infection of Ft in other cell types including macrophages where host cytopathic effects are observed within 24 hrs. This proposal seeks to fill our current knowledge gap by investigating aspects of Ft interactions with primary human ATII and ATI pneumocytes. We hypothesize that ATII and/or ATI pneumocytes may serve as an intracellular reservoir for virulent Ft within the human lung, allowing a subset of bacteria to be maintained within a protected niche where they are able to modulate the subsequent host response. To address this hypothesis, three specific aims have been proposed. First, a primary human ATII/ATI infection model for Ft will be established and used to characterize internalization, replication, and trafficking of the bacterium. Second, bacterial determinants expressed by Ft within lung pneumocytes will be identified using RNA-seq technology and by screening a >20,000-member Schu S4::himar1 transposon mutant library in A549 cells for defects in cell cytotoxicity suppression. Finally, a subset of pneumocyte-expressed genes identified above will be targeted for disruption and characterization using various in vitro and in vivo model systems of infection. Collectively, these studies are expected to provide novel insights into the importance of ATII/ATI pneumocytes in Ft pathogenesis. This information may lead to identification of novel therapeutic targets or vaccine candidates to combat Ft infection.

描述(由申请人提供：ABS Francisella tularsis(Ft)是一种革兰氏阴性兼性细胞内病原体，是图拉热症的病原体，图拉热症是一种与人类高发病率和死亡率相关的疾病。肺炎性图拉热症是一种最致命的疾病，由吸入雾化的Ft或Ft通过血液从远处的感染灶扩散到肺部而引起，在不治疗的情况下可导致高达50%的死亡率。Ft肺炎是一种急性炎症性感染，以肺泡上皮细胞的严重破坏和大量的肺泡和间质单核细胞浸润为特征，常导致肺损伤。肺泡巨噬细胞是Ft感染后的主要靶细胞类型，既是细菌复制的场所，也是随后传播的载体。然而，吸入性感染后，FT会遇到其他类型的细胞，包括肺上皮细胞。最近，在小鼠肺炎性图拉热症的早期阶段，报道了Ft与肺泡II型和I型(ATII和ATI)肺泡细胞的相互作用。这些相互作用在Ft生命周期中的意义尚不清楚，到目前为止还没有研究表明Ft与人肺泡肺泡细胞相互作用的重要性。这项建议中描述的初步研究表明，Ft感染、复制并持续存在于人ATII样细胞Lin A549中，即使在高复数感染时也不会诱导宿主细胞病变效应，这一表型与Ft在其他类型的细胞中感染后的情况形成对比，在巨噬细胞中，在24小时内观察到宿主细胞病变效应。这项建议试图通过研究Ft与原代人类ATII和ATI肺泡细胞的相互作用来填补我们目前的知识空白。我们假设ATII和/或ATI肺泡细胞可能作为人肺内毒力Ft的细胞内储存库，允许细菌的子集保持在受保护的生态位内，在那里它们能够调节随后的宿主反应。为了解决这一假设，提出了三个具体目标。首先，将建立Ft的人类ATII/ATI感染模型，并用于表征细菌的内化、复制和运输。其次，将使用RNA-seq技术和在A549细胞中筛选包含20,000个成员的Schu S4：：HIMAR1转座子突变文库来鉴定肺细胞内Ft表达的细菌决定簇，以寻找细胞毒性抑制方面的缺陷。最后，上面确定的肺泡细胞表达的基因的子集将被靶向干扰，并使用各种体外和体内感染模型系统进行表征。总而言之，这些研究有望为ATII/ATI肺泡细胞在Ft发病机制中的重要性提供新的见解。这些信息可能导致确定新的治疗靶点或候选疫苗来对抗Ft感染。