An air-liquid interface system to study Bordetella pertussis interactions with respiratory epithelia

研究百日咳博德特氏菌与呼吸道上皮细胞相互作用的气液界面系统

• 批准号: 10665943
• 负责人: Eric T Harvill
• 金额: $ 22.09万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-14 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10665943
• 关键词: Adherence; Affect; Air; Anti-Inflammatory Agents; Apical; Appearance; Architecture; Bacteria; Bacterial Adhesins; Bacterial Attachment Site; Behavior; Binding; Biological Assay; Biology; Bordetella; Bordetella pertussis; Cell Culture System; Cell Culture Techniques; Cell Line; Cells; Cessation of life; Cilia; Clinical; Clinical Data; Colloids; Columnar Epithelium; Complex; Culture Media; Cultured Cells; Cytopathology; Data; Disease; Epithelial Cells; Epithelium; Fibroblasts; Genetic; Goblet Cells; Growth; Health; Human; Immunity; In Vitro; Incidence; Individual; Infection; Infiltration; Inflammatory; Inflammatory Response; Interruption; Liquid substance; Lung; Macrophage; Mammalian Cell; Measurable; Measures; Mediating; Microbiology; Mucous Membrane; Mucous body substance; Mus; Nutrient; Pathogenesis; Penetration; Pertussis; Pertussis Toxin; Pertussis Vaccine; Population Heterogeneity; Predisposition; Production; Respiratory System; Role; Side; Signal Transduction; Site; Structure; Surface; System; Targeted Research; Techniques; Tight Junctions; Time; Tissues; Trachea; United States National Institutes of Health; Vaccines; Virulence Factors; Viscosity; airway epithelium; cell behavior; cell injury; cell type; cytotoxic; cytotoxicity; emerging pathogen; experience; experimental study; feeding; in vivo; monolayer; novel vaccines; pathogen; pathogenic bacteria; success; transmission process; unvaccinated

Bordetella pertussis, the bacterial pathogen responsible for “whooping cough” causes an estimated 24 million cases of vaccine-preventable illness per year, resulting in an excess of 170,000 deaths annually. Importantly, the incidence of whooping cough in nations with high vaccine coverage is on the rise, attributed to asymptomatic transmission that is enabled by the imperfect and waning immunity of current acellular pertussis vaccines. Due to these and other factors, both the CDC and NIH have listed B. pertussis as a priority (re)emerging pathogen of high concern. B. pertussis efficiently colonizes and grows within the human respiratory tract, requiring that it access and cross mucus and sol layers to find, tightly attach to and grow on ciliated epithelial cells. These abilities are critical to its remarkable success, but are difficult to study in detail in vivo. They have been studied primarily in submerged cell culture, with monolayers of host cells and B. pertussis all submerged in rich mammalian cell growth media. These conditions do not replicate the structure or function of the columnar airway epithelia and completely lack the overlying mucus, sol and air-interface of a natural airway, and the milieu in which B. pertussis naturally grows. Our team has decades of experience with Bordetella spp and with polarized primary culture from human broncho-tracheal tissues which contains representative cell types, most importantly including cilia beating within protective mucus and sol layers. Here, for the first time, we will use this air-liquid interface (ALI) system, combined with techniques in genetics and biology of B. pertussis, to probe the roles of key bacterial factors in host-pathogen interactions with realistic human respiratory epithelia. Specifically, we propose three aims to (1) identify factors that mediate infiltration through mucosal layers and enable bacterial growth on ciliated epithelia, (2) define the effect of B. pertussis factors on inducing and/or modulating epithelial cell produced pro/anti-inflammatory signals, and (3) determine how B. pertussis damages cells and disrupts the epithelial barrier. The data generated from these studies will reveal the roles of specific B. pertussis factors in various measurable aspects of their interactions with ciliated respiratory epithelia and should inform the choice of new vaccine targets capable of interrupting the airway interactions.

百日咳杆菌是引起“百日咳”的细菌病原体， 估计每年有2 400万例疫苗可预防的疾病， 每年有17万人死亡。重要的是，百日咳的发病率在高 疫苗覆盖率正在上升，这是由于无症状传播， 目前无细胞百日咳疫苗的免疫力不完善和减弱。由于这些和其它 CDC和NIH都列出了B。百日咳作为一个优先（重新）出现的病原体， 关心B。百日咳在人呼吸道内有效地定殖和生长， 需要它进入并穿过粘液和溶胶层，以找到，紧紧附着并生长 纤毛上皮细胞这些能力对于它的非凡成功至关重要，但也很困难 在体内进行详细研究。它们主要在浸没细胞培养中进行研究， 宿主细胞单层和B.百日咳疫苗全部浸没在丰富的哺乳动物细胞生长培养基中。 这些条件不能复制气道柱状上皮的结构或功能 并且完全缺乏天然气道的覆盖粘液、溶胶和空气界面， B.百日咳会自然生长。我们的团队拥有数十年的经验， 博德特氏菌属和来自人支气管-气管组织的极化原代培养物， 包含代表性的细胞类型，最重要的是包括纤毛跳动内的保护 粘液和溶胶层。在这里，我们将首次使用这种气液界面（ALI）系统， 结合B的遗传学和生物学技术。百日咳，探讨作用的关键 宿主-病原体相互作用中的细菌因子与真实的人类呼吸道上皮细胞。 具体而言，我们提出了三个目标：（1）确定介导渗透的因素， 粘膜层并使细菌在纤毛上皮细胞上生长，（2）确定B的作用。 百日咳因子对诱导和/或调节上皮细胞产生促炎/抗炎作用 信号，以及（3）确定B.百日咳破坏细胞并破坏上皮屏障。 这些研究产生的数据将揭示特定B的作用。百日咳因子 它们与纤毛呼吸道上皮细胞相互作用的各种可测量方面， 告知能够中断气道相互作用的新疫苗靶点的选择。