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.

百日咳博德特氏菌是导致“百日咳”的细菌病原体，会导致 据估计，每年有 2400 万例疫苗可预防的疾病，导致 重要的是，百日咳发病率高的国家每年有 17 万人死亡。 疫苗覆盖率正在上升，归因于无症状传播 由于这些和其他原因，当前无细胞百日咳疫苗的免疫力不完善且减弱。 CDC 和 NIH 均已将百日咳博德特氏菌列为高致病性（重新）出现的优先病原体 百日咳博德特氏菌在人类呼吸道内有效定植和生长， 要求它进入并穿过粘液和溶胶层以找到、紧密附着并生长 纤毛上皮细胞的这些能力对其取得显着的成功至关重要，但也很困难。 主要在深层细胞培养中对它们进行了详细的体内研究。 宿主细胞和百日咳博德特氏菌的单层全部浸没在丰富的哺乳动物细胞生长培养基中。 这些条件不能复制柱状气道上皮的结构或功能 并且完全缺乏自然气道的粘液、溶胶和空气界面，并且 我们的团队在百日咳杆菌自然生长的环境中拥有数十年的经验。 博德特氏菌 (Bordetella spp) 和来自人支气管气管组织的极化原代培养物 包含代表性细胞类型，最重要的是包括保护性范围内的纤毛跳动 在这里，我们将首次使用这种气液界面（ALI）系统， 结合百日咳博德特氏菌遗传学和生物学技术，探讨关键基因的作用 宿主-病原体与现实人类呼吸道上皮细胞相互作用中的细菌因素。 具体来说，我们提出三个目标：（1）确定通过以下方式介导渗透的因素： 粘膜层并使细菌能够在纤毛上皮上生长，(2) 定义 B. 诱导和/或调节上皮细胞产生促/抗炎作用的百日咳因子 信号，(3) 确定百日咳博德特氏菌如何损害细胞并破坏上皮屏障。 这些研究产生的数据将揭示特定百日咳博德特氏菌因子在 它们与纤毛呼吸道上皮细胞相互作用的各个可测量的方面，并且应该 告知选择能够中断气道相互作用的新疫苗靶标。