The Role of Hepoxilin A3 in Neutrophil Breach of the Infected Airway Mucosa

赫泊西林 A3 在中性粒细胞突破感染气道粘膜中的作用

• 批准号: 8813522
• 负责人: BRYAN P HURLEY
• 金额: $ 43.93万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-08 至 2016-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8813522
• 关键词: AIDS/HIV problem; Acute; Age; Air; Animal Model; Anti-Inflammatory Agents; Anti-inflammatory; Apical; Arachidonate 12-Lipoxygenase; Arachidonic Acids; Automobile Driving; Biological Models; Blood Circulation; CXC Chemokines; Cell Culture Techniques; Charge; Child; Chronic; Complement 5a; Complex; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Development; Disease; Eicosanoids; Enzymes; Epithelial; Epithelial Cells; Exhibits; Failure; Family; Future; Genes; Health; Human; Immigration; Immunologic Surveillance; In Vitro; Inborn Genetic Diseases; Individual; Infection; Inflammation; Inflammation Mediators; Inflammatory; Injury; Intervention; Investigation; Knowledge; Leukotriene B4; Leukotrienes; Lipids; Liquid substance; Lung; Lung Inflammation; Lung diseases; Malaria; Malignant Neoplasms; Membrane; Modeling; Molecular; Mucous Membrane; Mus; Myocardial Infarction; Neutrophil Infiltration; Organism; Outcome; Pathology; Pathway interactions; Phospholipase; Phospholipase A2; Phospholipids; Play; Pneumonia; Process; Production; Prostaglandins; Protein Isoforms; Pseudomonas aeruginosa; Publications; Recruitment Activity; Resolution; Respiratory System; Respiratory tract structure; Role; Severities; Signal Transduction; Site; Staging; Stroke; Structure of parenchyma of lung; Surface; Therapeutic; Tissues; Work; airway epithelium; base; design; in vitro Model; in vivo; inflammatory lung disease; information gathering; member; migration; monolayer; mouse model; neutrophil; novel; pathogen; response; therapeutic target

DESCRIPTION (provided by applicant): The mucosal epithelial surface serves as a protective barrier to external threats both in the physical sense and as a vital contributor to immune surveillance and orchestration. One of the hallmarks of infectious inflammatory mucosal disease in the respiratory tract is massive mobilization of neutrophils across the mucosal barrier into the airway lumen. Encounter with a pathogenic organism triggers a process whereby neutrophils migrate from the bloodstream to airspace in order to directly confront mucosal invaders through engulfment and release of noxious substances. Neutrophil accumulation can become over-exuberant resulting in host lung tissue damage as with the infectious process involving Pseudomonas aeruginosa in the diseases pneumonia and cystic fibrosis. The final step in neutrophil recruitment from the bloodstream to the airway involves migration across the epithelial barrier of the mucosal surface. Our studies have revealed a molecular mechanism that underlies this phenomenon. Using lung epithelial cells grown on permeable Transwell filters, we have modeled neutrophil trans-epithelial migration in response to infection. Our studies have demonstrated that treatment of lung epithelial monolayers with P. aeruginosa results in activation of phospholipase A2, which causes the release of arachidonic acid. Arachidonic acid is then converted to the neutrophil chemo-attractant hepoxilin A3 by the actions of 12-lipoxygenases / hepoxilin synthases. Hepoxilin A3 is released at the apical surface of lung epithelial monolayers and guides neutrophils across the epithelial barrier. In the current application, we aim to further explore the mechanism underlying hepoxilin A3 production in mouse and human lung epithelial cells by identifying both the specific phospholipase A2 and 12-lipoxygenase / hepoxilin synthase enzyme(s) responsible for generating hepoxilin A3 upon infection with pathogen. Furthermore, we propose to establish air-liquid interface cultures of primary mouse and human lung epithelial cells as well as lung epithelial cells with defective cystic fibrosis transmembrane conductance regulator. The ability of P. aeruginosa to instigate hepoxilin A3 production and neutrophil trans-epithelial migration will be assessed in these air-liquid models. Finally, we will evaluate hepoxilin A3 as a neutrophil chemo-attractant operating at the airway mucosal surface in vivo employing a mouse model of P. aeruginosa pneumonia. Multiple interventions to interfere with hepoxilin A3 synthesis, stability, and action will be utilized and correlated with outcomes that include severity of pathological tissue damage, expression of 12-lipoxygenases, and neutrophil accumulation in the infected airspace. These studies aim to further elucidate the role of this newly appreciated eicosanoid neutrophil chemo-attractant termed hepoxilin A3 that is produced at various mucosal surfaces. A better understanding of this novel innate inflammatory pathway holds tremendous promise towards unveiling a distinct class of unexplored targets to exploit therapeutically in order to alleviate destructive lung inflammation.

描述（由申请人提供）：粘膜上皮表面在物理意义上作为外部威胁的保护屏障，并作为免疫监视和协调的重要贡献者。呼吸道感染性炎性粘膜疾病的特征之一是嗜中性粒细胞大量动员穿过粘膜屏障进入气道腔。与致病生物体的相遇触发了一个过程，其中中性粒细胞从血流迁移到空气中，以便通过吞噬和释放有毒物质直接对抗粘膜入侵者。中性粒细胞的积累可能变得过度旺盛，导致宿主肺组织损伤，正如肺炎和囊性纤维化疾病中涉及铜绿假单胞菌的感染过程一样。中性粒细胞从血流募集到气道的最后一步涉及穿过粘膜表面的上皮屏障的迁移。我们的研究揭示了这种现象背后的分子机制。使用可渗透的Transwell过滤器上生长的肺上皮细胞，我们模拟了中性粒细胞跨上皮迁移感染。我们的研究已经证明，用铜绿假单胞菌处理肺上皮单层导致磷脂酶A2的活化，其引起花生四烯酸的释放。花生四烯酸然后通过12-脂氧合酶/肝氧素酶的作用转化为中性粒细胞化学引诱物肝氧素A3。肝氧素A3在肺上皮单层的顶端表面释放，并引导中性粒细胞穿过上皮屏障。在本申请中，我们的目的是通过鉴定在病原体感染时负责产生肝氧素A3的特异性磷脂酶A2和12-脂氧合酶/肝氧素合酶来进一步探索小鼠和人肺上皮细胞中肝氧素A3产生的潜在机制。此外，我们建议建立原代小鼠和人肺上皮细胞以及具有缺陷性囊性纤维化跨膜传导调节因子的肺上皮细胞的气液界面培养。将在这些空气-液体模型中评估铜绿假单胞菌激发肝氧素A3产生和中性粒细胞跨上皮迁移的能力。最后，我们将评估肝氧素A3作为一种中性粒细胞化学引诱剂在体内气道粘膜表面采用小鼠模型的铜绿假单胞菌肺炎。将采用多种干预措施干扰肝氧素A3的合成、稳定性和作用，并将其与包括病理组织损伤严重程度、12-脂氧合酶表达和感染空域中性粒细胞蓄积在内的结局相关。这些研究旨在进一步阐明这种新认识的类花生酸中性粒细胞化学引诱物（称为肝氧素A3）在各种粘膜表面产生的作用。更好地了解这种新的先天性炎症通路，对于揭示一种独特的未探索的靶点具有巨大的希望，可以在治疗上加以利用，以减轻破坏性的肺部炎症。