Regulation of Airway Lactoperoxidase Host Defense

气道乳过氧化物酶宿主防御的调节

• 批准号: 7386660
• 负责人: Gregory E. Conner
• 金额: $ 33.43万
• 依托单位: UNIVERSITY OF MIAMI SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-03-01 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7386660
• 关键词: Address; Aftercare; Agonist; Air; Alginates; Anti-Bacterial Agents; Antibiotics; Apical; Bacteria; Biological Assay; Bronchiectasis; Cell membrane; Chelating Agents; Chemistry; Chronic; Clinical; Computer Simulation; Consumption; Controlled Study; Count; Cyclic AMP; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Defect; Depressed mood; Disease; Down-Regulation; EF-Hand Domain; Elements; Enzymes; Epithelial; Epithelial Cells; Epithelium; Exhalation; Failure; Funding; Genes; Genetic; Genetic Transcription; Grant; Host Defense; Human; Hydrogen Peroxide; Hypochlorite; Indium; Individual; Infection; Inflammation; Ionophores; Kinetics; Lead; Liquid Chromatography; Liquid substance; Maps; Mediating; Messenger RNA; Metabolism; Modeling; Morphology; NADPH Oxidase; OSCN-; Pathogenesis; Patients; Peroxidase; Peroxidases; Peroxonitrite; Phenotype; Physiological; Play; Primary Ciliary Dyskinesias; Production; Proteins; Pseudomonas aeruginosa; Publishing; Race; Rate; Reactive Oxygen Species; Recurrence; Regulation; Relative (related person); Research Personnel; Resistance; Reverse Transcriptase Polymerase Chain Reaction; Role; Sheep; Signal Transduction; Source; Spectrometry, Mass, Electrospray Ionization; Sterility; Stimulus; Stress; Sulfur; Superoxides; Surface; System; Taurine; Temperature; Testing; Thapsigargin; Thiocyanates; Tyrosine; Up-Regulation; Vesicle; Virus; Work; Xanthine Oxidase; Xanthines; airway epithelium; airway hyperresponsiveness; airway inflammation; airway surface liquid; base; cystic fibrosis airway; cystic fibrosis airway epithelia; cystic fibrosis patients; disorder control; enzyme activity; fungus; grasp; hypothiocyanite; in vivo; insight; lactoperoxidase; mRNA Expression; mRNA Stability; mucoid; neutrophil; novel therapeutics; nuclear runoff assay; oxidation; programs; promoter; prospective; research study; response; thiocyanate; xanthine

The lactoperoxidase (LPO) system is a significant contributor to airway host defense against bacteria, viruses and fungi and also is a major consumer of airway H2O2 that in turn is associated with airway inflammation. A loss of LPO activity may lead to increased airway infection, while any increase in LPO could have injurious consequences. This application will test the hypothesis that all components of the LPO system, including LPO enzyme expression and secretion, SCN~ transport across the epithelium and H2O2 production, are coordinately regulated and that an absence of the LPO substrate, SCN~, in cystic fibrosis (CF) may contribute to pathogenesis. This hypothesis is supported by a computational model of LPO system activity in airway surface liquid that predicts any unbalance in the components will render the system inadequate for host defense. The hypothesis and model are supported by preliminary data showing: a) CF airway epithelia lack normal transport of LPO's substrate, SCN~, to the apical surface that may contribute to airway infections and inflammation; b) airway epithelia produce H2O2 through Duox, an NADPH oxidase that may be regulated by intracellular [Ca2+]; c) reactive oxygen species (ROS) increase expression of LPO; and d) Duox and LPO mRNAs are up-regulated in CF airway epithelia. Specific Aim 1 will address regulation of Duox H2O2 production in normal and CF airway epithelial cells by changes in [Ca2+]j, cAMP, SCN~ or proinflammatory and infection related stimuli. Specific Aim 2 will test the hypothesis that defective SCN~ transport in CF airway epithelia results in decreased SCN~ in CF airway secretions in vivo that then may compromise host defense. SCN~ and peroxidase endproducts will be determined in exhaled breath condensate using liquid chromatography in tandem with electrospray ionization-mass spectrometry. Specific Aim 3 will test the hypothesis that ROS and/or SCN" increases LPO mRNA expression in normal and CF airway epithelia by using turnover studies and nuclear runoff assays. The 5' end of LPO mRNA will be mapped by RACE and promoter elements in upstream sequences identified. Specific Aim 4 will test the hypothesis that depressed CF airway SCN" levels contribute to neutrophil-mediated oxidative damage and to the phenotype of colonizing bacteria observed in CF airways. Lav Summary: These experiments will provide new information about physiological defects in cystic fibrosis patients that may lead to new therapeutic approaches to treating this disease.

乳过氧化物酶 (LPO) 系统是气道宿主防御细菌的重要贡献者， 病毒和真菌，也是气道 H2O2 的主要消耗者，而 H2O2 又与气道相关 炎。 LPO 活性的丧失可能会导致气道感染增加，而 LPO 的任何增加都可能导致气道感染增加。 造成有害后果。该应用程序将测试 LPO 的所有组件的假设 系统，包括 LPO 酶的表达和分泌、SCN~ 跨上皮的转运和 H2O2 囊性纤维化中 LPO 底物 SCN~ 的缺失是协调调节的 (CF)可能有助于发病机制。 LPO 系统的计算模型支持了这一假设 气道表面液体的活动预测组件中的任何不平衡都会导致系统 不适合宿主防御。该假设和模型得到初步数据的支持，显示： a) CF 气道上皮缺乏 LPO 底物 SCN~ 到顶端表面的正常运输，这可能有助于 气道感染和炎症； b) 气道上皮通过 Duox（一种 NADPH 氧化酶）产生 H2O2 可能受细胞内[Ca2+]调节； c) 活性氧（ROS）增加LPO的表达；和 d) Duox 和 LPO mRNA 在 CF 气道上皮细胞中上调。具体目标 1 将解决以下问题的监管 通过改变 [Ca2+]j、cAMP、SCN~ 或 正常和 CF 气道上皮细胞产生 Duox H2O2 促炎和感染相关的刺激。具体目标 2 将检验 SCN 缺陷的假设~ CF 气道上皮中的转运导致体内 CF 气道分泌物中的 SCN~ 减少，然后可能 损害宿主防御。 SCN~ 和过氧化物酶终产物将在呼出气中测定 使用液相色谱法与电喷雾电离质谱联用进行冷凝。具体的 目标 3 将检验 ROS 和/或 SCN”增加正常和 CF 中 LPO mRNA 表达的假设 通过使用周转研究和核径流测定来检测气道上皮细胞。 LPO mRNA 的 5' 末端将是 由上游序列中的 RACE 和启动子元件进行定位。具体目标 4 将测试 假设 CF 气道 SCN" 水平降低会导致中性粒细胞介导的氧化损伤，并导致 CF气道中观察到的定植细菌的表型。 Lav 摘要：这些实验将提供 关于囊性纤维化患者生理缺陷的新信息可能会带来新的治疗方法 治疗这种疾病的方法。