Role of sodium-thiocyanate symporters in the antibacterial airway host defense

硫氰酸钠同向转运体在抗菌气道宿主防御中的作用

• 批准号: 7581377
• 负责人: Botond Banfi
• 金额: $ 33.75万
• 依托单位: UNIVERSITY OF IOWA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7581377
• 关键词: Acids; Adenoviruses; Affinity; Anti-Bacterial Agents; Antibiotics; Antiviral Agents; Apical; Bacteria; Bacterial Infections; Biochemical; Biological; Biological Assay; Cell Culture Techniques; Cell membrane; Cells; Cellular biology; Chemicals; Clinical; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Data; Disease; Drug Kinetics; Enzymes; Epithelial; Epithelial Cells; Epithelium; FDA approved; Gene Delivery; Generations; Gland; HOI; Harvest; Homologous Gene; Host Defense; Host Defense Mechanism; Human; Human Activities; Hydrogen Peroxide; Immune; Immunity; In Vitro; Infection; Ingestion; Intake; Iodides; Ion-Exchange Chromatography Procedure; Knowledge; Liquid substance; Lung; Mediating; Methods; Modeling; Molecular; Molecular Biology; Natural Immunity; Nose; OSCN-; Oral; Organ; Oxidases; Oxidation-Reduction; Patients; Pattern; Potassium Iodide; Production; Proteins; Publishing; RNA Interference; Reaction; Reactive Oxygen Species; Recurrence; Role; Route; Serous; Serum; Surface; System; Tablets; Testing; Therapeutic; Xenograft Model; Xenograft procedure; airway epithelium; antimicrobial; bactericide; base; catalase; cell type; cystic fibrosis airway epithelia; cystic fibrosis patients; defined contribution; healthy volunteer; human subject; hypothiocyanite; in vivo; in vivo Model; inhibitor/antagonist; insight; killings; lactoperoxidase; novel; oxidation; pathogen; prevent; public health relevance; research study; sodium thiocyanate; sodium-iodide symporter; symporter; tool; uptake

DESCRIPTION (provided by applicant): Recent data suggest that an oxidative host defense mechanism may prevent bacterial infections in the airway. This novel mechanism kills bacteria by producing bactericidal hypothiocyanite (OSCN-) in a lactoperoxidase (LPO)-catalyzed reaction: H2O2 + SCN- ? OSCN-. Accordingly, OSCN- production requires LPO secretion by the submucosal glands, H2O2 generation by the dual oxidase (Duox) enzymes of surface epithelia, and SCN transport across the airway epithelium. SCN- transport is mediated predominantly by CFTR in the apical plasma membrane. Thus, CF airway epithelia have reduced SCN- secretion and, consequently, are defective in OSCN- production and bacterial killing. The molecular identity of the basolateral SCN- transporter remains to be determined in an in vivo system. This gap in the current knowledge is hindering efforts to enhance oxidative innate immunity in the lung and to understand the clinical importance of the oxidative system. Pharmacological data suggest that - at least in cultured airway epithelia - the sodium-iodide symporter (NIS) is the basolateral SCN- transporter. However, our preliminary results indicate that both NIS and its close homolog slc5a8 may mediate SCN- secretion by airway epithelial cells. Furthermore, slc5a8 is expressed in the surface epithelium, whereas NIS is localized primarily to the serous cells of the submucosal glands. Our central hypothesis is that compartmentalized expression of slc5a8 and NIS creates two routes of SCN- secretion in the airway, and that the Duox/LPO/SCN- system is critical for antibacterial immunity. We also hypothesize that the secretory route involving NIS can be utilized to deliver iodide (I-) to the airway surface fluid, where Duox/LPO oxidizes I- to hypoiodous acid (H2O2 + I- ? HOI), a very effective antibacterial and antiviral molecule. Thus, our objective is to define the functional relevance of NIS and slc5a8 to SCN- secretion in the airway, and to evaluate the biological importance of the Duox/LPO enzymes to airway innate immunity in the presence of SCN- and I-. We propose the following Specific Aims: 1. Define the importance of compartmentalized NIS and slc5a8 expression to SCN- secretion in the airways. 2. Evaluate the importance of the Duox/LPO/SCN- system to bacterial killing in the human airway. 3. Explore the pharmacokinetics of I- in the airway, and the effect of airway fluid I- on bacterial killing. We will pursue these aims using 1) in vivo approaches such as human airway xenografts and human subject studies; 2) cell biology tools including primary airway epithelial cultures, adenovirus-mediated gene delivery, and bacterial killing assays; 3) molecular biology methods such as RNA interference; and 4) biochemical assays including ion-exchange chromatography and colorimetric redox reactions. This project is expected to identify the SCN- transporters that are required for OSCN- mediated bacterial killing in the airway, as well as to provide insight into the in vivo importance and therapeutic potential of OSCN-- and HOI-mediated host defense, which until now has not been explored. PUBLIC HEALTH RELEVANCE. The proposed project will explore a novel innate immune mechanism of airways that eliminates bacteria via the production of reactive oxygen species. Ex vivo experiments have demonstrated that this novel mechanism is defective in airway epithelia of patients with cystic fibrosis. Therefore, the proposed studies may ascertain the therapeutic potential of restoring or enhancing the oxidative host defense system in cystic fibrosis and other diseases that are characterized by recurrent airway infections.

描述（由申请人提供）：最近的数据表明，氧化宿主防御机制可能会防止气道中的细菌感染。这种新型机制通过在乳酸氧化酶（LPO）催化反应中产生杀菌性下氨酸（OSCN-）来杀死细菌：H2O2 + SCN-？ OSCN-。因此，OSCN的产生需要粘膜下腺的LPO分泌，表面上皮的双氧化酶（DUOX）酶产生H2O2，以及横跨气道上皮的SCN转运。 SCN转运主要由CFTR在顶端质膜中介导。因此，CF气道上皮降低了SCN分泌，因此在OSCN生产和细菌杀伤中有缺陷。基底外侧SCN-转运蛋白的分子身份仍有待确定在体内系统中。当前知识的这一差距正在阻碍增强肺部氧化先天免​​疫的努力，并了解氧化系统的临床重要性。药理学数据表明 - 至少在培养的气道上皮中 - 碘化钠分类剂（NIS）是基底外侧SCN-转运蛋白。但是，我们的初步结果表明，NIS及其紧密的同源性SLC5A8都可以介导气道上皮细胞的SCN分泌。此外，SLC5A8在表面上皮中表达，而NIS主要定位于粘膜下腺的浆液细胞。我们的中心假设是SLC5A8和NIS的分隔表达在气道中创建了两种SCN分泌途径，并且DUOX/LPO/SCN-系统对于抗菌免疫至关重要。我们还假设，涉及NIS的分泌途径可用于将碘化物（I-）递送到气道表面流体，DUOX/LPO在其中氧化I-至低约二酸（H2O2 + I- HOI），一种非常有效的抗体和抗病毒分子。因此，我们的目标是定义NIS和SLC5A8与气道中的SCN分泌的功能相关性，并评估DUOX/LPO酶在SCN和I-存在下气道先天免疫的生物学重要性。我们提出以下特定目的：1。定义隔室化的NIS和SLC5A8表达对气道中SCN分泌的重要性。 2。评估DUOX/LPO/SCN-系统对人类气道细菌杀死的重要性。 3。探索气道中I-的药代动力学以及气道液I-对细菌杀死的影响。我们将使用1）人体气道异种移植和人类学科研究等体内方法来追求这些目标； 2）细胞生物学工具，包括原发性气道上皮培养物，腺病毒介导的基因递送和细菌杀戮测定； 3）分子生物学方法，例如RNA干扰； 4）包括离子交换色谱和比色氧化还原反应在内的生化测定。预计该项目将确定OSCN介导的细菌在气道中所需的SCN-转运蛋白，并洞悉OSCN的体内重要性和治疗潜力以及HOI介导的宿主防御，直到现在尚未探索。公共卫生相关性。拟议的项目将探索一种新型的气道先天免疫机制，该机制通过产生活性氧，从而消除细菌。离体实验表明，这种新型机制在囊性纤维化患者的气道上皮中有缺陷。因此，拟议的研究可能会确定在囊性纤维化中恢复或增强氧化宿主防御系统的治疗潜力，以及以复发性气道感染为特征的其他疾病。