Protective activity of the lectin-like domain of TNF in permeability edema

TNF 凝集素样结构域对通透性水肿的保护活性

• 批准号: 8308361
• 负责人: Rudolf Lucas
• 金额: $ 37.13万
• 依托单位: AUGUSTA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2014-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8308361
• 关键词: 5 year old; Affect; Affinity Chromatography; Agonist; Alveolar; Antibiotic Therapy; Arginine; Bacterial Infections; Bacterial Pneumonia; Bacterial exotoxin; Cause of Death; Cells; Cessation of life; Child; Cholesterol; Complication; Data; Development; Disease; Down-Regulation; Edema; Elderly; Enzyme Uncoupling; Enzymes; Epithelial; Epithelial Cells; Event; Exotoxins; Functional disorder; Generations; Goals; Human; In Vitro; Infection; Influenza A virus; Knock-in Mouse; Lead; Lectin; Life; Liquid substance; Listeria monocytogenes; Listeria monocytogenes hlyA protein; Lung; Mediating; Membrane; Mus; NADPH Oxidase; Patients; Peptide N-glycohydrolase F; Peptides; Permeability; Phosphorylation; Play; Pneumonia; Production; Protein Kinase C; Research; Role; Secondary to; Small Interfering RNA; Sodium; Sodium Channel; Sterility; Streptococcus pneumoniae; Streptococcus pneumoniae plY protein; TNF gene; Testing; Tissues; Toxin; Vascular Permeabilities; Virulence Factors; Virus Diseases; Wild Type Mouse; aged; arginase; base; combat; epithelial Na+ channel; improved; in vivo; inhibitor/antagonist; mortality; mouse model; mutant; novel therapeutic intervention; novel therapeutics; patch clamp; programs; protein expression; public health relevance; rhoA GTP-Binding Protein; uptake

DESCRIPTION (provided by applicant): Severe pneumonia is the leading single cause of mortality in children aged less than five years worldwide and the sixth leading cause of death in seniors over age 65 in the US. Streptococcus pneumoniae represents the main etiological agent of the disease. Moreover, mortality after influenza A virus (IAV) infection has been suggested to be mainly due to secondary pneumoccocal infections. Intriguingly, death in pneumococcal-induced pneumonia can occur days after initiation of antibiotic therapy, when tissues are sterile and the pneumonia is clearing. This mortality correlates with the presence of pneumococcal virulence factors, the most important one of which is the pore-forming toxin pneumolysin (PLY). A major complication of severe pneumonia is permeability edema, characterized by a dramatically increased pulmonary endothelial hyperpermeability and an impaired alveolar liquid clearance (ALC). The enzyme PKC-(, which is activated by PLY-mediated Ca2+ influx, has recently been suggested to be implicated in the downregulation of the epithelial sodium channel (ENaC) expression, as well as in endothelial barrier dysfunction. Our preliminary data demonstrate that the lectin-like domain of TNF, mimicked by the TIP peptide, is able to blunt PLY-induced PKC-( activation, endothelial hyperpermeability and ALC dysfunction. Our overall hypothesis is that the lectin-like domain of TNF restores alveolar liquid clearance and improves barrier integrity during G+-infection- associated pneumonia, by means of blunting exotoxin-mediated PKC-( activation. We will test three specific aims to this purpose. First, we will unravel the mechanism by which the TIP peptide restores epithelial sodium channel activity and expression in PLY-treated alveolar epithelial cells. Second, we will determine at which step(s) in the cascade of events leading to PLY-induced endothelial dysfunction, the TIP peptide intervenes and what is the most upstream event affected by the peptide. Third, we will test the hypothesis that the lectin-like domain of TNF restores ALC and preserves pulmonary endothelial barrier integrity in PLY-treated mice, upon blunting PKC-( activation, making use of Triple mTNF knock-in mice, expressing a lectin-deficient mutant of TNF. The results of this research program can thus provide important information about mechanisms leading to permeability edema and dysfunctional ALC during bacterial pneumonia and can thus lead to the identification of novel therapeutic strategies. PUBLIC HEALTH RELEVANCE: One of the major complications of severe bacterial pneumonia is permeability edema, characterized by endothelial hyperpermeability and a dysfunctional alveolar liquid clearance (ALC) capacity. Our recent data from mouse models and in vitro studies using human endothelial and airway epithelial cells indicate that the TNF-derived TIP peptide, mimicking its lectin-like domain, restores ALC and endothelial barrier integrity upon treatment with the G+ bacterial exotoxin pneumolysin (PLY). This project aims to demonstrate that the protective activity of the TIP peptide in bacterial pneumonia relies on its capacity to blunt PLY-induced Protein Kinase C-( activation, which plays a crucial role in both the induction of endothelial hyperpermeability and in the down-regulation of the epithelial sodium channel, as such providing a basis for novel therapeutic strategies to combat severe bacterial pneumonia.

描述（由申请人提供）：重症肺炎是全球五岁以下儿童死亡的首要单一原因，也是美国 65 岁以上老年人死亡的第六大原因。肺炎链球菌是该疾病的主要病原体。此外，甲型流感病毒（IAV）感染后的死亡被认为主要是由于继发性肺炎球菌感染。有趣的是，肺炎球菌引起的肺炎的死亡可能发生在开始抗生素治疗几天后，此时组织已无菌且肺炎已痊愈。这种死亡率与肺炎球菌毒力因子的存在相关，其中最重要的一种是成孔毒素肺炎球菌溶血素（PLY）。重症肺炎的一个主要并发症是渗透性水肿，其特征是肺内皮细胞渗透性显着增加和肺泡液体清除率（ALC）受损。 PKC-( 由 PLY 介导的 Ca2+ 内流激活，最近被认为与上皮钠通道 (ENaC) 表达下调以及内皮屏障功能障碍有关。我们的初步数据表明， TNF 的凝集素样结构域（由 TIP 肽模拟）能够减弱 PLY 诱导的 PKC-( 激活、内皮细胞通透性过高和 ALC 功能障碍。假设是 TNF 的凝集素样结构域通过减弱外毒素介导的 PKC-( 激活，恢复肺泡液体清除率并改善 G+ 感染相关性肺炎期间的屏障完整性。我们将为此目的测试三个具体目标。首先，我们将揭示 TIP 肽恢复 PLY 处理的肺泡上皮细胞中上皮钠通道活性和表达的机制 其次，我们将确定级联反应中的哪个步骤。在导致 PLY 诱导的内皮功能障碍的事件中，TIP 肽会进行干预，而受该肽影响的最上游事件是什么。第三，我们将测试以下假设：在 PLY 处理的小鼠中，TNF 的凝集素样结构域在削弱 PKC-( 激活，利用三重​​ mTNF 敲入小鼠，表达凝集素-因此，该研究项目的结果可以提供有关细菌性肺炎期间导致通透性水肿和 ALC 功能障碍的机制的重要信息，从而可以确定新的治疗策略。 公共卫生相关性：严重细菌性肺炎的主要并发症之一是渗透性水肿，其特征是内皮细胞渗透性过高和肺泡液体清除（ALC）能力功能障碍。我们最近从小鼠模型和使用人内皮和气道上皮细胞进行的体外研究中获得的数据表明，TNF 衍生的 TIP 肽模仿其凝集素样结构域，在用 G+ 细菌外毒素肺炎球菌溶血素 (PLY) 治疗后可恢复 ALC 和内皮屏障完整性。 。该项目旨在证明 TIP 肽在细菌性肺炎中的保护活性依赖于其减弱 PLY 诱导的蛋白激酶 C-( 激活的能力，这在诱导内皮细胞通透性过高和下调中发挥着至关重要的作用上皮钠通道的变化，因此为对抗严重细菌性肺炎的新治疗策略提供了基础。