Alterations In Pulmonary Immune Function And Host Resistance In COX Null Mice

COX 无效小鼠肺免疫功能和宿主抵抗力的变化

• 批准号: 8336532
• 负责人: Darryl C Zeldin
• 金额: $ 76.53万
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8336532
• 关键词: Acute; Allergens; Allergic; Alveolar Cell; Alveolar Macrophages; Anabolism; Appearance; Attenuated; Biochemical; Bleomycin; Body Temperature Changes; Body Weight; Breathing; Breeding; Bronchoalveolar Lavage Fluid; Bronchoconstriction; CCL17 gene; Cell Adhesion Molecules; Cell Count; Cells; Clara cell; Data; Defect; Dinoprostone; Distal; Endotoxins; Eotaxin; Equilibrium; Exhibits; Fibrosis; Genes; Genotype; Goals; Histopathology; Host resistance; Human; Hydroxyproline; IgE; Immune response; In Vitro; Infection; Inflammation; Inflammatory; Inflammatory Response; Inflammatory Response Pathway; Influenza; Intercellular adhesion molecule 1; Interleukin-13; Interleukin-4; Interleukin-5; Knock-out; Knockout Mice; Leukotriene B4; Leukotrienes; Lipopolysaccharides; Liquid substance; Lung; Lung Compliance; Lung Inflammation; Lymphocyte; MUC5AC gene; Mechanics; Modeling; Mus; Ovalbumin; PTGS2 gene; Pharmaceutical Preparations; Physiological; Prostaglandin Production; Prostaglandin-Endoperoxide Synthase; Prostaglandins; Proteins; Relative (related person); Respiratory physiology; Role; Stimulus; T-Lymphocyte; Transgenes; Transgenic Mice; Trichrome stain; Type II Epithelial Receptor Cell; Up-Regulation; Vascular Cell Adhesion Molecule-1; Viral; Virus Diseases; Wild Type Mouse; airway hyperresponsiveness; allergic airway inflammation; cell type; chemokine; cyclooxygenase 1; cysteinyl-leukotriene; cytokine; environmental agent; eosinophil; immune function; in vivo; indexing; influenzavirus; methacholine; mortality; overexpression; promoter; respiratory; response; vanadium pentoxide

We are investigating the role of cyclooxygenases in basal lung function and in the pulmonary response to environmental agents. At baseline, lung prostaglandin E2 levels are lower in COX-1 null mice compared to either wild type or COX-2 null mice, but there are no significant differences in basal lung function or in lung histopathology between the genotypes. Following allergen (ovalbumin) sensitizationexposure, lung inflammatory indices are significantly greater in COX-1 null and COX-2 null mice compared to wild type mice. Airways of allergic COX-1 null mice have increased numbers of eosinophils and increased numbers of CD3CD4 lymphocytes (Th cells). Alveolar macrophages from allergic COX-1 null airways show biochemical and morphologic evidence of activation. Bronchoalveolar lavage fluid (BALF) from allergic COX-1 null mice contains significantly higher levels of the Th2 cytokines IL-4, IL-5 and IL-13, increased levels of LTB4 and the cysteinyl leukotrienes, and increased levels of the chemokines TARC and eotaxin. These changes in the COX-1 null mice are associated with increased BALF IgE levels and increased MUC5AC productionmucin secretion. Moreover, expression of the adhesion molecules VCAM-1 and ICAM-1 are increased in the lungs of both allergic COX-1 and allergic COX-2 null mice. Allergic COX-1 null mice have reduced lung compliance, increased allergen-induced bronchoconstriction and display hyperresponsiveness to inhaled methacholine. We have also examined the effects of disruption of COX genes on the pulmonary responses to other environmentally relevant agents including inhaled endotoxin (bacterial lipopolysaccharide, LPS), vanadium pentoxide, and influenza virus. Following LPS exposure, all mice exhibit increased bronchoconstriction and methacholine hyperresponsiveness; however, these changes are much more pronounced in both the COX-1 null and COX-2 null mice relative to wild type controls. Interestingly, there are no significant differences in BALF cells or lung histopathology between the genotypes following LPS exposure. Thus, the balance of COX-1 and COX-2 is important in regulating the physiologic but not the inflammatory responses to inhaled LPS. Following vanadium pentoxide (V2O5) exposure, COX-2 null mice, but not COX-1 null mice, have increased acute lung inflammation and develop more lung fibrosis (increased lung hydroxyproline and enhanced trichrome staining). We have also utilized a pulmonary influenza infectivity model to evaluate host resistance and to determine if there are defects in innate or adaptive immune responses to viral infection in COX-1 null and COX-2 null mice. Infection induced less severe illness in COX-2 null mice in comparison to wild type and COX-1 null mice as evidenced by body weight and body temperature changes. Mortality was significantly reduced in COX-2 null mice. COX-1 null mice had enhanced inflammation and earlier appearance of pro-inflammatory cytokines in the BAL fluid, whereas the inflammatory and cytokine responses were blunted in COX-2 null mice. However, lung viral titres were markedly elevated in COX-2 null mice relative to wild type and COX-1 null mice. Levels of prostaglandin E2 were reduced in COX-1 null airways whereas cysteinyl leukotrienes were elevated in COX-2 null airways following infection. Thus, deficiency of COX-1 and COX-2 leads to contrasting effects in the host response to influenza infection, and these differences are associated with altered production of prostaglandins and leukotrienes following infection. The response of COX-deficient mice varies depending on the environmental stimulus. More recently, we developed transgenic mice with lung-specific overexpression of human COX-1 (murine CC10 promoter driven). Whereas no differences in basal respiratory or lung mechanical parameters were observed, COX-1 transgenic mice had increased bronchoalveolar lavage fluid prostaglandin E2 content compared to wild type littermates and exhibited decreased airway responsiveness to inhaled methacholine. In an ovalbumin-induced allergic airway inflammation model, comparable upregulation of COX-2 protein was observed in the lungs of allergic wild-type and COX 1 transgenic mice. Furthermore, no genotype differences were observed in allergic mice in total cell number, eosinophil content and inflammatory cytokine content of bronchoalveolar lavage fluid, or in airway responsiveness to inhaled methacholine. To eliminate the presumed confounding effects of COX-2 upregulation, COX-1 transgenic mice were bred into a COX-2 null background. In these mice, presence of the COX-1 transgene did not alter allergen-induced inflammation but significantly attenuated allergen-induced airway hyperresponsiveness, coincident with reduced airway leukotriene levels. Collectively, these data indicate that COX-1 overexpression attenuates airway responsiveness under basal conditions but does not influence allergic airway inflammation. We are currently developing transgenic mice with overexpression of COX-2 in type II cells (SPA promoter driven) to examine the role of COX-2 in the distal airway. We are also developing mice with selective knockout of COX-2 in the lung (Clara cells, type II alveolar cells) to determine if systemic or local biosynthesis of prostaglandins is important in regulating the lung response to environmental agents. Finally, we are studying the role of COX-1 and COX-2 in differentiation of lung T-cells in vitro and in vivo.

我们正在研究环氧合酶在基础肺功能和肺部对环境因素的反应中的作用。在基线时，与野生型或 COX-2 缺失小鼠相比，COX-1 缺失小鼠的肺前列腺素 E2 水平较低，但基因型之间的基础肺功能或肺组织病理学没有显着差异。过敏原（卵清蛋白）致敏暴露后，COX-1 缺失和 COX-2 缺失小鼠的肺部炎症指数显着高于野生型小鼠。过敏性 COX-1 缺失小鼠的气道中嗜酸性粒细胞数量增加，CD3CD4 淋巴细胞（Th 细胞）数量增加。来自过敏性 COX-1 无效气道的肺泡巨噬细胞显示出激活的生化和形态学证据。来自过敏性 COX-1 缺失小鼠的支气管肺泡灌洗液 (BALF) 含有显着较高水平的 Th2 细胞因子 IL-4、IL-5 和 IL-13，较高水平的 LTB4 和半胱氨酰白三烯，以及较高水平的趋化因子 TARC 和嗜酸细胞趋化因子。 COX-1 缺失小鼠中的这些变化与 BALF IgE 水平增加和 MUC5AC 产生粘蛋白分泌增加相关。此外，过敏性COX-1和过敏性COX-2无效小鼠的肺部粘附分子VCAM-1和ICAM-1的表达均增加。过敏性 COX-1 缺失小鼠的肺顺应性降低，过敏原诱导的支气管收缩增加，并对吸入乙酰甲胆碱表现出高反应性。我们还研究了 COX 基因破坏对其他环境相关物质（包括吸入内毒素（细菌脂多糖，LPS）、五氧化二钒和流感病毒）肺部反应的影响。 LPS暴露后，所有小鼠都表现出支气管收缩和乙酰甲胆碱高反应性增加；然而，相对于野生型对照，这些变化在 COX-1 缺失和 COX-2 缺失小鼠中更为明显。有趣的是，LPS 暴露后各基因型之间的 BALF 细胞或肺组织病理学没有显着差异。因此，COX-1 和 COX-2 的平衡对于调节吸入 LPS 的生理反应很重要，但对于调节炎症反应并不重要。暴露于五氧化二钒 (V2O5) 后，COX-2 缺失小鼠（而非 COX-1 缺失小鼠）急性肺部炎症增加并出现更多肺纤维化（肺羟脯氨酸增加和三色染色增强）。我们还利用肺部流感感染模型来评估宿主抵抗力，并确定 COX-1 缺失和 COX-2 缺失小鼠对病毒感染的先天或适应性免疫反应是否存在缺陷。体重和体温变化证明，与野生型和 COX-1 缺失小鼠相比，COX-2 缺失小鼠感染引起的疾病较轻。 COX-2 缺失小鼠的死亡率显着降低。 COX-1缺失小鼠的炎症增强，BAL液中促炎细胞因子的出现更早，而COX-2缺失小鼠的炎症和细胞因子反应则减弱。 然而，相对于野生型和 COX-1 缺失小鼠，COX-2 缺失小鼠的肺部病毒滴度显着升高。 感染后，COX-1 无效气道中的前列腺素 E2 水平降低，而 COX-2 无效气道中的半胱氨酰白三烯水平升高。 因此，COX-1和COX-2的缺乏会导致宿主对流感感染的反应产生不同的影响，并且这些差异与感染后前列腺素和白三烯的产生改变有关。 COX 缺陷小鼠的反应因环境刺激而异。 最近，我们开发了肺特异性过度表达人 COX-1（鼠 CC10 启动子驱动）的转基因小鼠。虽然基础呼吸或肺机械参数没有观察到差异，但与野生型同窝小鼠相比，COX-1 转基因小鼠的支气管肺泡灌洗液前列腺素 E2 含量增加，并且表现出对吸入乙酰甲胆碱的气道反应性降低。 在卵清蛋白诱导的过敏性气道炎症模型中，在过敏性野生型和 COX 1 转基因小鼠的肺部观察到 COX-2 蛋白的上调。 此外，在过敏小鼠中，在支气管肺泡灌洗液的细胞总数、嗜酸性粒细胞含量和炎性细胞因子含量或对吸入乙酰甲胆碱的气道反应性方面没有观察到基因型差异。 为了消除 COX-2 上调的假定混杂效应，将 COX-1 转基因小鼠培育到 COX-2 无效背景中。 在这些小鼠中，COX-1转基因的存在并没有改变过敏原诱导的炎症，但显着减弱了过敏原诱导的气道高反应性，这与气道白三烯水平的降低相一致。 总的来说，这些数据表明 COX-1 过度表达会减弱基础条件下的气道反应性，但不会影响过敏性气道炎症。我们目前正在开发在 II 型细胞（SPA 启动子驱动）中过度表达 COX-2 的转基因小鼠，以检查 COX-2 在远端气道中的作用。 我们还在开发选择性敲除肺部 COX-2 的小鼠（Clara 细胞，II 型肺泡细胞），以确定前列腺素的全身或局部生物合成对于调节肺部对环境因素的反应是否重要。最后，我们正在研究 COX-1 和 COX-2 在体外和体内肺 T 细胞分化中的作用。