Modulation of neonatal alveolar macrophage by cftr mutation

cftr 突变对新生儿肺泡巨噬细胞的调节

• 批准号: 8822087
• 负责人: Lou Ann S Brown
• 金额: $ 23.4万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-22 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8822087
• 关键词: AGTR2 gene; Address; Alveolar Macrophages; Antioxidants; Attenuated; Bacteria; Breathing; Bronchoalveolar Lavage Fluid; Caring; Cell Line; Characteristics; Chronic; Cystic Fibrosis; Data; Depressed mood; Development; Diagnosis; Early identification; Epithelial Cells; Functional disorder; Glutathione; Health; Human; Immune response; Immunosuppression; Infant; Infection; Infiltration; Inflammation; Inflammatory; Inflammatory Response; Investigation; Knock-out; Laboratories; Lower Respiratory Tract Infection; Lung; Lung diseases; Macrophage Activation; Mediating; Microbe; Modeling; Morbidity - disease rate; Mus; Mutate; Mutation; Neonatal; Neutrophil Infiltration; Newborn Infant; Outcome; Oxidants; Oxidative Stress; Phagocytosis; Play; Pneumonia; Pulmonary Pathology; Respiratory Syncytial Virus Infections; Respiratory syncytial virus; Risk; Role; Signal Transduction; Symptoms; Therapeutic; Therapeutic Intervention; Up-Regulation; Viral; Virus; Virus Diseases; Work; arginase; children with cystic fibrosis; endoplasmic reticulum stress; extracellular; immune function; improved; mouse model; neonatal human; neonate; oxidant stress; pathogen; pathogen exposure; respiratory; response

DESCRIPTION (provided by applicant): In this proposal, we will address the often debated role of the neonatal alveolar macrophage (AM) as an important contributor to impaired immune responses in the developing cystic fibrosis (CF) lung. AM are important in the initial defenses against bacteria and viruses, including respiratory syncytial virus (RSV), an important viral pathogen which causes significant morbidity in the CF lung. As one of few laboratories focused on the effects of oxidative stress on the function of AM in the newborn lung, we have begun to evaluate the effects of cftr mutation on neonatal AM using a mouse cftr knockout gut corrected model (cftr KO). While cftr KO mice do not spontaneously develop characteristic pulmonary pathologies of human CF lung per se, they serve as excellent models to evaluate the intrinsic cftr alterations in neonatal AM, in the absence of pulmonary infection. In cftr KO neonatal mice, oxidative stress was increased in the bronchoalveolar lavage fluid (BALF), even in the absence of infection. Cftr KO AM had dramatic increases in TGFβ1 and the M2 marker arginase 1 (Arg1), and depressed phagocytosis. This suggests that the cftr mutation shifts the neonatal AM's activation state away from pathogen clearance (classically activated, M1) towards immunosuppression (alternative activation,M2). Interestingly, control neonatal AM treated with the BALF obtained from young children with CF also mimic the effects of the cftr KO AM with increased oxidant stress, TGFβ1 and M2 polarization. A central role for CF-related decreases in the antioxidant glutathione (GSH) was demonstrated by the ability of extracellular GSH to attenuate the increased TGFβ1 and Arg1 and improve phagocytosis in cftr KO AM and CFBALF-exposed AM. This suggested that oxidant-induced up- regulation of TGFβ1 modulated the neonatal cftr KO AM's inflammatory state; but increasing extracellular GSH blunted CF-induced TGFβ1 signaling and its downstream effects on AM polarity and function. Our central hypothesis is that decreased GSH pools due to cftr mutation increase AM oxidant stress and impair phagocytosis by increasing TGFβ1 thereby shifting neonatal AM towards M2 activation and diminishing the neonatal lung's defenses against RSV. In Aim 1, we will determine the effects of cftr mutations on baseline and RSV-stimulated state of manipulated cells lines, and primary neonatal AM isolated from cftr KO, ΔF508 and G551D mutated neonatal mice. In Aim 2, we will examine the mechanisms by which maintaining GSH pools modulates the immune functions of neonatal CF AM. Alterations in AM activation state and subsequent changes in inflammatory signaling may compromise RSV clearance. This impaired response by AM may then be the signal that promotes sustained neutrophil recruitment into the airspace. With early identification of the CF newborn, the development of potential early therapeutic interventions could modify pulmonary inflammation and infection before respiratory symptoms occur. This proposal will begin to address whether the neonatal AM is an appropriate target for the development of such potential therapeutic strategies.

描述(由申请人提供)：在这项提案中，我们将讨论新生儿肺泡巨噬细胞(AM)作为发展中的囊性纤维化(CF)肺免疫反应受损的重要贡献者所扮演的经常有争议的角色。AM在抵抗细菌和病毒包括呼吸道合胞病毒(RSV)的初始防御中起重要作用，RSV是一种重要的病毒病原体，会导致CF肺的显著发病率。作为少数几个关注氧化应激对新生儿肺AM功能影响的实验室之一，我们已经开始使用小鼠CFTR基因敲除肠道校正模型(CFTRKO)来评估CFTR突变对新生儿AM的影响。虽然CFTRKO小鼠本身并不会自发地发展成人类CFT肺的特征肺部病变，但在没有肺部感染的情况下，它们可以作为评估新生儿AM固有的CFTR性改变的极佳模型。在CFTRKO新生小鼠中，即使在没有感染的情况下，支气管肺泡灌洗液(BALF)中的氧化应激也会增加。CFTRKO AM中转化生长因子β1和M2标记的精氨酸酶1(Arg1)显著升高，吞噬功能降低。这表明CFTR突变将新生儿AM的激活状态从病原体清除(经典激活，M1)转移到免疫抑制(替代激活，M2)。有趣的是，对照组新生儿AM也模拟CFTRKO AM的氧化应激增加、转化生长因子β1和M2极化的效应。细胞外谷胱甘肽抑制CFTRKO AM和CFBALF暴露的AM中转化生长因子β1和Arg1的增加并改善吞噬功能是抗氧化剂谷胱甘肽减少的主要原因。提示氧化剂诱导的转化生长因子β1的上调调节了新生CFTRKO AM的炎症状态，但细胞外增加的谷胱甘肽抑制了CF诱导的转化生长因子β1信号转导及其对AM极性和功能的下游影响。我们的中心假设是，cftr突变导致的谷胱甘肽池减少增加了AM的氧化应激，并通过增加转化生长因子β1从而损害吞噬功能，从而使新生儿AM转向M2激活，并降低了新生儿肺对呼吸道合胞病毒的防御能力。在目标1中，我们将确定cftr突变对操纵细胞系和从cftr KO、ΔF508和G551D突变的新生小鼠分离的原代新生AM的基线和呼吸道合胞病毒刺激状态的影响。在目标2中，我们将研究维持GSH池调节新生儿CFAM免疫功能的机制AM激活状态的改变和随后炎症信号的改变可能会影响RSV的清除。AM的这种受损反应可能是促进中性粒细胞持续重新进入空域的信号。随着对CF新生儿的早期识别，潜在的早期治疗干预措施的发展可以在呼吸道症状出现之前改善肺部炎症和感染。这项提案将开始讨论新生儿AM是否是开发这种潜在治疗策略的合适靶点。