Role of the Respiratory Syncytial Virus Fusion Protein in Airway Mucus Induction

描述（由申请人提供）：我们的长期目标是阐明呼吸道合胞病毒（RSV）引起肺功能障碍的机制。在美国，RSV每年使> 10万婴儿住院。肺粘液是RSV疾病的标志。粘液与上皮细胞碎片混合阻塞气道。RSV感染诱导气道粘液的机制尚不清楚。本提案的目的是确定粘液诱导RSV毒株的病毒融合（F）蛋白在发病机制中的作用，并确定中性粒细胞和嗜碱性粒细胞在RSV诱导的粘液中的作用。 与实验室RSV毒株相反，RSV毒株品系19和2-20在小鼠中诱导粘液和细支气管炎。使用RSV反向遗传学系统，我们表明19号系的F蛋白在粘液诱导中起作用。我们确定了5个氨基酸的线19 F蛋白参与粘液诱导的候选人。我们将使用F突变体病毒来定义对体内粘液表达重要的19系F残基。RSV 2- 20在小鼠中具有毒性。我们假设RSV 2-20 F有助于气道粘液和毒力。我们将使用2 - 20 F嵌合RSV来确定2-20 F在发病机制中的作用。细胞系19和2-20在体内引起比实验室RSV株更大的上皮细胞病变效应（CPE）和脱落。我们将使用F突变体来确定RSV株系19 F和2-20 F在体内和体外对上皮细胞CPE、合胞体和脱落的影响。我们将使用融合试验在体外确定RSV株系19、2-20和实验室菌株F蛋白在上皮细胞中的融合性。我们假设19 F和2-20 F系在体内和体外诱导更大的合胞体和融合。 该研究还将确定嗜中性粒细胞和嗜碱性粒细胞在RSV细支气管炎中的作用。与上皮细胞损伤同时，RSV株系19和2-20在小鼠肺中诱导嗜肺病毒。我们假设中性粒细胞在19和2-20号线发病机制中有助于病毒清除和粘液表达。细胞系19诱导的粘液依赖于IL-13，一种TH 2细胞因子。IL 4是主要的TH 2驱动细胞因子。我们发现嗜碱性粒细胞是RSV感染中表达IL 4的主要细胞类型。我们假设嗜碱性粒细胞有助于RSV株19和2-20诱导的IL-13和粘液表达。这些RSV毛细支气管炎模型将促进我们对RSV毒株依赖性发病机制的理解，并可能导致急需的治疗。

DESCRIPTION (provided by applicant): Our long term goal is to elucidate mechanisms by which respiratory syncytial virus (RSV) causes lung dysfunction. In the USA, RSV hospitalizes >100K infants/year. Pulmonary mucus is a hallmark of RSV disease. Mucus mixed with epithelial cell debris blocks the airways. Mechanisms by which RSV infection induces airway mucus are not known. Goals of this proposal are to define the role of the viral fusion (F) proteins of mucus-inducing RSV strains in pathogenesis and to define the roles of neutrophils and basophils in RSV-induced mucus. In contrast to laboratory RSV strains, RSV strains line 19 and 2-20 induce mucus and bronchiolitis in mice. Using a RSV reverse genetics system, we showed that the F protein of line 19 plays a role in mucus induction. We identified five amino acids in the line 19 F protein as candidates for involvement in mucus induction. We will use F mutant viruses to define line 19 F residues that are important for mucus expression in vivo. RSV 2- 20 is virulent in mice. We hypothesize that RSV 2-20 F contributes to airway mucus and virulence. We will define the role of 2-20 F in pathogenesis using 2-20F-chimeric RSV. Line 19 and 2-20 cause greater epithelial cell cytopathic effect (CPE) and desquamation in vivo than lab RSV strains. We will use F mutants to define effects of RSV line 19 F and 2-20 F on epithelial cell CPE, syncytia, and desquamation in vivo and in vitro. We will define the fusogenicity of RSV line 19, 2-20, and lab strain F proteins in epithelial cells in vitro using a fusion assay. We hypothesize that line 19 F and 2-20 F induce greater syncytia and fusion in vivo and in vitro. The study will also determine the roles of neutrophils and basophils in RSV bronchiolitis. Concurrently wih epithelial cell damage, RSV line 19 and 2-20 induce neutrophilia in the lungs of mice. We hypothesize that neutrophils contribute to viral clearance and mucus expression in line 19 and 2-20 pathogenesis. Line 19- induced mucus is dependent on IL-13, a TH2 cytokine. IL 4 is the primary TH2-driving cytokine. We showed that basophils are the major IL 4-expressing cell type in RSV-infection. We hypothesize that basophils contribute to RSV line 19- and 2-20-induced IL-13 and mucus expression. These RSV bronchiolitis models will advance our understanding of RSV strain-dependent pathogenesis and may lead to much-needed therapies.