Models of rhinovirus-C respiratory infection and asthma

项目摘要 越来越多的证据表明，感染一种新发现的鼻病毒，RV-C，是阿索， 伴有严重的呼吸道感染和哮喘恶化，通常需要住院治疗。此外， 因此，最近的数据表明早期RV-C感染在哮喘发展中可能起作用。 尽管越来越多的人认识到RV-C是哮喘疾病的一个原因，但实际上对RV-C一无所知。 RV-C感染的发病机制。为了实现这一点，我们用RV-C15感染小鼠（RV-C15感染小鼠）。 过表达人RV-C受体钙粘蛋白相关家族变异体的克隆和HeLa-E8细胞 成员3，获自威斯康星州大学的James热尔恩）。我们的初步研究表明，RV-C15- 感染的小鼠表现出2型细胞因子和粘蛋白基因表达增加，BAL嗜酸性粒细胞和谱系阴性， 表达，CD 25+，CD 127 + 2型先天性淋巴样细胞（ILC 2）与RV-A1 B感染小鼠相比。此外，PI- 对患有自然RV-C感染的儿童的大量研究表明2型细胞因子的产生增加。 在本申请中，我们将检验一般假设，即在RV-C感染后，气道先天性细胞因子表达增加， 表达驱动ILC 2扩增和嗜酸性炎症和粘液化生的发展。测试 基于这一假设，我们提出以下具体目标： 具体目标1。确定上皮来源的先天性细胞因子对RV-C15诱导的细胞凋亡的贡献。 嗜酸性粒细胞气道炎症和高反应性（AHR）。我们假设：1）与 成熟小鼠的RV-A、RV-C感染诱导先天性细胞因子（IL-25、IL-33、TSLP）的更大的肺表达; 2)IL-25由双皮质素样激酶（DCLK）-1阳性气道簇细胞产生; 3）先天性细胞因子被重新激活。 4）RV-C与CDHR 3的结合激活不同的信号通路 导致先天性细胞因子表达。 具体目标2。确定肺ILC 2和巨噬细胞对RV-C诱导的气道的贡献 炎症和AHR。我们假设：1）RV-C感染成熟小鼠诱导先天性精氨酸脱氨酶， ILC 2s促进嗜酸性粒细胞炎症、巨噬细胞极化和AHR; 3)屋尘螨（HDM）和RV-C对嗜酸性粒细胞炎症和AHR有相加作用; 4）ILC 2 传递皮质类固醇抗性;和5）来自感染RV-C的人类受试者的鼻吸出物显示- 与来自RV-A感染受试者的样品相比，2型细胞因子和ILC 2的表达增加。 具体目标3。确定生命早期RV-C感染对已确定的哮喘表型的影响， 类型.我们已经发现，RV-A1 B感染6日龄小鼠，而不是成熟小鼠，诱导持久的μ- Cous化生和AHR，其依赖于产生IL-13的ILC 2。我们假设：1）RV-C感染- 与RV-A相比，6日龄小鼠的RV-C诱导更大和更持久的粘膜化生; 2）早期RV-C 感染增加了产生IL-25的气道簇细胞的数量。

PROJECT SUMMARY Accumulating evidence indicates that infections with a newly-discovered species of rhinovirus, RV-C, are asso- ciated with severe respiratory tract infections and asthma exacerbations often requiring hospitalization. In addi- tion, recent data suggest a possible role for early-life RV-C infections in asthma development. Despite increasing recognition of RV-C as a cause of asthmatic disease, virtually nothing is known about the pathogenesis of RV-C infections. To accomplish this, we infected mice with RV-C15 (the RV-C15 infec- tious clone and HeLa-E8 cells overexpressing a variant of the human RV-C receptor, cadherin related family member 3, were obtained from James Gern, University of Wisconsin). Our pilot studies show that RV-C15- infected mice show increased type 2 cytokine and mucin gene expression, BAL eosinophils and lineage-nega- tive, CD25+, CD127+ type 2 innate lymphoid cells (ILC2s) compared to RV-A1B-infected mice. In addition, pi- lot studies from children with natural RV-C infections show increased type 2 cytokine production. In this application, we will test the general hypothesis that, after RV-C infection, airway innate cytokine ex- pression drives ILC2 expansion and development of eosinophilic inflammation and mucous metaplasia. To test this hypothesis, we propose the following Specific Aims: Specific Aim 1. Determine the contribution of epithelial-derived innate cytokines to RV-C15-induced eosinophilic airway inflammation and hyperresponsiveness (AHR). We hypothesize that: 1) compared to RV-A, RV-C infection of mature mice induces greater lung expression of innate cytokines (IL-25, IL-33, TSLP); 2) IL-25 is produced by doublecortin-like kinase (DCLK)-1-positive airway tuft cells; 3) innate cytokines are re- quired for eosinophilic inflammation; 4) RV-C engagement of CDHR3 activates distinct signaling pathways leading to innate cytokine expression. Specific Aim 2. Determine the contribution of lung ILC2s and macrophages to RV-C-induced airway inflammation and AHR. We hypothesize that: 1) RV-C infection of mature mice induces innate cytokine-de- pendent expansion of ILC2s; 2) ILC2s promote eosinophilic inflammation, macrophage polarization and AHR; 3) house dust mite (HDM) and RV-C have additive effects on eosinophilic inflammation and AHR; 4) ILC2s convey corticosteroid resistance; and 5) nasal aspirates from human subjects infected with RV-C show in- creased expression of type 2 cytokines and ILC2s compared to samples from RV-A-infected subjects. Specific Aim 3. Determine the effects of early-life RV-C infection on the established asthma pheno- type. We have found that RV-A1B infection of six day-old mice, but not mature mice, induces long-lasting mu- cous metaplasia and AHR which is dependent on IL-13-producing ILC2s. We hypothesize that: 1) RV-C infec- tion of 6 day-old mice induces greater and more long-lasting mucous metaplasia than RV-A; 2) early-life RV-C infection increases the number of IL-25-producing airway tuft cells.