Models of rhinovirus-C respiratory infection and asthma

项目总结 越来越多的证据表明，感染一种新发现的鼻病毒RV-C也是如此- 与严重的呼吸道感染和哮喘加重有关，通常需要住院治疗。另外-- 最近的数据表明，早期RV-C感染在哮喘发展中可能起到了作用。 尽管越来越多的人认识到RV-C是哮喘的原因，但几乎对此一无所知 RV-C感染的发病机制。为了做到这一点，我们用RV-C15感染小鼠(RV-C15 infec- 高表达人RV-C受体变异体钙粘素相关家族的特异性克隆和HeLa-E8细胞 成员3，由威斯康星大学James Gern获得)。我们的初步研究表明，RV-C15- 感染小鼠表现出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- 活性、CD25+、CD127+2型固有淋巴样细胞(ILC2s)与RV-A1B感染小鼠比较。此外，圆周率- 对自然RV-C感染儿童的大量研究表明，2型细胞因子的产生增加。 在这个应用中，我们将检验一个普遍的假设，即在RV-C感染后，呼吸道固有的细胞因子ex. 压力驱动ILC2的扩张和发展为嗜酸性炎症和粘液化生。为了测试 在这一假设下，我们提出了以下具体目标： 特定目的1.确定上皮源性先天细胞因子在RV-C15诱导中的作用 嗜酸性呼吸道炎症和高反应性(AHR)。我们假设：1)与 成年小鼠感染RV-A、RV-C后，肺组织固有细胞因子(IL-25、IL-33、TSLP)表达增加； 2)IL-25由双皮质激素样激酶(DCLK)-1阳性的气道束细胞产生； 嗜酸性炎症；4)CDHR3的RV-C参与激活不同的信号通路 导致先天细胞因子的表达。 特定目的2.确定肺ILC2和巨噬细胞在RV-C诱导的气道中的作用 炎症和AHR。我们推测：1)成熟小鼠的RV-C感染诱导先天细胞因子去... 2)ILC2s促进嗜酸性炎症、巨噬细胞极化和AHR； 3)室内粉尘(HDM)和RV-C对嗜酸性粒细胞炎症和AHR有相加作用；4)ILC2 传递皮质类固醇耐药性；以及5)感染RV-C的人的鼻腔抽吸物显示- 与RV-A感染者的样本相比，2型细胞因子和ILC2的表达增加。 特定目的3.确定早期RV-C感染对已建立的哮喘表型的影响 打字。我们发现，RV-A1B感染6日龄小鼠，而不是成熟小鼠，可诱导持久的小鼠感染。 慢性上皮化生和AHR，依赖于产生IL-13的ILC2。我们假设：1)RV-C信息- 出生6天的小鼠比RV-A诱导更大和更持久的粘液化生；2)早期生命的RV-C 感染增加了产生IL-25的呼吸道绒毛细胞的数量。

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.