Association of bacteria and viruses with wheezy episodes in young children: prospective birth cohort study

Objective To study the association between wheezy symptoms in young children and the presence of bacteria in the airways

A Comprehensive Evaluation of Nasal and Bronchial Cytokines and Chemokines Following Experimental Rhinovirus Infection in Allergic Asthma: Increased Interferons (IFN-γ and IFN-λ) and Type 2 Inflammation (IL-5 and IL-13).

BACKGROUND: Rhinovirus infection is a major cause of asthma exacerbations. OBJECTIVES: We studied nasal and bronchial mucosal inflammatory responses during experimental rhinovirus-induced asthma exacerbations. METHODS: We used nasosorption on days 0, 2-5 and 7 and bronchosorption at baseline and day 4 to sample mucosal lining fluid to investigate airway mucosal responses to rhinovirus infection in patients with allergic asthma (n=28) and healthy non-atopic controls (n=11), by using a synthetic absorptive matrix and measuring levels of 34 cytokines and chemokines using a sensitive multiplex assay. RESULTS: Following rhinovirus infection asthmatics developed more upper and lower respiratory symptoms and lower peak expiratory flows compared to controls (all P<0.05). Asthmatics also developed higher nasal lining fluid levels of an anti-viral pathway (including IFN-γ, IFN-λ/IL-29, CXCL11/ITAC, CXCL10/IP10 and IL-15) and a type 2 inflammatory pathway (IL-4, IL-5, IL-13, CCL17/TARC, CCL11/eotaxin, CCL26/eotaxin-3) (area under curve day 0-7, all P<0.05). Nasal IL-5 and IL-13 were higher in asthmatics at day 0 (P<0.01) and levels increased by days 3 and 4 (P<0.01). A hierarchical correlation matrix of 24 nasal lining fluid cytokine and chemokine levels over 7days demonstrated expression of distinct interferon-related and type 2 pathways in asthmatics. In asthmatics IFN-γ, CXCL10/IP10, CXCL11/ITAC, IL-15 and IL-5 increased in bronchial lining fluid following viral infection (all P<0.05). CONCLUSIONS: Precision sampling of mucosal lining fluid identifies robust interferon and type 2 responses in the upper and lower airways of asthmatics during an asthma exacerbation. Nasosorption and bronchosorption have potential to define asthma endotypes in stable disease and at exacerbation

Bronchial mucosal inflammation and illness severity in response to experimental rhinovirus infection in COPD

Background: Respiratory viral infection causes chronic obstructive pulmonary disease (COPD) exacerbations. We previously reported increased bronchial mucosa eosinophil and neutrophil inflammation in patients with COPD experiencing naturally occurring exacerbations. But it is unclear whether virus per se induces bronchial mucosal inflammation, nor whether this relates to exacerbation severity. Objectives: We sought to determine the extent and nature of bronchial mucosal inflammation following experimental rhinovirus (RV)-16–induced COPD exacerbations and its relationship to disease severity. Methods: Bronchial mucosal inflammatory cell phenotypes were determined at preinfection baseline and following experimental RV infection in 17 Global Initiative for Chronic Obstructive Lung Disease stage II subjects with COPD and as controls 20 smokers and 11 nonsmokers with normal lung function. No subject had a history of asthma/allergic rhinitis: all had negative results for aeroallergen skin prick tests. Results: RV infection increased the numbers of bronchial mucosal eosinophils and neutrophils only in COPD and CD8+ T lymphocytes in patients with COPD and nonsmokers. Monocytes/macrophages, CD4+ T lymphocytes, and CD20+ B lymphocytes were increased in all subjects. At baseline, compared with nonsmokers, subjects with COPD and smokers had increased numbers of bronchial mucosal monocytes/macrophages and CD8+ T lymphocytes but fewer numbers of CD4+ T lymphocytes and CD20+ B lymphocytes. The virus-induced inflammatory cells in patients with COPD were positively associated with virus load, illness severity, and reductions in lung function. Conclusions: Experimental RV infection induces bronchial mucosal eosinophilia and neutrophilia only in patients with COPD and monocytes/macrophages and lymphocytes in both patients with COPD and control subjects. The virus-induced inflammatory cell phenotypes observed in COPD positively related to virus load and illness severity. Antiviral/anti-inflammatory therapies could attenuate bronchial inflammation and ameliorate virus-induced COPD exacerbations

Asthma-like airways inflammation is CD4+ T cell dependent.

<p>Tbet-/- mice were infected intranasally with RV1B or sham infected with PBS. In addition, mice were systemically depleted of CD4 expressing cells (anti-CD4), or treated with isotype control antibody (isotype) 3hrs prior to infection. Tissues were harvested at 7 days post-challenge. (A) Lung flow cytometry staining for CD3+CD4+ T cells. (B-D) Levels of cytokines IL-4 (B), IL-13 (C) and IL-17a (D) in lung tissue measured by Taqman qPCR. (E) Total BAL cell counts and (F) eosinophil counts in BAL measured by cytospin assay. (G) MUC5AC levels in BAL measured by ELISA. (H) Representative PAS staining for mucus in lung tissue sections. Scale bars 50μm. n = 12–15 mice/group (mice for which CD4+ cell depletion was not successful were excluded from all analyses (n = 5 of 30)). ***p<0.001, **p<0.01, *p<0.05, n.s. not significant.</p

RV-specific antibody responses in Tbet deficient and wild type mice.

<p>Wild type and Tbet-/- mice were infected intranasally with RV1B or sham infected with PBS. Blood was collected 14 days after infection. (A&B) RV1B-binding IgG2c (A) and IgG1 (B) in sera was measured by ELISA. (C) Neutralisation of RV1B infection of Ohio HeLa cells by pooled sera assessed by crystal violet cell viability staining. ATCC ctl: control reference guinea pig anti-sera. Top dashed line in C, uninfected cells control. Bottom dashed line in C, RV infected cells control. Data represent results from 5–6 pooled sera per treatment group in a single experiment, representative of 3 independent experiments.</p

A short-term mouse model that reproduces the immunopathological features of rhinovirus-induced exacerbation of COPD

Viral exacerbations of chronic obstructive pulmonary disease (COPD), commonly caused by rhinovirus (RV) infections, are poorly controlled by current therapies. This is due to a lack of understanding of the underlying immunopathological mechanisms. Human studies have identified a number of key immune responses that are associated with RV-induced exacerbations including neutrophilic inflammation, expression of inflammatory cytokines and deficiencies in innate anti-viral interferon. Animal models of COPD exacerbation are required to determine the contribution of these responses to disease pathogenesis. We aimed to develop a short-term mouse model that reproduced the hallmark features of RV-induced exacerbation of COPD. Evaluation of complex protocols involving multiple dose elastase and lipopolysaccharide (LPS) administration combined with RV1B infection showed suppression rather than enhancement of inflammatory parameters compared with control mice infected with RV1B alone. Therefore, these approaches did not accurately model the enhanced inflammation associated with RV infection in patients with COPD compared with healthy subjects. In contrast, a single elastase treatment followed by RV infection led to heightened airway neutrophilic and lymphocytic inflammation, increased expression of tumour necrosis factor (TNF)-α, C-X-C motif chemokine 10 (CXCL10)/IP-10 (interferon γ-induced protein 10) and CCL5 [chemokine (C-C motif) ligand 5]/RANTES (regulated on activation, normal T-cell expressed and secreted), mucus hypersecretion and preliminary evidence for increased airway hyper-responsiveness compared with mice treated with elastase or RV infection alone. In summary, we have developed a new mouse model of RV-induced COPD exacerbation that mimics many of the inflammatory features of human disease. This model, in conjunction with human models of disease, will provide an essential tool for studying disease mechanisms and allow testing of novel therapies with potential to be translated into clinical practice

Helper T cell responses in Tbet deficient and wild type mice.

<p>Wild type and Tbet-/- mice were infected intranasally with RV1B or sham infected with PBS. (A-D) Intranuclear flow cytometry staining for transcription factors Tbet (A), GATA-3 (B), RORγt (C) and FOXP3 (D) in CD3+CD4+ lung T cells, 2 and 7 days post-infection. (E-H) Intracellular flow cytometry staining for cytokines IFN-γ (E), IL-13 (F), IL-17A (G) and IL-10 (H) in lung CD3+CD4+ cells stimulated with PMA and ionomycin, on day 7 post-infection. (I-L) RNA was extracted from lung tissue harvested on day 7 post-infection and expression of IFN-γ (I), IL-13 (J), IL-17a (K) and IL-10 (L) mRNA was quantified by Taqman qPCR. n = 8–9 mice/group.***p<0.001, **p<0.01, *p<0.05, n.s. not significant.</p

Rhinovirus infection induces expression of airway remodelling factors in vitro and in vivo

BACKGROUND AND OBJECTIVE: A hallmark of asthma is airway remodelling, which includes increased deposition of extracellular matrix (ECM) protein. Viral infections may promote the development of asthma and are the most common causes of asthma exacerbations. We evaluated whether rhinovirus (RV) infection induces airway remodelling, as assessed by ECM deposition. METHODS: Primary human bronchial epithelial cells and lung parenchymal fibroblasts were infected with RV-2 or RV-16, or treated with RV-16 RNA, imiquimod (Toll-like receptor (TLR) 7/8 agonist) or polyinosinic : polycytidylic acid (poly I : C) (activator of TLR 3, retinoic-acid-inducible protein I and melanoma-differentiated-associated gene 5). Changes in ECM proteins and their transcription were measured by ELISA and quantitative real-time PCR. In addition, gene expression for ECM proteins was assessed in a mouse model of RV infection. RESULTS: RV infection increased deposition of the ECM protein, perlecan, by human bronchial epithelial cells, and collagen V and matrix-bound vascular endothelial growth factor were increased in both human bronchial epithelial cell and fibroblast cultures. Purified RV-16 RNA, poly I : C and imiquimod induced similar increases in ECM deposition to those observed with RV-infected fibroblasts. However, only poly I : C induced ECM deposition by bronchial epithelial cells, suggesting that RV-induced ECM deposition is mediated through TLR. Furthermore, gene expression for fibronectin and collagen I was increased in lung homogenates of mice infected with RV-1b. CONCLUSIONS: RV infection and TLR ligands promote ECM deposition in isolated cell systems and RV induces ECM gene expression in vivo, thus demonstrating that RV has the potential to contribute to remodelling of the airways through induction of ECM deposition

Experimental Rhinovirus Infection as a Human Model of Chronic Obstructive Pulmonary Disease Exacerbation

Rationale: Respiratory virus infections are associated with chronic obstructive pulmonary disease (COPD) exacerbations, but a causative relationship has not been proven. Studies of naturally occurring exacerbations are difficult and the mechanisms linking virus infection to exacerbations are poorly understood. We hypothesized that experimental rhinovirus infection in subjects with COPD would reproduce the features of naturally occurring COPD exacerbations and is a valid model of COPD exacerbations