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Patients with frequent exacerbations represent a chronic obstructive pulmonary disease (COPD) subgroup requiring better treatment options. The aim of this study was to determine the innate immune mechanisms that underlie susceptibility to frequent exacerbations in COPD. We measured sputum expression of immune mediators and bacterial loads in samples from patients with COPD at stable state and during virus-associated exacerbations. In vitro immune responses to rhinovirus infection in differentiated primary bronchial epithelial cells (BECs) sampled from patients with COPD were additionally evaluated. Patients were stratified as frequent exacerbators (≥2 exacerbations in the preceding year) or infrequent exacerbators (

Bacterial infection commonly complicates inflammatory airway diseases such as chronic obstructive pulmonary disease (COPD). The mechanisms of increased infection susceptibility and how use of the commonly prescribed therapy inhaled corticosteroids (ICS) accentuates pneumonia risk in COPD are poorly understood. Here, using analysis of samples from patients with COPD, we show that ICS use is associated with lung microbiota disruption leading to proliferation of streptococcal genera, an effect that could be recapitulated in ICS-treated mice. To study mechanisms underlying this effect, we used cellular and mouse models of streptococcal expansion with Streptococcus pneumoniae, an important pathogen in COPD, to demonstrate that ICS impairs pulmonary clearance of bacteria through suppression of the antimicrobial peptide cathelicidin. ICS impairment of pulmonary immunity was dependent on suppression of cathelicidin because ICS had no effect on bacterial loads in mice lacking cathelicidin (Camp−/−) and exogenous cathelicidin prevented ICS-mediated expansion of streptococci within the microbiota and improved bacterial clearance. Suppression of pulmonary immunity by ICS was mediated by augmentation of the protease cathepsin D. Collectively, these data suggest a central role for cathepsin D/cathelicidin in the suppression of antibacterial host defense by ICS in COPD. Therapeutic restoration of cathelicidin to boost antibacterial immunity and beneficially modulate the lung microbiota might be an effective strategy in COPD.

The respiratory tract surface is protected from inhaled pathogens by a secreted layer of mucus that is rich in mucin glycoproteins. Disrupted mucus production is a cardinal feature of chronic respiratory diseases but how this alteration affect interactions between mucins and pathogens is complex and poorly understood. Here, we identify a central and unexpected role for the major airway mucin MUC5AC in pathogenesis of virus-induced exacerbations of chronic obstructive pulmonary disease (COPD). Virus induction of MUC5AC is augmented in COPD compared to healthy subjects, is enhanced in frequent exacerbators and correlates with inflammation, symptom severity and secondary bacterial infection during exacerbation. MUC5AC is functionally related to inflammation as MUC5AC-deficient (Muc5ac-/-) mice had attenuated rhinovirus-induced airway inflammation whilst exogenous MUC5AC glycoprotein administration augmented virus-induced inflammatory responses and bacterial load. Mechanistically, MUC5AC-augmentation of rhinovirus-induced inflammation occurred through release of extracellular adenosine triphosphate (ATP). Therapeutic suppression of virus-induced MUC5AC release using an epidermal growth factor receptor (EGFR) inhibitor ameliorated exaggerated pro-inflammatory responses in a mouse COPD exacerbation model. Collectively, these studies demonstrate previously unrecognised pro-inflammatory effects of MUC5AC during infection and thus highlight a key unforeseen role in driving COPD exacerbation severity.

Background Respiratory virus infections are commonly associated with COPD exacerbations, but little is known about the mechanisms linking virus infection to exacerbations. Pathogenic mechanisms in stable COPD include oxidative and nitrosative stress and reduced activity of histone deacetylase-2 (HDAC2), but their roles in COPD exacerbations is unknown. We investigated oxidative and nitrosative stress (O&NS) and HDAC2 in COPD exacerbations using experimental rhinovirus infection. Methods Nine subjects with COPD (Global Initiative for Chronic Obstructive Lung Disease stage II), 10 smokers, and 11 nonsmokers were successfully infected with rhinovirus. Markers of O&NS-associated cellular damage, and inflammatory mediators and proteases were measured in sputum, and HDAC2 activity was measured in sputum and bronchoalveolar macrophages. In an in vitro model, monocyte-derived THP-1 cells were infected with rhinovirus and nitrosylation and activity of HDAC2 was measured. Results Rhinovirus infection induced significant increases in airways inflammation and markers of O&NS in subjects with COPD. O&NS markers correlated with virus load and inflammatory markers. Macrophage HDAC2 activity was reduced during exacerbation and correlated inversely with virus load, inflammatory markers, and nitrosative stress. Sputum macrophage HDAC2 activity pre-infection was inversely associated with sputum virus load and inflammatory markers during exacerbation. Rhinovirus infection of monocytes induced nitrosylation of HDAC2 and reduced HDAC2 activity; inhibition of O&NS inhibited rhinovirus-induced inflammatory cytokines. Conclusions O&NS, airways inflammation, and impaired HDAC2 may be important mechanisms of virus-induced COPD exacerbations. Therapies targeting these mechanisms offer potential new treatments for COPD exacerbations.

Rhinovirus infection is associated with the majority of asthma exacerbations. The role of fractalkine in anti-viral (type 1) and pathogenic (type 2) responses to rhinovirus infection in allergic asthma is unknown. To determine whether (1) fractalkine is produced in airway cells and in peripheral blood leucocytes, (2) rhinovirus infection increases production of fractalkine and (3) levels of fractalkine differ in asthmatic compared to non-asthmatic subjects. Fractalkine protein and mRNA levels were measured in bronchoalveolar lavage (BAL) cells and peripheral blood mononuclear cells (PBMCs) from non-asthmatic controls (n = 15) and mild allergic asthmatic (n = 15) subjects. Protein levels of fractalkine were also measured in macrophages polarised ex vivo to give M1 (type 1) and M2 (type 2) macrophages and in BAL fluid obtained from mild (n = 11) and moderate (n = 14) allergic asthmatic and non-asthmatic control (n = 10) subjects pre and post in vivo rhinovirus infection. BAL cells produced significantly greater levels of fractalkine than PBMCs. Rhinovirus infection increased production of fractalkine by BAL cells from non-asthmatic controls (P

Background: Fibre-optic bronchoscopy (FOB) is a research tool in asthma and COPD with a good safety record in stable patients. There is no data regarding adverse events for FOB in patients with asthma/COPD exacerbations. Aims: To evaluate the safety of FOB in subjects participating in experimental rhinovirus infection studies. Methods: We reviewed the records of these subjects from a single centre since 2000. Subjects had FOB at baseline and convalescence (stable) and during the virus-induced exacerbation (infection) with biopsies, brushings and BAL. Results: The clinical characteristics of the subjects are in Table 1 . There were no adverse events in the asthma studies and 3 in the COPD studies. Two subjects developed lower respiratory tract infections requiring antibiotics. In 1 subject (non-smoker) this occurred at the infection FOB and the subject was withdrawn. In the other subject (COPD) this occurred at the baseline FOB and he continued in the study and had 2 other uneventful FOB. One subject (smoker) had FOB terminated due to hypoxia. Conclusions: The incidence of adverse events in research FOB in asthma and COPD exacerbations is low.

Copyright © 2016 Russell et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Introduction. Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine associated with acute and chronic inflammatory disorders and corticosteroid insensitivity. Its expression in the airways of patients with chronic obstructive pulmonary disease (COPD), a relatively steroid insensitive inflammatory disease is unclear, however. Methods. Sputum, bronchoalveolar lavage (BAL) macrophages and serum were obtained from nonsmokers, smokers and COPD patients. To mimic oxidative stress-induced COPD, mice were exposed to ozone for six-weeks and treated with ISO-1, a MIF inhibitor, and/or dexamethasone before each exposure. BAL fluid and lung tissue were collected after the final exposure. Airway hyperresponsiveness (AHR) and lung function were measured using whole body plethysmography. HIF-1α binding to the Mif promoter was determined by Chromatin Immunoprecipitation assays. Results. MIF levels in sputum and BAL macrophages from COPD patients were higher than those from non-smokers, with healthy smokers having intermediate levels. MIF expression correlated with that of HIF-1α in all patients groups and in ozone-exposed mice. BAL cell counts, cytokine mRNA and protein expression in lungs and BAL, including MIF, were elevated in ozone-exposed mice and had increased AHR. Dexamethasone had no effect on these parameters in the mouse but ISO-1 attenuated cell recruitment, cytokine release and AHR. Conclusion MIF and HIF-1α levels are elevated in COPD BAL macrophages and inhibition of MIF function blocks corticosteroid-insensitive lung inflammation and AHR. Inhibition of MIF may provide a novel anti-inflammatory approach in COPD.

Acute cough is a major symptom of viral respiratory tract infection and causes excessive morbidity in human populations across the world. A wide variety of viruses play a role in the development of cough after acute infection and all of these manifest a similar clinical picture across different age groups. Despite the large disease burden surprisingly little is known about the mechanism of acute cough following viral infection. Both in vitro and in vivo experiments show that increased production of neuropeptides and leukotrienes mediate cough after viral infection, along with altered expression of neural receptors. Increased airway mucus production is also likely to play a significant role. This work is reviewed in this article. Following the recent development of a mouse model for rhinovirus infection and the establishment of experimental models of rhinovirus challenge in human subjects with both asthma and COPD the field is expanding to translate in vitro research into clinical studies and hopefully eventually into clinical practice. Developing a clearer understanding of the mechanisms underlying virus induced cough may lead to more specific and effective therapies.

Rhinoviruses are a major driver of the reduced lung function seen during exacerbations in a number of airway diseases including asthma and COPD. Viral infection is associated with increased inflammation in the lungs and increased oxidative stress in the airways. Monocyte and epithelial cells lines (THP-1 and BEAS2B respectively) were infected with rhinovirus strain RV16. Infection of both cell lines resulted in increased inflammation however, the oxidative stress responses were different. Infection of epithelial cells resulted in increased intracellular ROS whereas infection of monocytes decreased intracellular ROS. Infected BEAS2B cells showed decreased antioxidant capacity compared to that seen in THP-1 cells. In particular, infection of THP-1 cells increased expression of the antioxidant geneNRF2and reduced NOS2 and NOS3 gene expression. Using a pharmacological approach we demonstrated that RV16-induced inflammation and NRF2 expression was ERK/JNK-dependent in THP-1 cells, whereas, RV16-induced inflammation in BEAS-2B cells was p38 MAPK- and NF-κB-dependent. Whilst both monocytes and epithelial cells show similar RV-16-induced inflammatory responses, these cells show different oxidative stress responses to infection and the responses to infection are driven by distinct intracellular signalling pathways.

Rationale: Rhinoviruses are the major cause of asthma exacerbations; however, its underlying mechanisms are poorly understood. We hypothesized that the epithelial cell–derived cytokine IL-33 plays a central role in exacerbation pathogenesis through augmentation of type 2 inflammation. Objectives: To assess whether rhinovirus induces a type 2 inflammatory response in asthma in vivo and to define a role for IL-33 in this pathway. Methods: We used a human experimental model of rhinovirus infection and novel airway sampling techniques to measure IL-4, IL-5, IL-13, and IL-33 levels in the asthmatic and healthy airways during a rhinovirus infection. Additionally, we cultured human T cells and type 2 innate lymphoid cells (ILC2s) with the supernatants of rhinovirusinfected bronchial epithelial cells (BECs) to assess type 2 cytokine production in the presence or absence of IL-33 receptor blockade. Measurements and Main Results: IL-4, IL-5, IL-13, and IL-33 are all induced by rhinovirus in the asthmatic airway in vivo and relate to exacerbation severity. Further, induction of IL-33 correlates with viral load and IL-5 and IL-13 levels. Rhinovirus infection of human primary BECs induced IL-33, and culture of human T cells and ILC2s with supernatants of rhinovirus-infected BECs strongly induced type 2 cytokines. This induction was entirely dependent on IL-33. Conclusions: IL-33 and type 2 cytokines are induced during a rhinovirus-induced asthma exacerbation in vivo. Virus-induced IL-33 and IL-33–responsive T cells and ILC2s are key mechanistic links between viral infection and exacerbation of asthma. IL-33 inhibition is a novel therapeutic approach for asthma exacerbations

The BTS took over the entire Queen Elizabeth II Conference Centre in London again this year to host its Silver Jubilee year Winter Meeting. This, the biggest and most comprehensive meeting so far, was also the first to accommodate an additional day for allied health professionals, held in conjunction with the Association of Chartered Physiotherapists in Respiratory Care (ACPRC). In his Presidential address, “Beyond the prescription”, Professor Martyn Partridge focused on the way health care delivery might change in the future with more consultations being delivered in community-based clinics at times which would be more convenient to our patients. The BTS medal was jointly presented to Professor Peter Barnes and Dr Alistair Brewis for their outstanding contributions to respiratory medicine and, at the lively reception, Professor Sue Hill, Chief Scientific Officer at the Department of Health, presented the BTS Silver Jubilee Awards. These covered seven categories celebrating innovation and excellence in respiratory medicine care and service delivery and were a showcase of achievement through teamwork. Also at the reception, the BTS Young Investigator Prize was awarded to Dr David Simcock for his work on airway neovascularisation by airway smooth muscle in asthma.1 The BALR prize went to Dr Yang for his studies on altered gene regulation in familial pulmonary hypertension2 and the BLF prize winner was Dr Kewin for his work on a novel cytokine found to induce eosinophilic airway inflammation.3 Other abstracts submitted for prizes covered a wide range of topics such as statin treatment in hypoxic pulmonary hypertension,4 the search for molecules to block polymerisation of Z α1-antitrypsin5 and the role of vascular endothelial growth factor on the cell cycle of alveolar cells.6 In recognition of the increasing interest and research in COPD, a large proportion of the programme was …

Rhinovirus infection is followed by significantly increased frequencies of positive, potentially pathogenic sputum cultures in chronic obstructive pulmonary disease (COPD). However, it remains unclear whether these represent de novo infections or an increased load of organisms from the complex microbial communities (microbiome) in the lower airways.To investigate the effect of rhinovirus infection on the airway bacterial microbiome.Subjects with COPD (n = 14) and healthy control subjects with normal lung function (n = 17) were infected with rhinovirus. Induced sputum was collected at baseline before rhinovirus inoculation and again on Days 5, 15, and 42 after rhinovirus infection and DNA was extracted. The V3-V5 region of the bacterial 16S ribosomal RNA gene was amplified and pyrosequenced, resulting in 370,849 high-quality reads from 112 of the possible 124 time points.At 15 days after rhinovirus infection, there was a sixfold increase in 16S copy number (P = 0.007) and a 16% rise in numbers of proteobacterial sequences, most notably in potentially pathogenic Haemophilus influenzae (P = 2.7 × 10(-20)), from a preexisting community. These changes occurred only in the sputum microbiome of subjects with COPD and were still evident 42 days after infection. This was in contrast to the temporal stability demonstrated in the microbiome of healthy smokers and nonsmokers.After rhinovirus infection, there is a rise in bacterial burden and a significant outgrowth of Haemophilus influenzae from the existing microbiota of subjects with COPD. This is not observed in healthy individuals. Our findings suggest that rhinovirus infection in COPD alters the respiratory microbiome and may precipitate secondary bacterial infections.

Background Increased mortality rates in patients with chronic obstructive pulmonary disease (COPD) are largely due to severe infectious exacerbations. Impaired respiratory immunity is linked to the enhanced susceptibility to infections. Dendritic cells (DCs) direct host immune responses toward immunity or tolerance. Pulmonary CD1c + DCs elicit robust antiviral immune responses in healthy subjects. Nevertheless, their functional specialization in patients with COPD remains unexplored. Objective We sought to better understand the mechanisms that suppress respiratory immunity in patients with COPD by examining the immunostimulatory and tolerogenic properties of pulmonary CD1c + DCs. Methods We analyzed the expression of costimulatory and tolerogenic molecules by pulmonary CD1c + DCs from patients with COPD (CD1c + DC COPD ) and former smokers without COPD. We isolated lung CD1c + DCs and determined their ability to stimulate allogeneic T-cell responses. The suppressive effects of lung CD1c + DCs and CD1c + DC–primed T cells on mixed leukocyte reactions were examined. An experimental human model of COPD exacerbation was used to investigate the levels of critical immunosuppressive molecules in vivo . Results CD1c + DCs from patients with COPD hinder T-cell effector functions and favor the generation of suppressive IL-10–secreting CD4 + T cells that function through IL-10 and TGF-β. IL-27, IL-10, and inducible T-cell costimulator ligand signaling are essential for CD1c + DC COPD -mediated differentiation of IL-10–producing suppressive T cells. Exposure of lung CD1c + DCs from nonobstructed subjects to lungs of patients with COPD confers tolerogenic properties. IL-27 and IL-10 levels are increased in the lung microenvironment on rhinovirus-induced COPD exacerbation in vivo . Conclusion We identify a novel tolerogenic circuit encompassing suppressive CD1c + DCs and regulatory T cells in patients with COPD that might be implicated in impaired respiratory immunity and further highlight IL-10 and IL-27 as potent therapeutic targets.

Rationale: Chronic obstructive pulmonary disease (COPD) exacerbations are associated with virus (mostly rhinovirus) and bacterial infections, but it is not known whether rhinovirus infections precipitate secondary bacterial infections. Objectives: To investigate relationships between rhinovirus infection and bacterial infection and the role of antimicrobial peptides in COPD exacerbations. Methods: We infected subjects with moderate COPD and smokers and nonsmokers with normal lung function with rhinovirus. Induced sputum was collected before and repeatedly after rhinovirus infection and virus and bacterial loads measured with quantitative polymerase chain reaction and culture. The antimicrobial peptides secretory leukoprotease inhibitor (SLPI), elafin, pentraxin, LL-37, α-defensins and β-defensin-2, and the protease neutrophil elastase were measured in sputum supernatants. Measurements and Main Results: After rhinovirus infection, secondary bacterial infection was detected in 60% of subjects with COPD, 9.5% of smokers, and 10% of nonsmokers (P < 0.001). Sputum virus load peaked on Days 5–9 and bacterial load on Day 15. Sputum neutrophil elastase was significantly increased and SLPI and elafin significantly reduced after rhinovirus infection exclusively in subjects with COPD with secondary bacterial infections, and SLPI and elafin levels correlated inversely with bacterial load. Conclusions: Rhinovirus infections are frequently followed by secondary bacterial infections in COPD and cleavage of the antimicrobial peptides SLPI and elafin by virus-induced neutrophil elastase may precipitate these secondary bacterial infections. Therapy targeting neutrophil elastase or enhancing innate immunity may be useful novel therapies for prevention of secondary bacterial infections in virus-induced COPD exacerbations.

Introduction and Objectives Histone deacetylase (HDAC) enzymes have a role in suppressing inflammatory gene transcription. There is evidence for reduced HDAC activity in COPD which correlates with the severity of airflow obstruction. Increased inflammation found during exacerbations of COPD may result from a reduction in HDAC activity but this has not been studied in virus induced exacerbations. We sought to investigate HDAC activity following rhinovirus infection in an experimental model of COPD exacerbations. Methods Experimental rhinovirus challenge was performed in GOLD stage II COPD subjects and non-obstructed control smokers and non-smokers. Rhinovirus infection was confirmed with quantitative PCR performed on nasal lavage and sputum samples collected at baseline and days 3, 5, 9, 12, 15, 21 and 42 post virus inoculation. Sputum macrophages were isolated by adhesion and HDAC2 isoenzyme immunoprecipitated, prior to performing a HDAC activity assay. Results 11 non-smokers (NS), 10 smokers (Smk) and 9 COPD subjects were recruited. The mean (SEM) % predicted baseline FEV1 was 103 (4), 98 (4) and 69 (2)% for NS, Smk and COPD respectively. At baseline there was no difference in HDAC2 activity between the three study groups, the geometric mean (95% CI) activity was NS 22.39 (12.45 to 40.25); Smk 22.91 (18.28 to 28.71) and COPD 48.98 (26.04 to 92.13) (p=0.095). Following rhinovirus infection, in NS HDAC2 activity increased, in Smk it remained largely unchanged and there was a fall from baseline levels in COPD subjects at all time points, Abstract S14 figure 1. Conclusions HDAC2 activity was not reduced in stable COPD subjects compared to controls. This may reflect the mild disease state of the study population. During a rhinovirus induced exacerbation HDAC2 activity fell in COPD subjects, this represents a potential mechanism of excessive inflammation. The same pattern is not seen in control subjects.

Introduction and Objectives Acute exacerbations are a major cause of morbidity and mortality in COPD and current treatments are not very effective. Histone deacetylase 2 (HDAC2) is deficient in stable COPD and is likely to be a mechanism of corticosteroid resistance. It is not known whether impaired HDAC2 activity is an important mechanism in COPD exacerbations. Methods 9 subjects with GOLD stage II COPD, 10 smokers and 11 non-smokers were infected with rhinovirus 16. Macropahges from induced sputum and bronchoalveolar lavage (BAL) were collected before and following rhinovirus infection and HDAC2 activity measured. Virus load and inflammatory markers were measured in sputum supernantants. Results At baseline there were no differences in HDAC2 activity in sputum or BAL macrophages between the groups. Following infection HDAC2 activity in the smoking controls and non-smoking controls did not change significantly from baseline (Figure 1). In the COPD subjects there was a trend towards reduced HDAC2 activity in both sputum (ANOVA P = 0.064) and BAL macrophages (Paired t test P = 0.098). Sputum HDAC activity was significantly lower in the COPD subjects compared to non-smokers on days 5 and 42 (P Lower sputum macrophage HDAC2 activity at baseline was associated with greater sputum virus load (r = -0.82, P = 0.022) and higher sputum levels of neutrophil elastase (r = -0.81, P = 0.022) and TNF-α (r = -0.79, P = 0.028).HDAC2 activity in BAL macrophages at infection correlated inversely with peak NL virus load (r = -0.8, P = 0.0096), peak sputum GM-CSF (r = -0.67, P = 0.0499), TNF-α (r = -0.72, P = 0.03), neutrophil elastase (r = -0.67, P = 0.0499) and sputum nitrite levels (r = -0.78, P = 0.0125). Conclusions Following rhinovirus infection HDAC2 activity in airway macrophages is reduced and relates to airway inflammatory markers. Restoring HDAC activity is a potential therapeutic option for COPD exacerbations.

Rhinovirus (RV) infection is the most common cause of asthma exacerbations (AE), however mechanisms are poorly understood. Conventional sampling techniques such as bronchoalveolar lavage dilute many cytokines below limits of detection and consequently it has not been possible to measure key mediators of Th1, Th2 and Th17 pathways during virus-induced AE’s . In addition, IL-33, an epithelial-derived alarmin, has recently been shown to be essential for mouse virus-induced airway hyperresponsiveness, however the relationship between IL-33 and exacerbations in human asthma remains unknown. Using the human model of experimental RV induced AE along with novel techniques to sample upper and lower airway mucosal lining fluid (MLF) we investigated the roles of IL-33 and several other prominent cytokines in virus-induced AE’s. Methods 32 mild-to-moderate asthmatics and 14 healthy subjects were inoculated nasally with RV-16. Symptom scores were recorded daily. Bronchoscopies were performed 2 weeks prior to inoculation and on d4 post-inoculation. Novel techniques to sample MLF called ‘bronchosorption’ and ‘nasosorption’ were performed. Cytokines were measured in both bronchial and nasal samples at baseline and on d4 with further nasal sampling on days 2, 3, 5, 7, 10 and 42. Results In asthma, nasal IL-4, –5, –13, –17, –33, and IFN-γ levels were significantly increased during infection compared to baseline (all P Conclusion Sampling MLF permits the direct measurement of previously undetectable mediators across multiple inflammatory pathways. Increased IL-33 and Th2 induction are associated with increased AE severity. IL-33 correlated strongly with Th2 cytokine levels and may represent a novel target for the treatment of virus-induced AE’s. In addition, nasal Th2 inflammation correlated with bronchial levels whilst baseline levels predicted the magnitude of Th2 induction during the AE. Therefore it may be possible to use nasosorption to guide therapy with anti-IL-5 and anti-IL-13 mAb treatments.

Background Rhinovirus (RV) infection is the most common cause of asthma exacerbations (AE). Mechanisms underlying this remain poorly understood. A human model of experimental RV induced AE has been developed however published studies have only recruited subjects with mild, well-controlled asthma naive to inhaled corticosteroid (ICS) therapy. The influence of asthma severity and baseline control on outcome remains unknown. For these studies to be more representative of those who experience virus-induced AE there is a need to establish the safety of using this model in subjects with moderate, poorly-controlled asthma and to investigate clinical outcomes. Method 48 adults - 14 healthy, 14 mild asthmatic, and 18 moderate asthmatic (defined by GINA) were recruited and inoculated nasally with RV-16. Daily symptom scores and spirometry were recorded throughout the study. Asthma control at baseline was recorded using the ACQ. Nasal lavage (NL) for viral shedding was performed on days 0, 2, 3, 4, 5, 7, 10. Clinical infection was confirmed by demonstration of RV16 RNA by RT-PCR in NL and/or serum titre of RV-16 specific antibodies greater than 1:4 on d42. Results 11/14 healthy, 11/14 mild asthmatic and 17/18 moderate asthmatic volunteers met criteria for infection. Both groups of asthmatics developed greater lower respiratory symptoms, falls in FEV1, and airway hyper-responsiveness (AHR) compared to healthy volunteers (all P = P = P = P = Conclusion RV infection results in more severe chest symptoms and falls in lung function in moderate asthma than in mild asthma. Within the moderate group the poorly-controlled asthmatics experienced the most severe exacerbations. This occurred despite therapy with ICS. This is the first study to experimentally inoculate both moderate, poorly-controlled and milder well-controlled asthmatics. Both severity and baseline control appear to influence the outcome of virus-induced AE. Measures to improve control will significantly reduce the likelihood of a severe virus-induced AE and lessen the healthcare costs associated with them.

Introduction and Objectives There is increasing evidence that the majority of acute exacerbations of COPD (AECOPD) are caused by virus infection, and rhinoviruses are the most frequently identified species. Bacteria are also responsible for AECOPD but the relationship between these two is poorly understood. To investigate this further bacterial culture was performed in sputum samples collected from GOLD stage II COPD subjects (n=9) and non-obstructed smoking (n=10) and non-smoking (n=11) controls enrolled in a rhinovirus challenge study. Methods Rhinovirus infection was confirmed with quantitative PCR performed on nasal lavage and sputum samples collected at baseline and days 3, 5, 9, 12, 15, 21 and 42 post virus inoculation. Semi-quantitative bacterial detection was performed in sputum samples by a CPA-accredited microbiological laboratory. Any subject that had bacteria detected on or after day 9 was defined as “bacteria positive” and those with none detected were defined “bacteria negative”. Species frequently associated with respiratory illness were defined as pathogenic ( S pneumoniae, H influenzae, M catarrhalis and S aureus ) and any others as non-pathogenic. Results No bacteria were detected in baseline sputum samples. Peak virus load occurred on day 9 with maximum bacterial colonies identified on day 15. There were significantly more bacteria positive subjects in the COPD group (67%) with the majority of control subjects (81%) being classified as bacteria negative, (Abstract S15 figure 1, χ 2 p=0.04). COPD subjects with bacteria detected at any time point in the study had significantly more pathogenic species in their sputum samples (n=8/8) compared to controls (n=1/9), (χ 2 p=0.0001). No non-pathogenic bacteria were detected in COPD subjects. Conclusions Detection of bacteria is common after rhinovirus infection, with the peak occurring 6 days after maximum virus load. COPD subjects are more likely to have pathogenic bacteria detected than controls following virus infection. Mechanisms responsible for this phenomenon merit further investigation.

Introduction and Objectives The majority of acute exacerbations of COPD are associated with viral infection and rhinoviruses are the most frequently detected species. Exacerbations represent a major unmet health need and mechanisms are poorly understood. The association between nitrative stress and virus induced exacerbations of COPD are unclear. To investigate this we used an experimental rhinovirus challenge study. Methods Experimental rhinovirus challenge was performed in COPD (GOLD stage II) subjects (COPD, n=9), and non-obstructed control smokers (Smk, n=10) and non-smokers (NS, n=11). Rhinovirus infection was confirmed with quantitative PCR performed on nasal lavage and sputum samples collected at baseline and days 3, 5, 9, 12, 15, 21 and 42 post virus inoculation. Nitrite concentration was measured in sputum supernatant using the Griess assay, as a marker of total nitrative stress. Results At baseline the geometric mean (95% CI) nitrite levels were similar between the groups studied (NS 5.13 (3.27 to 8.05); Smk 2.82 (1.37 to 5.80) and COPD 4.17 (3.04 to 5.72); p=0.281). Nitrite levels were significantly higher in COPD subjects on day 15 when compared to both control groups (geometric mean (95% CI) NS 7.94 (7.59 to 8.31); Smk 7.59 (4.41 to 13.04) and COPD 20.98 (13.92 to 31.36); p=0.008). (Abstract S50 figure 1). At every time point sampled there was a significant increase in nitrite concentration from baseline in COPD subjects, but not controls. The area under the curve for nitrite concentration over the time course for NS, Smk and COPD subjects was 18, 43 and 76 respectively (p Conclusions Rhinovirus infection is associated with increased nitrative stress in COPD subjects compared to smoking and non-smoking controls. This may play a role in COPD exacerbations.

Summary Background Bacteria expressing phosphorylcholine (ChoP) co-opt host-expressed platelet-activating factor receptor (PAFR) to adhere to lower airway cells. Cigarette smoke and rhinovirus (RV) infection upregulate PAFR-dependent bacterial adhesion to airway cells in vitro , and in healthy adults smoking increases the proportion of PAFR positive bronchial epithelial cells. To date the effect of chronic obstructive pulmonary disease (COPD) on smoke-induced PAFR is unknown. We therefore sought to test the hypothesis that bronchial PAFR mRNA expression is increased in smokers with chronic obstructive pulmonary disease (COPD), and further increases after RV infection. Methods Endobronchial biopsies were obtained by fibreoptic bronchoscopy from healthy non-smokers, smokers without airway obstruction, and smokers with COPD, before and after infection with rhinovirus (RV) serotype 16. Endobronchial PAFR mRNA expression was assessed by quantitative PCR and expressed as a ratio of glyceraldehyde-3-phosphate dehydrogenase. The distribution of PAFR was assessed by immunohistochemistry. Results Baseline PAFR mRNA expression was increased ( p n = 16), and smokers with COPD ( n = 14) compared with non-smokers ( n = 18). In RV16 infected subjects there was no increase in PAFR mRNA expression in either non-smokers ( n = 9), smokers ( n = 8), or smokers with COPD ( n = 7). PAFR immunoreactivity in all 3 groups was predominately restricted to the bronchial epithelium and submucosal glands. Conclusions Endobronchial PAFR mRNA is increased in both smokers without airway obstruction and smokers with COPD. We found preliminary evidence that RV16 infection does not increase PAFR mRNA expression in either smokers or smokers with COPD.