Joseph Footitt

S14 Histone deacetylase activity is reduced in COPD subjects during rhinovirus induced 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.Abstract S14 figure 1 Time course of sputum macrophage HDAC2 activity in individual groups normalised for baseline. Data presented mean±SEM. (Paired t test between day 0 and post infection day † in NS p<0.05; ‡ in COPD p<0.05, ‡‡ in COPD p<0.01). 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.

Celebrating 25 years of the BTS: the Silver Jubilee Meeting

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).nnIn 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.nnThe 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.6nnIn recognition of the increasing interest and research in COPD, a large proportion of the programme was …

S112 HDAC activity in macrophages in experimental rhinovirus infection in COPD

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 < 0.05), and there was a trend towards lower levels of HDAC in BAL macrophages at infection compared to the non-smokers (P = 0.095) and smokers (P = 0.059) (Figure 1). Abstract S112 Figure 1. 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.

S63 Baseline Asthma Control and Severity Influences the Outcome of Virus-Induced Asthma Exacerbations

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=<0.01). These changes were significantly greater in the moderate asthmatics than in the mild asthmatics (P=<0.05). Poorly-controlled asthmatics experienced greater chest symptoms (P=<0.01) and RV-induced falls in lung function (P=<0.05) compared to subjects with well-controlled asthma. 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.

S15 Detection of bacteria in sputum following experimental rhinovirus infection is more common in COPD than controls subjects

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, χ2 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), (χ2 p=0.0001). No non-pathogenic bacteria were detected in COPD subjects.Abstract S15 Figure 1 Number of subjects in each study group with bacteria negative or bacteria positive sputum samples on or following day 9, (χ2 p=0.04). Conclusions Detection of bacteria is common after rhinovirus infection, with the peak occurring 6u2005days 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.

S50 Nitrative stress is increased in COPD exacerbations following experimental rhinovirus infection

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<0.05).Abstract S50 Figure 1 Time course of sputum supernatant nitrite levels in COPD subjects and non-obstructed non-smoking (NS) and smoking (Smk) controls. Data presented mean +/-SEM. (* One-way ANOVA p<0.05). 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.