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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.

The role of neutrophils in exacerbations of asthma is poorly understood. We examined the effect of withdrawal of inhaled corticosteroids on sputum inflammatory indexes in a double-blind study in patients with moderate, stable asthma.Following a 2-week run in period, 24 subjects were randomized to receive either budesonide (400 microg bid) or placebo, and the study was continued for another 10 weeks.Loss of asthma control developed in 8 of 12 patients over the 10-week period of steroid withdrawal, whereas only 1 of 10 patients with budesonide treatment had exacerbations. Those with an exacerbation had increased sputum interleukin (IL)-8 (p0.0001) and increased sputum neutrophil numbers (p0.0001) compared to those without an exacerbation. The significant elevation in sputum IL-8 and neutrophil counts initially occurred 2 weeks prior to an exacerbation. Sputum neutrophilia correlated positively with changes in IL-8 levels (r(2) = 0.76, p = 0.01).Rapid withdrawal of inhaled corticosteroids results in an exacerbation of asthma that is preceded by an increase in sputum neutrophils and IL-8 concentrations, in contrast to an increase in eosinophils reported in previous studies in which inhaled steroids are slowly tapered.

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.

Glucocorticoids are the most effective antiinflammatory agents for the treatment of chronic inflammatory diseases even though some diseases, such as chronic obstructive pulmonary disease (COPD), are relatively glucocorticoid insensitive. However, the molecular mechanism of this glucocorticoid insensitivity remains uncertain. We show that a defect of glucocorticoid receptor (GR) deacetylation caused by impaired histone deacetylase (HDAC) 2 induces glucocorticoid insensitivity toward nuclear factor (NF)-kappaB-mediated gene expression. Specific knockdown of HDAC2 by RNA interference resulted in reduced sensitivity to dexamethasone suppression of interleukin 1beta-induced granulocyte/macrophage colony-stimulating factor production. Loss of HDAC2 did not reduce GR nuclear translocation, GR binding to glucocorticoid response element (GRE) on DNA, or GR-induced DNA or gene induction but inhibited the association between GR and NF-kappaB. GR becomes acetylated after ligand binding, and HDAC2-mediated GR deacetylation enables GR binding to the NF-kappaB complex. Site-directed mutagenesis of K494 and K495 reduced GR acetylation, and the ability to repress NF-kappaB-dependent gene expression becomes insensitive to histone deacetylase inhibition. In conclusion, we show that overexpression of HDAC2 in glucocorticoid-insensitive alveolar macrophages from patients with COPD is able to restore glucocorticoid sensitivity. Thus, reduction of HDAC2 plays a critical role in glucocorticoid insensitivity in repressing NF-kappaB-mediated, but not GRE-mediated, gene expression.

Background: Combination inhaled therapy with long-acting beta(2) agonists (LABAs) and corticosteroids is beneficial in treating asthma and chronic obstructive pulmonary disease (COPD). Objective: In asthma, LABAs enhance glucocorticoid receptor (GR) nuclear translocation in the presence of corticosteroids. Whether this biological mechanism occurs in COPD, a relatively corticosteroid-resistant disease, is uncertain. Methods: Eight patients with mild/moderate COPD participated in a double-blind, placebo-controlled, crossover study and inhaled single doses of fluticasone propionate (FP) 100 mu g, FP 500 mu g, salmeterol xinafoate (SLM) 50 mu g, and combination FP 100 mu g + SLM 50 mu g. One hour postinhalation, sputum was induced, nuclear proteins isolated from purified macrophages, and levels of activated nuclear GR quantified by using a GR-glucocorticoid response element ELISA-based assay. Results: Nuclear GR significantly increased after the inhalation of FP 500 mu g(P

Study objectives: Airway neutrophil levels are increased in patients with severe asthma and during asthma exacerbations. Long-acting 2-agonists (LABAs), such as formoterol, reduce the number of asthma exacerbations. While 2-agonists may affect neutrophil function in vitro ,i t is uncertain whether they have effects on neutrophilic inflammation in asthmatic patients in vivo. Design: In a double-blind randomized crossover study, we evaluated the effects of 4 weeks of treatment with formoterol (Turbuhaler), 24 g bid, compared to placebo on sputum neutrophil numbers and interleukin (IL)-8 levels in asthmatic patients. Therapy with budesonide (administered via Turbuhaler), 400 g bid for 4 weeks, was added at the end as a “gold standard” antiinflammatory effect comparison. Patients: We studied 15 steroid-nao¨ve nonsmoking patients who ranged from 19 to 51 years of age and had mild persistent asthma. Results: Formoterol therapy significantly reduced sputum IL-8 levels and neutrophil numbers compared to placebo. There was a significant correlation between the reduction in sputum IL-8 levels and the number of neutrophils, indicating that formoterol may attenuate neutrophilic airway inflammation by inhibiting IL-8 production. Conclusions: Our data suggest that the LABA formoterol reduces neutrophilic airway inflammation in patients with mild asthma and that this might be beneficial in preventing asthma exacerbations. (CHEST 2005; 128:1936–1942)