P J Barnes

Chronic obstructive pulmonary disease: molecular and cellular mechanisms

Chronic obstructive pulmonary disease is a leading cause of death and disability, but has only recently been extensively explored from a cellular and molecular perspective. There is a chronic inflammation that leads to fixed narrowing of small airways and alveolar wall destruction (emphysema). This is characterised by increased numbers of alveolar macrophages, neutrophils and cytotoxic T‐lymphocytes, and the release of multiple inflammatory mediators (lipids, chemokines, cytokines, growth factors). A high level of oxidative stress may amplify this inflammation. There is also increased elastolysis and evidence for involvement of several elastolytic enzymes, including serine proteases, cathepsins and matrix metalloproteinases. The inflammation and proteolysis in chronic obstructive pulmonary disease is an amplification of the normal inflammatory response to cigarette smoke. This inflammation, in marked contrast to asthma, appears to be resistant to corticosteroids, prompting a search for novel anti-inflammatory therapies that may prevent the relentless progression of the disease.

Outcomes for COPD pharmacological trials: from lung function to biomarkers

The American Thoracic Society/European Respiratory Society jointly created a Task Force on “Outcomes for COPD pharmacological trials: from lung function to biomarkers” to inform the chronic obstructive pulmonary disease research community about the possible use and limitations of current outcomes and markers when evaluating the impact of a pharmacological therapy. Based on their review of the published literature, the following document has been prepared with individual sections that address specific outcomes and markers, and a final section that summarises their recommendations.

Scientific rationale for inhaled combination therapy with long-acting β2-agonists and corticosteroids

The addition of an inhaled long-acting β2-agonist (LABA) to an inhaled corticosteroid (ICS) gives optimal control of asthma in most patients and two fixed combination inhalers (salmeterol/fluticasone and formoterol/budesonide) are increasingly used as a convenient controller in patients with persistent asthma. There is a strong scientific rationale for the combination of these two drug classes. ICS suppress the chronic inflammation of asthma and reduce airway hyperresponsiveness and this is achieved at low doses in most patients. LABA act on different aspects of the pathophysiology of asthma. In addition to their bronchodilator action, LABA also inhibit mast cell mediator release, plasma exudation and may reduce sensory nerve activation. Thus these two classes of drug address complementary aspects of the pathophysiology of asthma that neither drug class is able to achieve alone. There are several positive interactions between LABA and ICS. Corticosteroids increase the expression of β2-receptors by increasing gene transcription. Experimentally this protects against the loss of β2-receptors in response to long-term exposure to β2-agonists. While this is unlikely to be important in bronchodilator responses to β2-agonists, in view of the large β-receptor reserve, it is probably important in preventing loss of β-agonist effects on the nonbronchodilator actions of LABA discussed earlier. β2-Agonists may potentiate the molecular mechanism of corticosteroid actions, with increased nuclear localization of glucocorticoid receptors and additive or sometimes synergistic suppression of inflammatory mediator release. Thus LABA and ICS may optimize each others beneficial actions in the airways, but the low systemic effects of these drugs do not result in any increase in adverse effects. Long-acting β2-agonists corticosteroid inhaler therapy is therefore a logical advance and results in effective control of asthma in the majority of patients without significant adverse effects. This simplified approach to long-term asthma therapy has a strong scientific rationale.

Increased nitric oxide in exhaled air of normal human subjects with upper respiratory tract infections

Viral infection may induce the expression of nitric oxide (NO) synthase, resulting in increased NO formation that has an antiviral effect. NO may be produced by various cells of the upper and lower respiratory tract, and may be detected in the exhaled air. We have studied the levels of exhaled NO in 18 normal subjects during symptomatic upper respiratory tract infections and during recovery 3 weeks later. Exhaled NO was measured using a modified chemiluminescence analyser. At the time of symptoms of upper respiratory tract infection, the peak exhaled NO values were 315 +/- 57 ppb (mean +/- SEM) and decreased to 87 +/- 9 ppb during recovery. Recovery values of exhaled NO were similar to those reported in age-matched normal control subjects (88 +/- 3 ppb, n = 72). These findings suggest that symptomatic upper respiratory tract infections markedly increase the concentration of NO in exhaled air. This may reflect the induction of nitric oxide synthase (NOS) in upper and lower respiratory tract, and may be relevant to viral exacerbations of asthma.

Increased expression of nuclear factor-κB in bronchial biopsies from smokers and patients with COPD

The expression of nuclear factor (NF)-κB is an indicator of cellular activation and of inflammatory mediator production. The aim of the present study was to characterise the expression and localisation of p65, the major subunit of NF-κB, in the bronchial mucosa of patients with chronic obstructive pulmonary disease (COPD), and to examine the relationship between p65 expression and disease status. Bronchial biopsies were obtained from 14 smokers with COPD, 17 smokers with normal lung function and 12 nonsmokers with normal lung function. The number of p65 positive (+) cells was quantified by immunohistochemistry and the expression of p65 in bronchial biopsies from the three groups was examined by Western blotting (WB). Smokers with normal lung function and patients with COPD had increased numbers of p65+ cells in the epithelium and increased p65 nuclear expression. In COPD patients the number of epithelial p65+ cells correlated with the degree of airflow limitation. WB analysis showed an increase in p65 in smokers with normal lung function and COPD patients (p<0.05). Bronchial biopsies in smokers with normal lung function and chronic obstructive pulmonary disease patients show increased expression of p65 protein, predominantly in the bronchial epithelium. Disease severity is associated with an increased epithelial expression of nuclear factor-κB.

Increased levels of the chemokines GROα and MCP-1 in sputum samples from patients with COPD

Background: Patients with chronic obstructive pulmonary disease (COPD) have increased numbers of neutrophils and macrophages in their lungs. Growth related oncogene-α (GROα) attracts neutrophils, whereas monocyte chemoattractant protein-1 (MCP-1) attracts monocytes that can differentiate into macrophages. The aim of this study was to determine the concentration of GROα and MCP-1 in bronchoalveolar lavage (BAL) fluid and sputum from non-smokers, healthy smokers and patients with COPD, and to see if there was a correlation between the concentrations of these chemokines, lung function, and numbers of inflammatory cells. Methods: BAL fluid and sputum from non-smokers (n=32), healthy smokers (n=36), and patients with COPD (n=40) were analysed for the presence of GROα and MCP-1 using ELISA. Cells counts were performed on the samples and correlations between the concentrations of these chemokines, lung function, and inflammatory cells observed. Results: Median (SE) GROα and MCP-1 levels were significantly increased in sputum from patients with COPD compared with non-smokers and healthy smokers (GROα: 31 (11) v 2 (2) v 3 (0.8) ng/ml; MCP-1: 0.8 (0.4) v 0.2 (0.1) v 0.1 (0.04) ng/ml, p<0.05), but not in BAL fluid. There were significant negative correlations between both GROα and MCP-1 levels in sputum and forced expiratory volume in 1 second (FEV1) % predicted (GROα: r=–0.5, p<0.001; MCP-1: r=–0.5, p<0.001), together with significant positive correlations between GROα and MCP-1 and neutrophil numbers in sputum (GROα: r=0.6, p<0.001; MCP-1: r=0.4, p<0.01). Conclusion: These results suggest that GROα and MCP-1 are involved in the migration of inflammatory cells, thus contributing to the inflammatory load associated with COPD.

Protective effects of a glucocorticoid on downregulation of pulmonary beta 2-adrenergic receptors in vivo.

We investigated the in vivo effects of a glucocorticoid on beta-agonist-induced downregulation of beta 1- and beta 2-adrenergic receptors (determined by [125I]iodocyanopindolol binding), mRNA expression (assessed by Northern blotting), and gene transcription (using nuclear run-on assays) in rat lung. Dexamethasone (Dex) (0.2 mg/kg/d, days 1-8) increased beta 1- and beta 2-receptor numbers by 70 and 69% above control, respectively, but did not change their mRNA expression. Isoproterenol (Iso) (0.96 mg/kg/d, days 2-8) decreased beta 1- and beta 2-receptor numbers by 48 and 51%, respectively, and also reduced mRNA expression by 69 and 57%, respectively. The combination of Dex and Iso resulted in no net change in beta 2-receptor number and its mRNA expression, although there was a significant reduction in beta 1-receptor number and mRNA expression. The mapping of beta 1- and beta 2-receptors by receptor autoradiography confirmed these findings over alveoli, epithelium, endothelium, and airway and vascular smooth muscle. We also measured the activation of the transcription factor, cyclic AMP response element binding protein (CREB) using an electrophoretic mobility shift assay. CREB-like DNA-binding activity was decreased after Iso treatment but this decrease was prevented after treatment with Dex. Nuclear run-on assays revealed that the transcription rate of the beta 1-receptor gene did not alter after Dex treatment, but was reduced after Iso treatment. The transcription rate of the beta 2-receptor gene was increased after Dex treatment by approximately twofold, but there was no change after Iso treatment. We conclude that glucocorticoids can prevent homologous downregulation of beta 2-receptor number and mRNA expression at the transcriptional level without affecting beta 1-receptors and that the transcription factor CREB may be involved in this phenomenon. Such an effect may have clinical implications for preventing the development of tolerance to beta 2-agonists in asthmatic patients treated with beta-agonist bronchodilators.

COPD: current therapeutic interventions and future approaches

Although long-acting bronchodilators have been an important advance for the management of chronic obstructive pulmonary disease (COPD), these drugs do not deal with the underlying inflammatory process. No currently available treatments reduce the progression of COPD or suppress the inflammation in small airways and lung parenchyma. Several new treatments that target the inflammatory process are now in clinical development. Some therapies, such as chemokine antagonists, are directed against the influx of inflammatory cells into the airways and lung parenchyma that occurs in COPD, whereas others target inflammatory cytokines such as tumour necrosis factor-α. Broad spectrum anti-inflammatory drugs are now in phase III development for COPD, and include phosphodiesterase-4 inhibitors. Other drugs that inhibit cell signalling include inhibitors of p38 mitogen-activated protein kinase, nuclear factor-κB and phosphoinositide-3 kinase-γ. More specific approaches are to give antioxidants, inhibitors of inducible nitric oxide synthase and leukotriene B4 antagonists. Other treatments have the potential to combat mucus hypersecretion, and there is also a search for serine proteinase and matrix metalloproteinase inhibitors to prevent lung destruction and the development of emphysema. More research is needed to understand the cellular and molecular mechanisms of chronic obstructive pulmonary disease and to develop biomarkers and monitoring techniques to aid the development of new therapies.

Relative corticosteroid insensitivity of alveolar macrophages in severe asthma compared with non-severe asthma

Background: About 5–10% of patients with asthma suffer from poorly controlled disease despite corticosteroid (CS) treatment, which may indicate the presence of CS insensitivity. A study was undertaken to determine whether relative CS insensitivity is present in alveolar macrophages from patients with severe asthma and its association with p38 mitogen-activated protein kinase (MAPK) activation and MAPK phosphatase-1 (MKP-1). Methods: Fibreoptic bronchoscopy and bronchoalveolar lavage (BAL) were performed in 20 patients with severe asthma and 19 with non-severe asthma and, for comparison, in 14 normal volunteers. Alveolar macrophages were exposed to lipopolysaccharide (LPS, 10 μg/ml) and dexamethasone (10−8 and 10−6 M). Supernatants were assayed for cytokines using an ELISA-based method. p38 MAPK activity and MKP-1 messenger RNA expression were assayed in cell extracts. Results: The inhibition of LPS-induced interleukin (IL)1β, IL6, IL8, monocyte chemotactic protein (MCP)-1 and macrophage inflammatory protein (MIP)-1α release by dexamethasone (10−6 M) was significantly less in macrophages from patients with severe asthma than in macrophages from patients with non-severe asthma. There was increased p38 MAPK activation in macrophages from patients with severe asthma. MKP-1 expression induced by dexamethasone and LPS, expressed as a ratio of LPS-induced expression, was reduced in severe asthma. Conclusion: Alveolar macrophages from patients with severe asthma demonstrate CS insensitivity associated with increased p38 MAPK activation that may result from impaired inducibility of MKP-1.

Exhaled nitric oxide from lung periphery is increased in COPD

Single constant flow exhaled nitric oxide (eNO) cannot distinguish between the sources of NO. The present study measured eNO at multiple expired flows (MEFeNO) to partition NO into alveolar (Calv,NO) and bronchial (Jaw,NO) fractions to investigate peripheral lung contribution to eNO in chronic obstructive lung disease (COPD). MEFeNO were made in 81 subjects including 18 nonsmokers, 16 smokers and 47 COPD patients of different severity by the classification of the Global Initiative for Chronic Obstructive Lung Disease (GOLD): 0 (n = 14), 1 (n = 7), 2 (n = 11), 3 (n = 8) and 4 (n = 7). COPD severity was correlated with an increased Calv,NO regardless of the patients smoking habit or current treatment. The levels of Calv,NO (in ppb) were 1.4±0.09 in nonsmokers, 2.1±0.1 in smokers categorised as GOLD stage 0 (smokers-GOLD0), 3.3±0.18 in GOLD1–2 and 3.4±0.1 in GOLD3–4. Jaw,NO levels (pL·s−1) were higher in nonsmokers than smokers-GOLD0 (716.2±33.3 versus 464.7±41.8), GOLD3–4 (609.4±71). Diffusion of NO in the airways (Daw,NO pL·ppb−1s−1) was higher (p<0.05) in GOLD3–4 than in nonsmokers (15±1.2 versus 11±0.5) and smokers-GOLD0 (11.6±0.5). MEFeNO measurements were reproducible, free from day-to-day and diurnal variation and were not affected by bronchodilators. In conclusion, chronic obstructive pulmonary disease is associated with elevated alveolar nitric oxide. Measurements of nitric oxide at multiple expired flows may be useful in monitoring inflammation and progression of chronic obstructive pulmonary disease, and the response to anti-inflammatory treatment.