A genome-wide analysis of the response to inhaled ß2-agonists in chronic obstructive pulmonary disease.

Short-acting ß2-agonist bronchodilators are the most common medications used in treating chronic obstructive pulmonary disease (COPD). Genetic variants determining bronchodilator responsiveness (BDR) in COPD have not been identified. We performed a genome-wide association study (GWAS) of BDR in 5789 current or former smokers with COPD in one African-American and four white populations. BDR was defined as the quantitative spirometric response to inhaled ß2-agonists. We combined results in a meta-analysis. In the meta-analysis, single-nucleotide polymorphisms (SNPs) in the genes KCNK1 (P=2.02 × 10(-7)) and KCNJ2 (P=1.79 × 10(-7)) were the top associations with BDR. Among African Americans, SNPs in CDH13 were significantly associated with BDR (P=5.1 × 10(-9)). A nominal association with CDH13 was identified in a gene-based analysis in all subjects. We identified suggestive association with BDR among COPD subjects for variants near two potassium channel genes (KCNK1 and KCNJ2). SNPs in CDH13 were significantly associated with BDR in African Americans.The Pharmacogenomics Journal advance online publication, 27 October 2015; doi:10.1038/tpj.2015.65.

Anti-inflammatory effects of atorvastatin treatment in chronic obstructive pulmonary disease. A controlled pilot study.

UNLABELLED: Observational studies have suggested that statins may have beneficial effects on outcomes in chronic obstructive pulmonary disease (COPD) patients. These effects may be mediated through an anti-inflammatory effect of statins. The purpose of this pilot-study was to determine whether statins have an anti-inflammatory effect on the lungs of COPD patients. We conducted randomized, controlled, parallel group pilot-study to compare the effects of atorvastatin (n=12) or placebo (n=6) on lung inflammation in patients with mild to moderate COPD. The primary endpoint was change in CD45+ cells expression measured by immunohistochemistry and changes in expression of genes measured using microarrays in lung biopsy (TBB) samples before and after 12 weeks of treatment with atorvastatin 40 mg/day. All subjects had spirometry, lung volumes, diffusing capacity of the lungs for carbon monoxide (DLCO), St George's Respiratory Questionnaire (SGRQ), 6 minute walk distance (6 MWD), serum lipids, hs-CRP, induced sputum (IS), bronchoscopy and TBB carried out at baseline and after treatment. TBB specimens were processed for histology, immunohistochemistry and genome-wide association studies (GWAS) profiling. Seventeen subjects completed the study. There was a significant improvement in SGRQ with mean SGRQ decreased by 12 points after treatment with atorvastatin (P=0.012). Atorvastatin treatment produced a significant 34% reduction in sputum neutrophil count, and a 57% reduction in CD45+ cells in lung biopsies (expressed as integrated optical density -IOD; median IOD 62.51% before, 27.01% after atorvastatin treatment, P=0.008). In patients' lung tissue atorvastatin treatment produced downregulation of key genes involved in inflammatory processes, immune response, and leukocyte activation. These data demonstrate the pulmonary anti-inflammatory effects of atorvastatin in COPD patients with the potential for beneficial clinical effects. TRIAL REGISTRATION: ClinicalTrials.gov: NCT01748279.

NiO and Co3O4 nanoparticles induce lung DTH-like responses and alveolar lipoproteinosis.

Lung exposure to metal oxide nanoparticles (NPs) comprising soluble metal haptens may produce T-helper cell type 1 (Th1)- and Th17-associated delayed-type hypersensitivity (DTH) responses and pulmonary alveolar proteinosis (PAP). In order to study this, haptenic metal oxide NPs (NiO, Co(3)O(4), Cr(2)O(3) and CuO) were instilled into the lungs of female Wistar rats, and the immunoinflammatory responses were assessed at 24 h and 4 weeks post-instillation. Primary culture of alveolar macrophages from Wistar rats was used to evaluate the effect of the NPs on the ability to clear surfactant. NiO NPs induced chronic interstitial inflammation and pro-inflammatory Th1 and Th17 immune responses characterised by increases in the cytokines monocyte chemotactic protein (MCP)-1/CCL2, interleukin (IL)-12 p40, interferon-γ and IL-17A, whilst similar pathological responses induced by Co(3)O(4) NPs were associated with increases in MCP-1/CCL2 and IL-12 p40. However, neither Cr(2)O(3) nor CuO NPs elicited immunoinflammatory reactions. PAP was induced by both NiO and Co(3)O(4) NPs during the chronic phase. PAP was associated with over-production of surfactant by proliferation of type II cells and impaired clearance of surfactant by macrophages. These findings have implications for the risk management of occupational NP exposure and provide evidence that haptenic metal oxide NPs can induce chronic progressive lung immune responses via a DTH-like mechanism.

Oral antioxidant supplementation does not prevent acute mountain sickness: double blind, randomized placebo-controlled trial.

BACKGROUND: Acute mountain sickness may be caused by cerebrovascular fluid leakage due to oxidative damage to the endothelium. This may be reduced by oral antioxidant supplementation. AIM: To assess the effectiveness of antioxidant supplementation for the prevention of acute mountain sickness (AMS). DESIGN: A parallel-group double blind, randomized placebo-controlled trial. METHODS: The study was conducted in a university clinical research facility and a high altitude research laboratory. Eighty-three healthy lowland volunteers ascended to 5200 m on the Apex 2 high altitude research expedition. The treatment group received a daily dose of 1 g l-ascorbic acid, 400 IU of alpha-tocopherol acetate and 600 mg of alpha-lipoic acid (Cultech Ltd., Wales, UK) in four divided doses. Prevalence of AMS was measured using the Lake Louise Consensus score sheet (LLS). Secondary outcomes were AMS severity measured using a novel visual analogue scale, arterial oxygen saturation and pulmonary artery systolic pressure (PASP). RESULTS: Forty-one subjects were allocated to the antioxidant group, and 42 to the placebo group. There was no difference in AMS incidence or severity between the antioxidant and placebo groups using the LLS at any time at high altitude. At the pre-determined comparison point at Day 2 at 5200 m, 69% of the antioxidant group (25/36) and 66% of the placebo group (23/35) had AMS using the LLS criteria (P = 0.74). No differences were observed between the groups for PASP, oxygen saturation, presence of a pericardial effusion or AMS assessed by VAS. CONCLUSION: This trial found no evidence of benefit from antioxidant supplementation at high altitude. TRIAL REGISTRATION NUMBER: NCT00664001.

Computed tomographic emphysema distribution: relationship to clinical features in a cohort of smokers.

Computed tomography (CT) scanning allows precise assessment of both the extent and distribution of emphysema. There has been little work on the relationship between the distribution of emphysema and clinical features of the disease. The current study investigated the association between clinical features and distribution of emphysema. A total of 129 patients with smoking-related chronic obstructive pulmonary disease underwent CT assessment of the extent and distribution of their emphysema (core/rind and upper/lower zone predominance). Emphysema was found predominantly in the upper/core zone and this distribution was related to the extent of disease. Core predominance was associated with lower forced expiratory volume in one second (FEV(1)), FEV(1)/forced vital capacity ratio and body mass index (BMI); and with higher BODE (BMI, airflow obstruction, dyspnoea and exercise capacity) index and Medical Research Council dyspnoea score. Upper-zone predominance was associated with female sex and an increased total St George's Respiratory Questionnaire score. Using multiple linear regression age, sex and whole lung emphysema severity were independently associated with core/rind distribution, while sex and whole lung emphysema severity were independently related to upper/lower distribution. Distribution of emphysema related best to clinical features when divided into core/rind predominance. However, the effects were not independent of the extent of emphysema. Increased age and female sex were related to disease distribution independent of emphysema severity. These findings may be related to differences in development of emphysema.

Genetic variants of microsomal epoxide hydrolase and glutamate-cysteine ligase in COPD.

The genetic factors that contribute to the development of chronic obstructive pulmonary disease (COPD) are poorly understood. Many candidate genes have been proposed, including enzymes that protect the lung against oxidative stress, such as microsomal epoxide hydrolase (EPHX1) and glutamate-cysteine ligase (GCL). To date, most reported findings have been for EPHX1, particularly in relation to functional variants associated with fast and slow metabolism of epoxide intermediates. The present study aimed to identify any association of variation in these genes with COPD susceptibility or severity. In total, 1,017 white COPD patients and 912 nondiseased age and sex matched smoking controls were genotyped for six single nucleotide polymorphisms (SNPs) in EPHX1 (including the fast and slow variants and associated haplotypes), and eight SNPs in the two genes encoding GCL. GCL is a rate-limiting enzyme in the synthesis of glutathione, a major contributor to anti-oxidant protection in the lung. No association of variation was found in EPHX1 or GCL with susceptibility to COPD or disease severity. This is the largest reported study to date and is well powered to detect associations that have been previously suggested. The current data indicate that these genetic variants are unlikely to be related to susceptibility or disease severity in white chronic obstructive pulmonary disease patients.

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.

Evaluation of COPD Longitudinally to Identify Predictive Surrogate End-points (ECLIPSE).

Chronic obstructive pulmonary disease (COPD) is a heterogeneous disease and not well understood. The forced expiratory volume in one second is used for the diagnosis and staging of COPD, but there is wide acceptance that it is a crude measure and insensitive to change over shorter periods of time. Evaluation of COPD Longitudinally to Identify Predictive Surrogate End-points (ECLIPSE) is a 3-yr longitudinal study with four specific aims: 1) definition of clinically relevant COPD subtypes; 2) identification of parameters that predict disease progression in these subtypes; 3) examination of biomarkers that correlate with COPD subtypes and may predict disease progression; and 4) identification of novel genetic factors and/or biomarkers that both correlate with clinically relevant COPD subtypes and predict disease progression. ECLIPSE plans to recruit 2,180 COPD subjects in Global Initiative for Chronic Obstructive Lung Disease categories II-IV and 343 smoking and 223 nonsmoking control subjects. Study procedures are to be performed at baseline, 3 months, 6 months and every 6 months thereafter. Assessments include pulmonary function measurements (spirometry, impulse oscillometry and plethysmography), chest computed tomography, biomarker measurement (in blood, sputum, urine and exhaled breath condensate), health outcomes, body impedance, resting oxygen saturation and 6-min walking distance. Evaluation of COPD Longitudinally to Identify Predictive Surrogate End-points is the largest study attempting to better describe the subtypes of chronic obstructive pulmonary disease, as well as defining predictive markers of its progression.

Increased arterial stiffness in patients with chronic obstructive pulmonary disease: a mechanism for increased cardiovascular risk.

RATIONALE: Chronic obstructive pulmonary disease (COPD) is associated with a 2-3-fold increase in the risk of ischaemic heart disease, stroke and sudden death. The mechanisms responsible for this association are not clear and appear to be independent of smoking history. OBJECTIVE: We test the hypothesis that patients with COPD have increased arterial stiffness and blood pressure in comparison with age and smoking matched controls. METHODS: In a prospective case control study, we recruited 102 patients with COPD and 103 healthy controls matched for age and smoking status. Patients were assessed by clinical history and spirometry, with arterial stiffness and blood pressure determined using radial artery applanation tonometry and sphygmomanometry. RESULTS: Patients with COPD had increased arterial stiffness compared with matched controls, with elevated augmentation pressure (17 (1) vs 14 (1) mm Hg; p = 0.005) and a reduced time to wave reflection (131 (1) vs 137 (2) ms; p = 0.004). These differences were associated with increases in both diastolic (82 (1) vs 78 (1) mm Hg; p = 0.005) and systolic blood pressure (147 (2) vs 132 (2) mm Hg; p<0.001). Serum C reactive protein concentrations were threefold higher in patients (6.1 (0.9) vs 2.3 (0.4) mg/l; p = 0.001). Data are presented as mean (SEM). CONCLUSIONS: Patients with COPD have increased arterial stiffness and blood pressure in comparison with controls matched for age and smoking status. We speculate that increased systemic inflammation and vascular dysfunction could potentially explain the excess cardiovascular morbidity and mortality associated with COPD.

Interacting effects of particulate pollution and cold temperature on cardiorespiratory mortality in Scotland.

OBJECTIVES: To determine whether the effect of black smoke on cardiorespiratory mortality is modified by cold temperatures. METHODS: Poisson regression models were used to investigate the relationship between lagged black smoke concentration and daily mortality, and whether the effect of black smoke on mortality was modified by cold temperature for three Scottish cities from January 1981 to December 2001. MAIN RESULTS: For all-cause respiratory and non-cardiorespiratory mortality, there was a significant association between mortality and lagged black smoke concentration. Generally the maximum black smoke effect occurred at lag 0, although these estimates were not statistically significant. A 10 mugm(-3) increase in the daily mean black smoke concentration on any given day was associated with a 1.68% (95% CI 0.72 to 2.65) increase in all-cause mortality and a 0.43% (95% CI -0.97 to 1.86), 5.36% (95% CI 2.93 to 7.84) and 2.13% (95% CI 0.82 to 3.47) increase in cardiovascular, respiratory and non-cardiorespiratory mortality, respectively, over the ensuing 30-day period. The effect of black smoke on mortality did not vary significantly between seasons (cool and warm periods). For all-cause, cardiovascular and non-cardiorespiratory mortality the inclusion of interaction terms did not improve the models, although for all-cause and non-cardiorespiratory mortality there was a suggestion for interaction between temperature and recent black smoke exposure. CONCLUSIONS: The results of this study suggested a greater effect of black smoke on mortality at low temperatures. Since extremes of cold and particulate pollution may coexist, for example during temperature inversion, these results may have important public health implications.

Nanoparticle-driven DNA damage mimics irradiation-related carcinogenesis pathways.

The epidemiological association between cancer and exposure to ambient air pollution particles (particles with a 50% cut-off aerodynamic diameter of 10 microm (PM(10))) has been related to the ability of PM(10) and its constituent nanoparticles (NPs) to cause reactive oxidative species (ROS)-driven DNA damage. However, there are no data on the molecular response to these genotoxic effects. In order to assess whether PM(10), NP and ROS-driven DNA damage induce carcinogenesis pathways, A549 cells were treated with tert-butyl-hyperperoxide (Tbh), urban dust (UD), carbon black (CB), nanoparticulate CB (NPCB), benzo(a)pyrene (BaP) and NPCB coated with BaP for

Cytoplasm-nuclear trafficking of CREB and CREB phosphorylation at Ser133 during therapy of chronic obstructive pulmonary disease.

cAMP responsive element binding protein (CREB) plays an important role in transcriptional machinery. CREB signaling is altered in patients with asthma. However, the role of CREB in chronic obstructive pulmonary disease (COPD) is less clear. In the present study we assessed changes in subcellular CREB distribution and activation (CREB-P) in 35 stable COPD patients treated with formoterol (F), formoterol+budesonide (F/ICS), and formoterol+budesonide+theophylline (F/ICS/Th) b.i.d. for 4 weeks, using SDS-PAGE/WB in cytosol and nuclear extracts of induced sputum cells. The expression of CREB was increased after F/ICS in both cytosolic and nuclear fractions by about 40% and 24%, respectively (P<0.001, P<0.01), while CREB-P increased after F/ICS by about 50% (P<0.01) in both compartments. These changes were not affected by theophylline. In F/ICS-treated patients, relative accumulation of CREB in cytosol was observed. These findings indicate, that poor response to ICS therapy may be related to increased CREB-associated signaling.

Nanoparticle carbon black driven DNA damage induces growth arrest and AP-1 and NFkappaB DNA binding in lung epithelial A549 cell line.

To assess whether nanoparticle (NP) driven DNA damage induces the expression of proinflammatory transcription factors such as NFB and AP-1 A549, lung epithelial cells were treated with Carbon Black (CB), nanoparticulate CB (NPCB), NPCB coated with BaP (BaP-NPCB) for various times ranging from 30 min to 24 h. DNA strand break was determined by the comet assay and cell cycle status was analyzed using flow cytometry. Nuclear extracts were used for WB analysis of P approximately Ser15-p53. EMSA was used to detect DNA binding. Tested NP caused single strand breaks and significantly altered cell cycle kinetics. NF-kappaB and AP-1 DNA binding were increased at early time points (2.3 and 2.6 fold at 1 hour, respectively). Effects were also found on Ser15-p53 phosphorylation. N-acetylcysteine blocked NP driven effects. In conclusion, NPCB and BaP-NPCB induce DNA damage, activating p53, proteins related to DNA repair and proinflammatory transcription factors.

Telomere shortening in smokers with and without COPD.

Telomeres are complex DNA-protein structures located at the end of eukaryotic chromosomes. Telomere length shortens with age in all replicating somatic cells. It has been shown that tobacco smoking enhances telomere shortening in circulating lymphocytes. The present study investigated whether this effect was further amplified in smokers who develop chronic obstructive pulmonary disease. Telomere length was determined by fluorescence in situ hybridisation in circulating lymphocytes harvested from 26 never-smokers, 24 smokers with normal lung function and 26 smokers with moderate-to-severe airflow obstruction (forced expiratory flow in one second 48+/-4% predicted). In contrast to never-smokers, telomere length significantly decreased with age in smokers. There was also a dose-effect relationship between the cumulative long-life exposure to tobacco smoking (pack-yrs) and telomere length. The presence and/or severity of chronic airflow obstruction did not modify this relationship. The results of the current study confirm that smoking exposure enhances telomere shortening in circulating lymphocytes. It also demonstrates a dose-effect relationship between exposure to tobacco smoking and telomere length, but failed to show that this effect is amplified in smokers who develop chronic obstructive pulmonary disease.

The potential impact of anaemia of chronic disease in COPD.

Anaemia of chronic disease (ACD), with chronically low levels of circulating haemoglobin, is an immune driven abnormality that occurs in many inflammatory diseases, and also in chronic heart failure. Although chronic obstructive pulmonary disease (COPD) is "traditionally" associated with polycythaemia, the systemic inflammation that is now recognised as a feature of COPD makes it a possible cause of ACD. If present in COPD, anaemia could worsen dyspnoea and limit exercise tolerance. Preliminary evidence suggests that anaemia in COPD patients may be more prevalent than expected, concerning 10-15% of patients suffering from severe forms of the disease. A database study conducted in 2,524 COPD patients being prescribed long-term oxygen therapy has shown that a low haematocrit is a strong predictor of survival in this population, before body mass index, and is associated with more hospitalisations and a longer cumulative duration of hospitalisation. COPD patients with low haemoglobin levels have a poorer prognosis than COPD patients with normal haemoglobin levels in the event of acute gastrointestinal bleeding or after elective aneurysm repair. Raising haemoglobinaemia through transfusion decreases minute ventilation and work of breathing in COPD patients. These preliminary evidences point to the need to study the prevalence of anaemia, and its physiological and clinical impact in chronic obstructive pulmonary disease. When this body of knowledge is available, the question of the putative benefits of raising haemoglobinaemia in chronic obstructive pulmonary disease will have to be addressed.