Shared genetic aetiology of respiratory diseases: a genome-wide multitraits association analysis.

OBJECTIVE: This study aims to explore the common genetic basis between respiratory diseases and to identify shared molecular and biological mechanisms. METHODS: This genome-wide pleiotropic association study uses multiple statistical methods to systematically analyse the shared genetic basis between five respiratory diseases (asthma, chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis, lung cancer and snoring) using the largest publicly available genome wide association studies summary statistics. The missions of this study are to evaluate global and local genetic correlations, to identify pleiotropic loci, to elucidate biological pathways at the multiomics level and to explore causal relationships between respiratory diseases. Data were collected from 27 November 2022 to 30 March 2023 and analysed from 14 April 2023 to 13 July 2023. MAIN OUTCOMES AND MEASURES: The primary outcomes are shared genetic loci, pleiotropic genes, biological pathways and estimates of genetic correlations and causal effects. RESULTS: Significant genetic correlations were found for 10 paired traits in 5 respiratory diseases. Cross-Phenotype Association identified 12 400 significant potential pleiotropic single-nucleotide polymorphism at 156 independent pleiotropic loci. In addition, multitrait colocalisation analysis identified 15 colocalised loci and a subset of colocalised traits. Gene-based analyses identified 432 potential pleiotropic genes and were further validated at the transcriptome and protein levels. Both pathway enrichment and single-cell enrichment analyses supported the role of the immune system in respiratory diseases. Additionally, five pairs of respiratory diseases have a causal relationship. CONCLUSIONS AND RELEVANCE: This study reveals the common genetic basis and pleiotropic genes among respiratory diseases. It provides strong evidence for further therapeutic strategies and risk prediction for the phenomenon of respiratory disease comorbidity.

Burden of COPD in China and the global from 1990 to 2019: a systematic analysis for the Global Burden of Disease Study 2019.

OBJECTIVE: To investigate the current disease burden of chronic obstructive pulmonary disease (COPD) in China and globally using the Global Burden of Disease (GBD) data in 2019, as well as to analyse the changes in its risk factors, providing a scientific basis for the formulation of a comprehensive prevention and control strategy for COPD in China. STUDY DESIGN: An observational study based on the GBDs. METHODS: Based on the GBD 2019 database, we obtained data on incidence, prevalence, mortality, disability-adjusted life years (DALYs) and corresponding age-standardised rates of COPD in China and the global, and analysed and described the changing trends of COPD burden in China and the global from 1990 to 2019. RESULTS: In 2019, the total number of COPD deaths in China was 1.04 (95% uncertainty intervals (95% UI): 0.89-1.27) million cases, the number of patients with COPD was 45.16 (95% UI: 41.13-49.62) million cases, and the number of new cases was 4.0 (95% UI: 3.6-4.4) million cases. DALYs were 74.4 (95% UI: 68.2-80.2) million years. Compared with 1990, the number of new incident cases and the overall prevalence of COPD in China in 2019 increased by 66.20% and 66.76%, respectively, which is lower than the overall global level. CONCLUSION: From 1990 to 2019, the age-standardized prevalence rate (ASPR), the age-standardized incidence rate (ASIR) and the age-standardized death rate (ASDR) in China and the global all showed a downward trend, and the rate of decline in China was much higher than the overall level of the world, indicating that China has made specific achievements in the prevention and treatment of COPD, but overall the disease burden of COPD is still hefty, and the number of affected individuals is still increasing.

Remote exposure to secondhand tobacco smoke is associated with lower exercise capacity through effects on oxygen pulse, a proxy of cardiac stroke volume.

BACKGROUND: Past exposure to secondhand tobacco smoke (SHS) is associated with exercise limitation. Pulmonary factors including air trapping contribute to this limitation but the contribution of cardiovascular factors is unclear. OBJECTIVE: To determine the contribution of cardiovascular mechanisms to SHS-associated exercise limitation. METHODS: We examined the cardiovascular responses to maximum-effort exercise in 245 never-smokers with remote, prolonged occupational exposure to SHS and no known history of cardiovascular disease. We estimated the contribution of oxygen-pulse (proxy for cardiac stroke volume) and changes in systolic blood pressures (SBP), diastolic blood pressures and heart rate (HR) towards exercise capacity, and examined whether the association of SHS with exercise capacity was mediated through these variables. RESULTS: At peak exercise (highest workload completed (WattsPeak)=156±46 watts (135±33 %predicted)), oxygen consumption and oxygen-pulse (O2-PulsePeak) were 1557±476 mL/min (100±24 %predicted) and 11.0±3.0 mL/beat (116±25 %predicted), respectively, with 29% and 3% participants not achieving their predicted normal range. Oxygen saturation at peak exercise was 98%±1% and remained >93% in all participants. Sixty-six per cent showed hypertensive response to exercise. In models adjusted for covariates, WattsPeak was associated directly with O2-PulsePeak, HRPeak and SBPPeak and inversely with SHS, air trapping (residual volume/total lung capacity) and rise of SBP over workload (all p<0.01). Moreover, SHS exposure association with WattsPeak was substantially (41%) mediated through its effect on O2-PulsePeak (p=0.038). Although not statistically significant, a considerable proportion (36%) of air trapping effect on WattsPeak seemed to be mediated through O2-PulsePeak (p=0.078). The likelihood of having baseline respiratory symptoms (modified Medical Research Council score ≥1) was associated with steeper rise in SBP over workload (p<0.01). CONCLUSION: In a never-smoker population with remote exposure to SHS, abnormal escalation of blood pressure and an SHS-associated reduction in cardiac output contributed to lower exercise capacity.

Implications of using the GLI-2012, GOLD and Australian COPD-X recommendations in assessing the severity of airflow limitation on spirometry among an Indigenous population with COPD: an Indigenous Australians perspective study.

BACKGROUND: Assessment of airflow limitation (AFL) is crucial in the clinical evaluation of patients with chronic obstructive pulmonary disease (COPD). However, in the absence of normative reference values among adult Australian Indigenous population, the implications of utilising the Global Lung Function Initiative (GLI-2012), Global Initiative for Chronic Obstructive Lung Disease (GOLD) and the Australian concise COPD-X recommended severity classifications is not known. Moreover, spirometry values (forced vital capacity (FVC) and forced expiratory volume in 1 s (FEV1)) are observed to be 20%-30% lower in an apparently healthy Indigenous population in comparison to Caucasian counterparts. METHODS: Adult Indigenous patients diagnosed to have COPD on spirometry (postbronchodilator (BD) FEV1/FVC <0.7 ((GOLD, (COPD-X)) and ≤lower limit of normal (others/mixed reference equations) for GLI-2012) were assessed for AFL severity classifications on Post-BD FEV1 values (mild, moderate, severe, very severe) as per the recommended classifications. RESULTS: From a total of 742 unique patient records of Indigenous Australians, 253 were identified to have COPD via GOLD/COPD-X criteria (n=238) or GLI-2012 criteria (n=238) with significant agreeance between criteria (96%, κ=0.901). Of these, the majority were classified as having moderate or severe/very-severe AFL with significant variability across classification criteria (COPD-X (40%-43%), GOLD (33%-65%), GLI-2012 (18%-75%)). The FVC and FEV1 values also varied significantly between classification criterion (COPD-X/GOLD/GLI-2012) within the same AFL category, with COPD-X 'moderate' AFL almost matching 'severe' AFL categorisation by GOLD or GLI-2012. CONCLUSIONS: Health professionals caring for Indigenous patients with COPD should be aware of the clinical implications and consequences of utilising various recommended AFL classifications in the absence of validated spirometry reference norms among adult Indigenous patients.

Role of the IL-33/ST2 axis in cigarette smoke-induced airways remodelling in chronic obstructive pulmonary disease.

BACKGROUND: Efficient therapy and potential prophylaxis are confounded by current ignorance of the pathogenesis of airway remodelling and blockade in COPD. OBJECTIVE: To explore the role of the IL-33/ST2 axis in cigarette smoke (CS) exposure-induced airways remodelling. METHODS: C57BL/6, BALB/c and IL-1RL1 -/- mice exposed to CS were used to establish an animal surrogate of COPD (air-exposed=5~8, CS-exposed=6~12). Hallmarks of remodelling were measured in mice. Cigarette smoke extract (CSE)-induced proliferation and protein production in vitro by fibroblasts in the presence of anti-interleukin-33 (anti-IL-33) or hST2 antibodies were measured. Expression of IL-33 and ST2 and other remodelling hallmarks were measured, respectively, in bronchoalveolar lavage fluid (BALF) (controls=20, COPD=20), serum (controls=59, COPD=90) and lung tissue sections (controls=11, COPD=7) from patients with COPD and controls. RESULTS: Wild-type mice exposed to CS elevated expression of hallmarks of tissue remodelling in the lungs and also in the heart, spleen and kidneys, which were significantly abrogated in the IL-1RL1 -/- mice. Fibroblasts exposed to CSE, compared with control, exhibited early cellular translocation of IL-33, accompanied by proliferation and elevated protein synthesis, all inhabitable by blockade of IL-33/ST2 signalling. Expression of IL-33 and ST2 and hallmarks of tissue remodelling were significantly and proportionally elevated in BALF, serum and tissue samples from patients with COPD. CONCLUSIONS: Exposure to CS induces remodelling changes in multiple organs. The data support the hypothesis that CS-induced lung collagen deposition is at least partly a result of CS-induced IL-33 translocation and release from local fibroblasts.

BET proteins are associated with the induction of small airway fibrosis in COPD.

RATIONALE: In COPD, small airway fibrosis occurs due to increased extracellular matrix (ECM) deposition in and around the airway smooth muscle (ASM) layer. Studies of immune cells and peripheral lung tissue have shown that epigenetic changes occur in COPD but it is unknown whether airway mesenchymal cells are reprogrammed. OBJECTIVES: Determine if COPD ASM cells have a unique epigenetic response to profibrotic cytokine transforming growth factor ß1 (TGF-ß1). METHODS: Primary human ASM cells from COPD and non-COPD smoking patients were stimulated with TGF-ß1. Gene array analysis performed to identify differences in ECM expression. Airway accumulation of collagen 15α1 and tenascin-C proteins was assessed. Aforementioned ASM cells were stimulated with TGF-ß1 ± epigenetic inhibitors with qPCR quantification of COL15A1 and TNC. Global histone acetyltransferase (HAT) and histone deacetylase (HDAC) activity were assessed. chromatin immunoprecipitation (ChIP)-qPCR for histone H3 and H4 acetylation at COL15A1 and TNC promoters was carried out. Effects of bromoterminal and extraterminal domain (BET) inhibitor JQ1(+) on expression and acetylation of ECM target genes were assessed. MEASUREMENTS AND MAIN RESULTS: COPD ASM show significantly higher COL15A1 and TNC expression in vitro and the same trend for higher levels of collagen 15α1 and tenascin-c deposited in COPD airways in vivo. Epigenetic screening indicated differential response to HDAC inhibition. ChIP-qPCR revealed histone H4 acetylation at COL15A1 and TNC promoters in COPD ASM only. ChIP-qPCR found JQ1(+) pretreatment significantly abrogated TGF-ß1 induced histone H4 acetylation at COL15A1 and TNC. CONCLUSIONS: BET protein binding to acetylated histones is important in TGF-ß1 induced expression of COL15A1 and TNC and maintenance of TGF-ß1 induced histone H4 acetylation in cell progeny.

DNA methylation is associated with airflow obstruction in patients living with HIV.

INTRODUCTION: People living with HIV (PLWH) suffer from age-related comorbidities such as COPD. The processes responsible for reduced lung function in PLWH are largely unknown. We performed an epigenome-wide association study to investigate whether blood DNA methylation is associated with impaired lung function in PLWH. METHODS: Using blood DNA methylation profiles from 161 PLWH, we tested the effect of methylation on FEV1, FEV1/FVC ratio and FEV1 decline over a median of 5 years. We evaluated the global methylation of PLWH with airflow obstruction by testing the differential methylation of transposable elements Alu and LINE-1, a well-described marker of epigenetic ageing. RESULTS: Airflow obstruction as defined by a FEV1/FVC<0.70 was associated with 1393 differentially methylated positions (DMPs), while 4676 were associated with airflow obstruction based on the FEV1/FVC

Elevated plasma level of Pentraxin 3 is associated with emphysema and mortality in smokers.

BACKGROUND: Pentraxin 3 (PTX3) influences innate immunity and inflammation, host defence, the complement cascade and angiogenesis. PTX3 expression in lung and blood of subjects with tobacco exposure, and its potential relationship with disease pattern and clinical outcome are poorly understood. METHODS: Using independent platforms and cohorts, we identified associations of PTX3 gene expression in lung tissue and plasma from current and former tobacco smokers (with and without chronic obstructive pulmonary disease, COPD) to disease phenotypes including quantitative CT determined emphysema, lung function, symptoms and survival. Two putative regulatory variants of the PTX3 gene were examined for association with COPD manifestations. The relationship between plasma PTX3 and hyaluronic acid levels was further examined. RESULTS: PTX3 gene expression in lung tissue was directly correlated with emphysema severity (p<0.0001). Circulating levels of PTX3 were inversely correlated with FEV1 (p=0.006), and positively associated with emphysema severity (p=0.004) and mortality (p=0.008). Two PTX3 gene regulatory variants were associated with a lower risk for emphysema and expiratory airflow obstruction, and plasma levels of PTX3 and hyaluronic acid were related. CONCLUSIONS: These data show strong and overlapping associations of lung and blood PTX3 levels, and PTX3 regulatory gene variants, with the severity of airflow obstruction, emphysema and mortality among smokers. These findings have potential implications regarding the pathogenesis of smoking-related lung diseases and warrant further exploration for the use of PTX3 as a predictive biomarker.

Characterisation of gas exchange in COPD with dissolved-phase hyperpolarised xenon-129 MRI.

To investigate whether hyperpolarised xenon-129 MRI (HXeMRI) enables regional and physiological resolution of diffusing capacity limitations in chronic obstructive pulmonary disease (COPD), we evaluated 34 COPD subjects and 11 healthy volunteers. We report significant correlations between airflow abnormality quantified by HXeMRI and per cent predicted forced expiratory volume in 1 s; HXeMRI gas transfer capacity to red blood cells and carbon monoxide diffusion capacity (%DLCO); and HXeMRI gas transfer capacity to interstitium and per cent emphysema quantified by multidetector chest CT. We further demonstrate the capability of HXeMRI to distinguish varying pathology underlying COPD in subjects with low %DLCO and minimal emphysema.

Associations of CT evaluations of antigravity muscles, emphysema and airway disease with longitudinal outcomes in patients with COPD.

Multiple CT indices are associated with disease progression and mortality in patients with COPD, but which indices have the strongest association remain unestablished. This longitudinal 10-year observational study (n=247) showed that the emphysema severity on CT is more closely associated with the progression of airflow limitation and that a reduction in the cross-sectional area of erector spinae muscles (ESMCSA) on CT is more closely associated with mortality than the other CT indices, independent of patient demographics and pulmonary function. ESMCSA is a useful CT index that is more closely associated with long-term mortality than emphysema and airway disease in patients with COPD.

FN3K expression in COPD: a potential comorbidity factor for cardiovascular disease.

INTRODUCTION: Cigarette smoking and oxidative stress are common risk factors for the multi-morbidities associated with chronic obstructive pulmonary disease (COPD). Elevated levels of advanced glycation endproducts (AGE) increase the risk of cardiovascular disease (CVD) comorbidity and mortality. The enzyme fructosamine-3-kinase (FN3K) reduces this risk by lowering AGE levels. METHODS: The distribution and expression of FN3K protein in lung tissues from stable COPD and control subjects, as well as an animal model of COPD, was assessed by immunohistochemistry. Serum FN3K protein and AGE levels were assessed by ELISA in patients with COPD exacerbations receiving metformin. Genetic variants within the FN3K and FN3K-RP genes were evaluated for associations with cardiorespiratory function in the Subpopulations and Intermediate Outcome Measures in COPD Study cohort. RESULTS: This pilot study demonstrates that FN3K expression in the blood and human lung epithelium is distributed at either high or low levels irrespective of disease status. The percentage of lung epithelial cells expressing FN3K was higher in control smokers with normal lung function, but this induction was not observed in COPD patients nor in a smoking model of COPD. The top five nominal FN3K polymorphisms with possible association to decreased cardiorespiratory function (p<0.008-0.02), all failed to reach the threshold (p<0.0028) to be considered highly significant following multi-comparison analysis. Metformin enhanced systemic levels of FN3K in COPD subjects independent of their high-expression or low-expression status. DISCUSSION: The data highlight that low and high FN3K expressors exist within our study cohort and metformin induces FN3K levels, highlighting a potential mechanism to reduce the risk of CVD comorbidity and mortality.

Protective effect of club cell secretory protein (CC-16) on COPD risk and progression: a Mendelian randomisation study.

BACKGROUND: The anti-inflammatory pneumoprotein club cell secretory protein-16 (CC-16) is associated with the clinical expression of chronic obstructive pulmonary disease (COPD). We aimed to determine if there is a causal effect of serum CC-16 level on the risk of having COPD and/or its progression using Mendelian randomisation (MR) analysis. METHODS: We performed a genome-wide association meta-analysis for serum CC-16 in two COPD cohorts (Lung Health Study (LHS), n=3850 and ECLIPSE, n=1702). We then used the CC-16-associated single-nucleotide polymorphisms (SNPs) as instrumental variables in MR analysis to identify a causal effect of serum CC-16 on 'COPD risk' (ie, case status in the International COPD Genetics Consortium/UK-Biobank dataset; n=35 735 COPD cases, n=222 076 controls) and 'COPD progression' (ie, annual change in forced expiratory volume in 1 s in LHS and ECLIPSE). We also determined the associations between SNPs associated with CC-16 and gene expression using n=1111 lung tissue samples from the Lung Expression Quantitative Trait Locus Study. RESULTS: We identified seven SNPs independently associated (p<5×10-8) with serum CC-16 levels; six of these were novel. MR analysis suggested a protective causal effect of increased serum CC-16 on COPD risk (MR estimate (SE) -0.11 (0.04), p=0.008) and progression (LHS only, MR estimate (SE) 7.40 (3.28), p=0.02). Five of the SNPs were also associated with gene expression in lung tissue (at false discovery rate <0.1) of several genes, including the CC-16-encoding gene SCGB1A1. CONCLUSION: We have identified several novel genetic variants associated with serum CC-16 level in COPD cohorts. These genetic associations suggest a potential causal effect of serum CC-16 on the risk of having COPD and its progression, the biological basis of which warrants further investigation.

Extracorporeal CO2 removal (ECCO2R) in patients with stable COPD with chronic hypercapnia: a proof-of-concept study.

Domiciliary non-invasive ventilation (NIV) effectively reduces arterial carbon dioxide pressure (PaCO2) in patients with stable hypercapnic chronic obstructive pulmonary disease, but a consistent percentage of them may remain hypercapnic. We hypothesised that extracorporeal CO2 removal (ECCO2R) may lower their PaCO2 Ten patients hypercapnic despite ≥6 months of NIV underwent a 24-hour trial of ECCO2R. Six patients completed the ECCO2R-trial with a PaCO2 drop ranging between 23% and 47%. Time to return to baseline after interruption ranged 48-96 hours. In four patients, mechanical events led to ECCO2R premature interruption, despite a decreased in PaCO2 This time window 'free' from hypercapnia might allow to propose the concept of 'CO2 dialysis'.

Per cent low attenuation volume and fractal dimension of low attenuation clusters on CT predict different long-term outcomes in COPD.

BACKGROUND: Fractal dimension (D) characterises the size distribution of low attenuation clusters on CT and assesses the spatial heterogeneity of emphysema that per cent low attenuation volume (%LAV) cannot detect. This study tested the hypothesis that %LAV and D have different roles in predicting decline in FEV1, exacerbation and mortality in patients with COPD. METHODS: Chest inspiratory CT scans in the baseline and longitudinal follow-up records for FEV1, exacerbation and mortality prospectively collected over 10 years in the Hokkaido COPD Cohort Study were examined (n=96). The associations between CT measures and long-term outcomes were replicated in the Kyoto University cohort (n=130). RESULTS: In the Hokkaido COPD cohort, higher %LAV, but not D, was associated with a greater decline in FEV1 and 10-year mortality, whereas lower D, but not %LAV, was associated with shorter time to first exacerbation. Multivariable analysis for the Kyoto University cohort confirmed that lower D at baseline was independently associated with shorter time to first exacerbation and that higher LAV% was independently associated with increased mortality after adjusting for age, height, weight, FEV1 and smoking status. CONCLUSION: These well-established cohorts clarify the different prognostic roles of %LAV and D, whereby lower D is associated with a higher risk of exacerbation and higher %LAV is associated with a rapid decline in lung function and long-term mortality. Combination of %LAV and fractal D may identify COPD subgroups at high risk of a poor clinical outcome more sensitively.

Integrated proteogenomic approach identifying a protein signature of COPD and a new splice variant of SORBS1.

Translation of genomic alterations to protein changes in chronic obstructive pulmonary disease (COPD) is largely unexplored. Using integrated proteomic and RNA sequencing analysis of COPD and control lung tissues, we identified a protein signature in COPD characterised by extracellular matrix changes and a potential regulatory role for SUMO2. Furthermore, we identified 61 differentially expressed novel, non-reference, peptides in COPD compared with control lungs. This included two peptides encoding for a new splice variant of SORBS1, of which the transcript usage was higher in COPD compared with control lungs. These explorative findings and integrative proteogenomic approach open new avenues to further unravel the pathology of COPD.

Anaemia and iron dysregulation: untapped therapeutic targets in chronic lung disease?

Hypoxia is common in many chronic lung diseases. Beyond pulmonary considerations, delivery of oxygen (O2) to the tissues and subsequent O2 utilisation is also determined by other factors including red blood cell mass and iron status; consequently, disruption to these mechanisms provides further physiological strains on an already stressed system. O2 availability influences ventilation, regulates pulmonary blood flow and impacts gene expression throughout the body. Deleterious effects of poor tissue oxygenation include decreased exercise tolerance, increased cardiac strain and pulmonary hypertension in addition to pathophysiological involvement of multiple other organs resulting in progressive frailty. Increasing inspired O2 is expensive, disliked by patients and does not normalise tissue oxygenation; thus, other strategies that improve O2 delivery and utilisation may provide novel therapeutic opportunities in patients with lung disease. In this review, we focus on the rationale and possibilities for doing this by increasing haemoglobin availability or improving iron regulation.

Minimum important difference of the incremental shuttle walk test distance in patients with COPD.

We aimed to describe the minimum important difference (MID) of the incremental shuttle walk test (ISWT) in patients with COPD using both distribution and anchor-based methods. Two cohorts were used (n=613) with eligibility criteria of a clinical diagnosis of COPD, an FEV1/FVC <70% and an ISWT (after familiarisation) before and after a 7-week course of pulmonary rehabilitation (PR). The MID of the ISWT using the distribution method was 36.1 m. The area under the curve to discriminate between perceived 'improvement' and 'no improvement' after PR for a change in ISWT of 35 m was 0.66 (0.58-0.73). The MID of the ISWT is therefore between 35.0 and 36.1 m.

Lung volume indices predict morbidity in smokers with preserved spirometry.

BACKGROUND: Abnormal lung volumes that reflect air trapping are common in COPD. However, their significance in smokers with preserved spirometry (normal FEV1 to FVC ratio) is unclear. METHODS: Using the Veterans Administration Informatics and Computing Infrastructure database, we identified 7479 patients at risk for COPD (ever smokers >40 years of age without restrictive lung disease) who had preserved spirometry and concomitant lung volume measurements, and examined their subsequent health records for clinical diagnoses of COPD, healthcare utilisation, follow-up spirometry and mortality. RESULTS: Air trapping was prevalent, with 31% of patients having residual volume to total lung capacity ratio (RV:TLC) greater than the upper limit of normal (ULN). RV:TLC varied widely from 14% to 77% (51% to 204% of predicted) across the normal ranges of FEV1:FVC and FEV1. Patients with RV:TLC greater than the ULN were more likely to receive subsequent clinical diagnoses of COPD (HR (95% CI)=1.55 (1.42 to 1.70), p<0.001) and had higher all-cause mortality (HR (95% CI)=1.41 (1.29 to 1.54), p<0.001). They had higher rates of respiratory medication prescriptions and hospital and intensive care unit admissions. Other air trapping and static hyperinflation indices showed similar associations with health outcomes. Additionally, high-normal RV:TLC was associated with intermediate adverse health outcomes compared with low-normal and abnormal RV:TLC. Abnormal RV:TLC predicted higher likelihood of progression to spirometric COPD (OR (95% CI)=1.30 (1.03 to 1.65), p=0.027). CONCLUSION: In this study of the Veterans Affairs electronic health records, air trapping was common in smokers with preserved spirometry and predicted adverse respiratory outcomes and progression to overt COPD.