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Original Research

Airway inflammation in Japanese COPD patients compared with smoking and nonsmoking controls

, , , , &
Pages 185-192 | Published online: 23 Jan 2015

Abstract

Purpose

To assess the importance of inflammation in chronic obstructive pulmonary disease (COPD) by measuring airway and systemic inflammatory biomarkers in Japanese patients with the disease and relevant control groups.

Patients and methods

This was the first study of its type in Japanese COPD patients. It was a non-treatment study in which 100 participants were enrolled into one of three groups: nonsmoking controls, current or ex-smoking controls, and COPD patients. All participants underwent standard lung function assessments and provided sputum and blood samples from which the numbers of inflammatory cells and concentrations of biomarkers were measured, using standard procedures.

Results

The overall trends observed in levels of inflammatory cells and biomarkers in sputum and blood in COPD were consistent with previous reports in Western studies. Increasing levels of neutrophils, interleukin 8 (IL-8), surfactant protein D (SP-D), and Krebs von den Lungen 6 (KL-6) in sputum and clara cell 16 (CC-16), high-sensitivity C-reactive protein (hs-CRP), and KL-6 in serum and plasma fibrinogen were seen in the Japanese COPD patients compared with the non-COPD control participants. In sputum, significant correlations were seen between total cell count and matrix metalloproteinase 9 (MMP-9; P<0.001), neutrophils and MMP-9 (P<0.001), macrophages and KL-6 (P<0.01), total cell count and IL-8 (P<0.05), neutrophils and IL-8 (P<0.05), and macrophages and MMP-9 (P<0.05). Significant correlations were also observed between some inflammatory cells in sputum and biomarkers in serum, with the most significant between serum CC-16 and both total cell count (P<0.005) and neutrophils (P<0.005) in sputum.

Conclusion

These results provide evidence for the first time that COPD in Japanese patients is a multicomponent disease, involving both airway and systemic inflammation, in addition to airway obstruction. Therefore, intervention with anti-inflammatory therapy may provide additional benefit in disease management of COPD in Japan.

Introduction

Chronic obstructive pulmonary disease (COPD) is the fourth leading cause of death globally, with a projected increase in burden resulting from continued exposure to COPD risk factors and an aging population.Citation1 From 1995 to 2005, a survey by the Japanese Ministry of Health and Welfare reported a prevalence of COPD of 0.2%–0.4% in Japan,Citation2 and there is a belief that COPD in Japan is different than in the West.Citation3 However, recent data indicate the prevalence of airflow limitation to be 10.9%, suggesting under-recognition of COPD in Japan,Citation3 and the phenotypic distribution is similar to that previously described in Western studies.Citation4 COPD is an inflammatory lung disease caused by long-term inhalation exposure to noxious substances, most commonly tobacco smoke.Citation1,Citation4 It is characterized by incompletely reversible airflow limitation, resulting from varying degrees of small airways disease and emphysema, and it is progressive.Citation1 Inflammation of the respiratory tract may be involved both in the development of COPD and the progression of airflow limitation.Citation1,Citation5 Indeed, Japanese and world guidelines define COPD as an inflammatory disease of the lung.Citation1,Citation2 However, increasing evidence suggests COPD symptoms are also associated with abnormal systemic inflammatory responses, making COPD a multicomponent disease.Citation6Citation9 Increased airway and systemic inflammation have been associated with COPD exacerbations and physiological changes such as hyperinflation.Citation10,Citation11

To this end, inflammatory biomarkers have become a useful means of characterizing the extent of both airway and systemic inflammation associated with COPD and for measuring the anti-inflammatory effects of treatment, especially given the high variability observed in the rate of decline of forced expiratory volume in 1 second (FEV1) as an endpoint.Citation12Citation14 However, serum and sputum inflammatory biomarkers in Japanese patients with COPD have not been studied extensively. In this study, we have selected a panel of biomarkers based on previous studies, such as ECLIPSE (Evaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints), in COPD. For example, in exacerbations of COPD, it has been reported that inflammatory cells such as neutrophils and eosinophils are increased in sputum.Citation11 Interleukin 8 (IL-8) in sputum is associated with increased chemotaxis of activated neutrophils and eosinophils.Citation15 Matrix metalloproteinase 9 (MMP-9) has also been related to the pathogenesis of COPD,Citation15 and clara cell secretory protein 16 (CC-16) in blood and pulmonary surfactant protein (SP) A and Krebs von den Lungen 6 (KL-6) have been postulated to represent a biomarker, especially for fibrosis of the lung.

The novelty and importance of the present study is that it is the first to assess both airway and systemic inflammation (using induced sputum and serum samples) in Japanese COPD patients compared with smoking and nonsmoking control participants. It proved impossible in this population to include a group of COPD patients without a smoking history.

Materials and methods

Subjects

A total of 100 Japanese men and women, aged 40 years or older, were enrolled who were either nonsmokers (nonsmoking control participants) who had not smoked for the past 6 months with a pack history of 1 pack a year or less and no diagnosis of COPD, current or ex-smokers (smoking control participants) with a pack history of 10 or more years and no diagnosis of COPD, or current or ex-smokers with a diagnosis of COPD (COPD patients) with a pack history of 10 or more years and cough or sputum 2 or fewer weeks before entry to the study. The diagnosis of COPD was defined according to the Global Initiative for Chronic Obstructive Lung Disease (GOLD) criteria, with a prebronchodilator FEV1/forced vital capacity (FVC) lower than 0.7.Citation1 Participants assigned as nonsmoking control patients and smoking control patients had no airway obstruction (prebronchodilator FEV1/FVC ≥0.7). Participants were excluded if they had a diagnosis of bronchial asthma or any respiratory disorder other than COPD; had undergone lung volume reduction or lung transplant; had a chest X-ray (or computed tomography [CT] scan) indicating major disease other than COPD that may interfere with study assessments; had a respiratory infection within 4 weeks before screening; used inhaled corticosteroids and systemic corticosteroids or low-dose xanthines (use at a regular dose was allowable) 2 or fewer weeks before screening; had a bacterial infection, viral infection (including viral hepatitis), or systemic inflammation at enrolment; or had a known α1-antitrypsin deficiency. All participants gave, in writing, their informed consent and permission to use their samples.

Study design

This was a non-treatment study conducted between November 2012 (first subject, first visit) and November 2013 (statistical analysis completed) at the Hiroshima University Hospital, Japan. The study was conducted in accordance with the ethical principles of the Declaration of Helsinki and was approved by the Ethics Committee of Hiroshima University Hospital.

Participants attended the study center once for an initial medical examination and study-related assessments. If participants did not reach their cough and sputum status, they could return to the site for a retest 2 or more weeks after the initial visit. Undesirable medical events, including serious adverse events, were monitored during and between both study visits.

Assessments

At the study visit, participants underwent standard lung function assessments (FEV1, FVC, % predicted FEV1 and FEV1/FVC)Citation16 and had a chest X-ray if one had not been performed within 6 months of the study visit. Impulse Oscillation System (IOS) parameters were measured, as previously described.Citation17,Citation18 Values at an airway resistance of 5 Hz (R5) and 20 Hz (R20) and airway reactance of 5 Hz (X5) were used for analysis. Airway resistance and reactance were assessed as measurements averaged over several tidal breaths (whole-breath analysis). Sputum specimens were obtained according to guidelines of the European Respiratory Society’s Task Force, as previously described.Citation19,Citation20 In brief, a sputum induction procedure was conducted using 3%–5% hypertonic saline given at 5 minute intervals for a maximum of 15 minutes via an ultrasonic nebulizer NE-U12 (Omron Co, Kyoto, Japan). Inflammatory cell counts (neutrophil, eosinophil, lymphocyte, macrophage, and total inflammatory cell) from induced sputum were measured using fixed cell samples by Diff-Quick staining in the Hiroshima University laboratory. Inflammatory biomarkers were measured using standard immunochemical assays by SRL Medisearch Inc (Sagamihara, Japan) collected from sputum supernatant; IL-8 [enzyme-linked immunosorbent assay {ELISA}], KL-6 [electro-chemiluminescence immunoassay], SP-A [enzyme immunoassay {EIA}], SP-D [EIA], hs-CRP [latex-enhanced nephelometry], or MMP-9 [EIA], serum; IL-8 [ELISA], KL-6 [enzyme-linked immune-culture assay], CC-16 [ELISA], SP-A [EIA], SP-D [EIA], hs-CRP [latex-enhanced nephelometry], and plasma; fibrinogen, measured using the thrombin clotting time test with a detection limit of 20 mg/dL. CC-16 was not measured in sputum, as previous studiesCitation14 showed it was not a descriptive factor in COPD.

Statistical methods

The target number of participants was 100, of whom 50 were COPD patients, 30 were smoking control participants, and 20 were nonsmoking control participants. Inflammatory cell counts, the percentage of inflammatory cells, and concentration of inflammatory biomarkers were analyzed using the Dunnett’s multiple comparison nonparametric test to compare the COPD, non-COPD smoker, and non-COPD nonsmoker groups. Stratified analyses were then conducted according to subject baseline factors. The per protocol (PP) set was the primary data set for analysis. Correlation between variables was assessed by Spearman’s rank correlation coefficient.

Results

Subject population

Demographic data are summarized in . The three subject groups were generally similar with respect to age, height, and weight; the smoking control participant group and COPD group were similar with respect to the number of pack-years. However, there was a predictable statistically significant difference between COPD patients versus both smoking control participants and nonsmoking control participants with respect to FEV1, FEV1 % predicted, and FEV1/FVC. There was also a statistically significant difference between COPD patients and both smoking control participants and nonsmoking control participants with respect to all the IOS parameters, with the exception of R20, a marker of mean airway resistance of central airways, for COPD patients versus smoking control participants.

Table 1 Summary of demographics and spirometry measurements

The PP population comprised 97 of 100 participants, as three participants from the COPD patient group were excluded from PP analysis because of concomitant illnesses of fibrosis of the lung, asthma, and fibrosis of the lung and kidney cancer, respectively. One participant experienced a serious adverse event (bronchial asthma attack) during the sputum induction procedure and was not enrolled in the study.

Inflammatory cells in sputum

Sputum inflammatory cell data are summarized in .

Table 2 Summary of inflammatory cells and biomarkers in sputum and blood

Although no statistically significant differences were observed in total cell counts between COPD patients and non-COPD control participants, there was a trend toward an increase in COPD. The mean sputum total cell count ± standard deviation in the COPD group, smoking control participants, and nonsmoking control participants were 66.5×105±80.8 cells/mL, 45.8×105±40.4 cells/mL and 39.5×105±30.4 cells/mL, respectively.

Similarly, no statistically significant differences were observed between the COPD patients and non-COPD control participants in relation to induced sputum neutrophils and macrophages. However, the mean percentage ± standard deviation of neutrophils in the COPD group, smoking control participants, and nonsmoking control participants were 76.3%±15.7%, 69.5%±20.3%, and 67.4%±24.3%, respectively, again indicating a trend toward an increase in COPD. In contrast, the absolute numbers of macrophages were 8.7×105, 9.5×105, and 12.3×105 cells/mL in the COPD patients, smoking control participants, and nonsmoking control participants, and the mean percentages were 16.6%±13.3%, 23.1%±18.0%, and 25.4%±23.7%, respectively, suggesting a marked decrease in COPD patients.

Levels of biomarkers in sputum and blood

Biomarker data in sputum and blood are summarized in .

Median sputum IL-8 levels were significantly higher in COPD patients (551.0 pg/mL) than in the smoking control participants (263.0 pg/mL) (P<0.05). Although overall, the levels of MMP-9, SP-A, SP-D, and KL-6 in sputum were not significantly different between COPD patients and the control groups, levels of SP-D and KL-6 tended to be higher in the COPD patients compared with both non-COPD control groups.

Mean levels of serum CC-16, hs-CRP, and KL-6 and plasma fibrinogen were also higher in COPD patients compared with the non-COPD control participants, with the lowest values generally seen in the nonsmoking group. However, none of the differences seen between the COPD patients and control groups were significantly different for any of the serum biomarkers.

Correlation between sputum inflammatory cells and biomarkers in sputum and serum in COPD patients

A summary of correlations in the COPD patients is given in and .

Table 3 Summary of correlation between sputum inflammatory cell counts and sputum biomarkers in patients with COPD

Table 4 Summary of correlation coefficient between sputum inflammatory cell counts and serum biomarkers in COPD patients

Significant correlations were observed between some inflammatory cells in sputum and biomarkers in sputum () in the COPD patients, with the most significant correlations seen between total cell count and MMP-9 (P<0.001), neutrophils and MMP-9 (P<0.001), and macrophages and KL-6 (P<0.01). Other significant correlations were observed between total cell count and IL-8 (P<0.05), neutrophils and IL-8 (P<0.05), and macrophages and MMP-9 (P<0.05) ().

Although not showing a clear association between lung and systemic biomarkers of inflammation, which has been reported previously, significant correlations were observed between some inflammatory cells in sputum and biomarkers in serum (), with the most significant correlations observed between serum CC-16 and both sputum total cell count (P<0.005) and neutrophils (P<0.005).

Discussion

COPD is a complex disease involving many different types of inflammatory and structural cells, with increasing evidence that both pulmonary and systemic inflammatory processes are involved.Citation4,Citation21 To add to the complexity, COPD is also a heterogeneous disease with interpatient variability.Citation22 Despite this complexity and heterogeneity, the overall trends and correlations observed in this study in levels of sputum inflammatory cells and sputum and serum biomarkers support the presence of pulmonary and systemic inflammation in Japanese COPD patients.

In induced sputum, the higher total cell count and neutrophil levels tended to be observed in the COPD patients compared with the non-COPD control participants; this agrees with previous dataCitation4 and provides evidence of pulmonary inflammatory responses occurring in the airways of Japanese COPD patients. However, there was a statistically significant difference between COPD patients and both smoking control participants and nonsmoking control participants with respect to FEV1, FEV1 % predicted, and FEV1/FVC, suggesting spirometry is essential in the diagnosis and assessment of severity of COPD. Indeed, studies have shown that sputum neutrophils have been directly correlated to the rate of annual decline in FEV1.Citation23 Furthermore, IOS parameters also differ between COPD patients and both smoking control participants and nonsmoking control participants. We note that IOS is a novel and noninvasive method of evaluating respiratory resistance and reactance.

Macrophages play a pivotal role in COPD pathophysiology. They are activated by cigarette smoke extract to release inflammatory mediators, thus providing a cellular mechanism by which smoking is linked with inflammation.Citation4 Marked increases in the number of macrophages in patients with COPD have been reported.Citation4,Citation24 However, in our study, macrophage levels in induced sputum tended to decrease in COPD patients compared with the non-COPD control participants, perhaps indicating that macrophages were being consumed by the inflammatory process. This concurs with several previous reports, where there was a decrease in the number of macrophages in COPD.Citation25,Citation26

The significantly higher levels of sputum IL-8 in COPD patients compared with in the smoking control group and its correlation with the numbers of neutrophils suggest IL-8 may be secreted into sputum and then acts as a chemoattractant for neutrophils migrating into the airway. Neutrophils are also a major source of IL-8.Citation4,Citation27 Similarly, there is a strong correlation between MMP-9 and the number of neutrophils and neutrophils are known to secrete MMP-9.

Higher levels of sputum SP-D, a lung type II pneumocyte-derived protein, were also observed in COPD patients compared with non-COPD control participants, which is in agreement with previous data in Japanese subjects.Citation19 SP-D has been implicated in the regulation of inflammation and pulmonary host defense and was shown to be elevated in COPD patients compared with non-COPD control participants with prolonged cough; in addition, SP-D levels are positively correlated with smoking status.Citation19

Serum CC-16 levels generally increase after acute exposure to smoke; however, there are conflicting data with regard to serum levels of CC-16 in COPD.Citation13 For example, in the ECLIPSE cohort, serum CC16 levels were decreased in COPD relative to non-COPD control participants, and demographic variability across the study groups was implicated as a potential contributing factor.Citation13 Serum CRP is also considered one of the signature biomarkers of systemic inflammation.Citation6,Citation28 Although levels are variable, serum hs-CRP tends to be higher in COPD patients compared with in healthy smokers or nonsmokers.Citation29 However, as with CC-16, results can be conflicting and may be a result of variable patient selection and differences in the classification of COPD across studies.Citation6

As for sputum, serum KL-6 levels also tended to be higher in the COPD patient group than in the non-COPD control participants. Circulating levels of KL-6 are considered to be a sensitive marker for fibrotic lung diseases and are used in clinical practice in Japan, alongside SP-A and SP-D, for the identification of idiopathic pulmonary fibrosis and other types of interstitial lung diseases.Citation19,Citation30 The increase in KL-6 in COPD in this study is of interest.

Although this study provides evidence that there is both pulmonary and systemic inflammation in Japanese COPD patients, our results were unable to provide robust evidence of any significant relationships between the pulmonary and systemic components, which agrees with other data.Citation6,Citation15,Citation31,Citation32 The main strength of our study was that the COPD patients enrolled represented a typical Japanese patient population. Our study also had limitations, including the small sample size, which restricted the statistical power of the findings. In addition, not all participants had a CT scan during enrolment. In fact, we were able to obtain the CT scan results of only 10/50 COPD patients, and therefore some of the COPD patients included in the analysis may have had undetected lung fibrosis that may have affected biomarker concentrations. Furthermore, the validation and definition of the specificity of these proteins in various phenotypes of COPD (bronchitis vs emphysema predominance) will require further study. Variability in biomarker levels, as seen in our study, especially in induced sputum, and even in stable COPD patients, is well-documented and can confound results.Citation15,Citation33 Finally, measuring biomarker levels in induced sputum rather than biopsy samples means the results are more likely to reflect inflammation in the central, rather than lower and peripheral, airways.Citation19

Conclusion

This is the first study to investigate both airway and systemic inflammation in COPD in Japanese subjects compared with the respective smoking and nonsmoking control groups. Our results provide further evidence that COPD in Japanese patients is a multicomponent disease, involving both airway and systemic inflammation, in addition to airway obstruction.

These results suggest that intervention with anti- inflammatory therapy may provide additional benefit for disease management of COPD in Japan. This now needs to be considered in a controlled setting.

Author contributions

All authors were involved in the acquisition and analysis of data, drafting and critical revision of the manuscript, and the final approval of the proof to be published.

Acknowledgments

Funding for this study was provided by GlaxoSmithKline (NCT0356642). All listed authors meet the criteria for authorship set forth by the International Committee for Medical Journal Editors. The authors wish to acknowledge the following individuals for their contributions: members of Hiroshima University who participated in subject recruitment: Hiroshi Murai, Yoshinori Haruta, Kazunori Fujitaka, Shinichiro Ohshimo, Hiroshi Iwamoto, and Taku Nakashima. The authors would also like to thank the following members of the GlaxoSmithKline team who developed the study protocol: Etsuko Hayashi, Hideo Kikkawa, and Daisuke Yoshimoto; SRL Medisearch Inc, for performing the biomarker analyses; and the Institute of Japanese Union of Scientists and Engineers, for performing the data management and statistical analysis.

Disclosure

AK and TH were employees of GlaxoSmithKline at the time of the study. NK discloses that his university has received research grants from the commercial entity that sponsored the study. NH discloses having received honoraria, and his university has received research grants from the commercial entity that sponsored the study. NI discloses having received honoraria/consulting fees from GlaxoSmithKline for participating on the advisory board meeting, and his university has received research grants from the commercial entity that sponsored the study. MJ is an independent respiratory consultant employed by GlaxoSmithKline to help in the design and conduct of the study and is a shareholder in GlaxoSmithKline, but has no other relationships or activities that could appear to have influenced the submitted work. Medical writing and editorial support in the form of development of draft outline, development of manuscript (all drafts), assembling tables, and collating author comments was provided by Dr Kathryn White of Cathean Ltd, and was funded by GlaxoSmithKline.

References

  • Global Initiative for Chronic Obstructive Lung DiseaseGlobal Strategy for the Diagnosis, Management, and Prevention of Chronic Obstructive Pulmonary DiseaseGlobal Initiative for Chronic Obstructive Lung Disease Available from: http://www.goldcopd.com/uploads/users/files/GOLD_Report_2014_Oct30.pdfAccessed December 15, 2014
  • Japanese Respiratory SocietyGuidelines for the Diagnosis and Treatment of COPD4th editionJapanese Respiratory Society2013 Available from: https://www.jrs.or.jp/modules/guidelines/index.php?content_id=1Accessed April 19, 2013
  • FukuchiYNishimuraMIchinoseMCOPD in Japan: the Nippon COPD Epidemiology studyRespirology20049445846515612956
  • BarnesPJMediators of chronic obstructive pulmonary diseasePharmacol Rev200456451554815602009
  • HoggJCChuFUtokaparchSThe nature of small-airway obstruction in chronic obstructive pulmonary diseaseN Engl J Med2004350262645265315215480
  • MacNeeWSystemic inflammatory biomarkers and co-morbidities of chronic obstructive pulmonary diseaseAnn Med201345329130023110517
  • HuertasAPalangePCOPD: a multifactorial systemic diseaseTher Adv Respir Dis20115321722421429981
  • AgustíASystemic effects of chronic obstructive pulmonary disease: what we know and what we don’t know (but should)Proc Am Thorac Soc20074752252517878464
  • van EedenSFSinDDChronic obstructive pulmonary disease: a chronic systemic inflammatory diseaseRespiration200875222423818042978
  • WedzichaJASeemungalTACOPD exacerbations: defining their cause and preventionLancet2007370958978679617765528
  • FujimotoKYasuoMUrushibataKHanaokaMKoizumiTKuboKAirway inflammation during stable and acutely exacerbated chronic obstructive pulmonary diseaseEur Respir J200525464064615802337
  • BarnesNCQiuY-SPavordIDSCO30005 Study GroupAntiinflammatory effects of salmeterol/fluticasone propionate in chronic obstructive lung diseaseAm J Respir Crit Care Med2006173773674316424444
  • LomasDASilvermanEKEdwardsLDMillerBECoxsonHORTal-SingerREvaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints (ECLIPSE) investigatorsEvaluation of serum CC-16 as a biomarker for COPD in the ECLIPSE cohortThorax200863121058106318757456
  • VestboJEdwardsLDScanlonPDECLIPSE InvestigatorsChanges in forced expiratory volume in 1 second over time in COPDN Engl J Med2011365131184119221991892
  • DickensJAMillerBEEdwardsLDSilvermanEKLomasDATal-SingerREvaluation of COPD Longitudinally to Identify Surrogate Endpoints (ECLIPSE) study investigatorsCOPD association and repeatability of blood biomarkers in the ECLIPSE cohortRespir Res201112114615522054035
  • MillerMRHankinsonJBrusascoVATS/ERS Task ForceStandardisation of spirometryEur Respir J200526231933816055882
  • SugiyamaAHattoriNHarutaYCharacteristics of inspiratory and expiratory reactance in interstitial lung diseaseRespir Med2013107687588223582576
  • NakagawaMHattoriNHarutaYEffect of increasing respiratory rate on airway resistance and reactance in COPD patientsRespirology Epub2014923
  • IshikawaNHattoriNTanakaSLevels of surfactant proteins A and D and KL-6 are elevated in the induced sputum of chronic obstructive pulmonary disease patients: a sequential sputum analysisRespiration2011821101821474912
  • PaggiaroPLChanezPHolzOSputum inductionEur Respir J Suppl2002373s8s12361361
  • BarnesNReducing inflammation in COPD: the evidence buildsThorax2007621192792817965073
  • AgustiACalverleyPMACelliBEvaluation of COPD Longitudinally to Identify Predictive Surrogate Endpoints (ECLIPSE) investigatorsCharacterisation of COPD heterogeneity in the ECLIPSE cohortRespir Res201011112213520831787
  • StănescuDSannaAVeriterCAirways obstruction, chronic expectoration, and rapid decline of FEV1 in smokers are associated with increased levels of sputum neutrophilsThorax19965132672718779129
  • ToungoussovaOMiglioriGBFoschino BarbaroMPChanges in sputum composition during 15 min of sputum induction in healthy subjects and patients with asthma and chronic obstructive pulmonary diseaseRespir Med200710171543154817258444
  • IwamotoHGaoJKoskelaJDifferences in plasma and sputum biomarkers between COPD and COPD-asthma overlapEur Respir J201443242142923794464
  • MakrisDVrekoussisTIzoldiMIncreased apoptosis of neutrophils in induced sputum of COPD patientsRespir Med200910381130113519329291
  • BazzoniFCassatellaMARossiFCeskaMDewaldBBaggioliniMPhagocytosing neutrophils produce and release high amounts of the neutrophil-activating peptide 1/interleukin 8J Exp Med199117337717741997655
  • Pinto-PlataVMMüllerovaHTosoJFC-reactive protein in patients with COPD, control smokers and non-smokersThorax200611232816143583
  • van DurmeYMVerhammeKMAarnoudseAJC-reactive protein levels, haplotypes, and the risk of incident chronic obstructive pulmonary diseaseAm J Respir Crit Care Med2009179537538219096002
  • IshikawaNHattoriNYokoyamaAKohnoNUtility of KL-6/MUC1 in the clinical management of interstitial lung diseasesRespir Investig2012501313
  • BizetoLMazzoliniABRibeiroMStelmachRCukierANunesMPTInterrelationship between serum and sputum inflammatory mediators in chronic obstructive pulmonary diseaseBraz J Med Biol Res200841319319818327434
  • RöpckeSHolzOLauerGRepeatability of and relationship between potential COPD biomarkers in bronchoalveolar lavage, bronchial biopsies, serum, and induced sputumPLoS ONE2012710e4620723056262
  • DuvoixADickensJHaqIBlood fibrinogen as a biomarker of chronic obstructive pulmonary diseaseThorax201368767067622744884