Chronic obstructive pulmonary disease (COPD) and asthma represent a significant challenge to the respiratory physician in their management and diagnosis, with significant overlap in their clinical features, physiological parameters and airway inflammation [¹, ²]. COPD is characterised by chronic irreversible airflow obstruction, structural changes such as small airway destruction, fibrosis and neutrophilic airway inflammation [³, ⁴, ⁵, ⁶]. However, up to 40% of COPD subjects display eosinophilic airway inflammation [⁷] which is associated with elevated levels of interleukin (IL)-5. Treatment with corticosteroid therapy reduces eosinophilic inflammation and IL-5 levels [⁸]. In contrast, asthma is characterised by variable airflow obstruction, airway hyper-responsiveness, a thickened basement membrane and eosinophilic airway inflammation [⁴]. Both non-eosinophilic and neutrophilic asthma have been described and these are hypothesised to confer steroid resistance [⁹, ¹⁰]. The use of induced sputum has led to further insights into the pulmonary inflammation of asthma and COPD [¹¹, ¹²], allowing characterisations of pro-inflammatory cytokines and chemokines. Importantly, COPD and asthma have often been studied independently using limited mediator panels [⁸, ¹³]. In light of the increasing recognition of the heterogeneity of asthma and COPD and their potential overlap [², ¹⁴], it is important to investigate whether there are different pulmonary mediator responses and whether airway inflammatory phenotypes confer similar cytokine and chemokine expression in both asthma and COPD.

In this study we sought to investigate the sputum cytokine and chemokine pattern in subjects with asthma and COPD using a novel sputum processing method and to compare the inflammatory response within and between diseases and in relation to sputum phenotypes.

One hundred and three subjects were recruited (asthma, n = 54; COPD, n = 49). The baseline demographics of all subjects are presented in table 1. Asthma severity showed that 13, 24, 30 and 33% of subjects were GINA I, II, III and IV/V. The GOLD classification for COPD severity identified 19, 33, 33 and 15% as having GOLD I, II, III and IV. The mean (SEM) post-bronchodilator FEV₁ % predicted for the asthma and COPD subjects was 87% (4) and 54% (4), respectively. Sputum mediator analysis using the MSD and Luminex platforms was available in 37 asthmatic and 23 COPD subjects. The clinical characteristics of these subjects with available sputum mediator analysis are presented in table 2. Analytes with increased recovery in D-PBS-treated sputum compared to DTT-treated sputum were IL-1β, 4, 5, 10, 13, IFN-γ, TNFRI, GM-CSF, CCL2, 3, 4, 5, 13 and 17 (online suppl. fig. 1; for all online supplementary material, see www.karger.com/doi/10.1159/000330667). The mediators that were measured in the sputum and their lower and upper quantification limits are listed in online supplementary table 1.

Correlations of all measured sputum markers are presented as a heat map illustration in figure 1. Sputum markers that are closely correlated are identified as intense bright colours (white: r = 1.0). There was a strong correlation identified between TNFRI, TNFRII and IL-6R (r = 0.93, r = 0.90 and r = 0.90); MMP8, MMP9 and IL-1β (r = 0.94, r = 0.91 and r = 0.87), and CXCL10 with CXCL11 (r = 0.92).

In this study we have shown that sputum cellular patterns of airway inflammation are distinct across disease severity in asthma and COPD. However, sputum cell profiles are similar between severe asthma and COPD. Likewise, the sputum mediator profiles are largely similar between asthma and COPD, with a few mediators increased in COPD. Importantly, differential mediator expression is more marked between eosinophilic and non-eosinophilic sub-phenotypes, which is independent of disease.

Mediator expression between COPD and asthma has confirmed elevated levels of the pro-inflammatory mediators IL-6 and IL-8 in COPD [¹⁹, ²⁰], whilst there is increased expression of Th2 cytokines such as IL-4, IL-5 and IL-13 in asthma and allergic disease [²¹, ²²]. Bronchial biopsies studies have shown predominately neutrophilic inflammation in COPD and eosinophilic inflammation in asthma [⁵, ²³]. However an overlap in cellular inflammation exists [²⁴]; indeed we have shown a similar overlap in airway cellular inflammation with little difference in airway inflammation and in particular airway inflammatory phenotypes between moderate-to-severe asthma and COPD. It has been recognised that in persistent severe asthma there is often neutrophilic airway inflammation with elevated levels of IL-8 [²⁵] and this type of inflammation confers steroid non-responsiveness [¹⁰, ²⁶], albeit this maybe confounded itself by the effects of inhaled and oral corticosteroids [²⁷], which have been shown to alter mediator expression [²⁸]. Here we have shown that the neutrophilic airway inflammation that is present in severe asthma is similar to the degree of neutrophilic inflammation that exists in COPD. The identification of airway inflammatory phenotypes has been effectively used to reduce exacerbations in asthma and COPD [²⁹, ³⁰], whilst cluster analysis has determined airway inflammatory phenotypes within clinical phenotypes of asthma [¹⁴].

We have also validated a sputum processing method that improves the recovery of previously difficult to measure cytokines [¹³]. The use of multiplex platforms has provided the opportunity to assess inflammatory mediators in small volumes. In our study the initial incubation of selected sputum plugs with D-PBS followed by standard sputum processing techniques improved the recovery of cytokines including previously difficult to measure Th2 cytokines [⁸] and improved the recovery of several cytokines satisfying the European Respiratory Guidelines on mediator recovery [¹⁷].

In our study we have shown that the chemokine and cytokine expressions in asthma and COPD are broadly similar, with a few mediators increased in COPD. Indeed the differential expression of mediators between eosinophilic and non-eosinophilic airway inflammation is more marked and we could not demonstrate any differential expression between asthma and COPD after stratification for inflammatory airway cellular phenotypes. This mediator expression between airway inflammatory phenotypes has been demonstrated separately in COPD [⁸] and asthma [¹⁰]. Although it has been widely perceived that asthma and COPD are distinct entities [³¹], the observation of similarities in their clinical phenotypes [²], and airway inflammometry [¹], increasingly suggests that one needs to view these airway inflammatory diseases together as a spectrum of disease with similar clinical symptoms, airway inflammation and physiology [³², ³³]. A phenotypic approach can propagate the development of targeted treatment strategies including the development of monoclonal antibody therapies [³⁴] and further the understanding in the pathogenesis of the disease [¹¹, ¹², ³⁵, ³⁶].

A limitation in this study is that we used subjects with asthma and COPD and did not compare the mediator response with relation to healthy controls, and in particular smoking and non-smoking controls. However we have been able to investigate asthma and COPD across all severities and applied our analysis to a widely observed clinical spectrum of disease; smoking healthy controls have previously demonstrated mediator expression and airway inflammation similar to subjects with COPD [²³]. In this study we did not fully characterise the airway disease phenotype using computed tomography or gas diffusion, and this and the effect of treatment could affect the inflammatory profiles seen. Serum and bronchoalveolar lavage cytokine expression with radiological characterisation of COPD have previously shown that mediator association is independent of lung function [³⁷] and Th2 cytokine expression is higher in subjects with radiological emphysema [³⁸, ³⁹]. Thus, it is interesting to note that similar mediator expression was seen within inflammatory groups in different disease expressions.

Although we were unable to do so in this study, inspection of mediators that are released, expressed on the cell surface or act as a surrogate measure of cell activity would provide further characterisation of eosinophilic and neutrophilic airway inflammatory patterns in COPD and asthma above that already shown using sputum mediator analysis in this study.

In conclusion, we have described and validated a novel sputum processing method which improves recovery and detection of several cytokines and chemokines, including that of previously difficult to measure Th2 cytokines. We have demonstrated that there are limited differences in the expression of sputum cytokines and chemokines between asthma and COPD, and there is greater differential expression between eosinophilic and non-eosinophilic sub-phenotypes, which may be independent of disease. Whether these airway mediator phenotypes confer a particular pathogenesis and clinical phenotype, and subsequent responses to treatment, needs to be further investigated in larger cohorts of subjects with airway disease.

This study was funded by AstraZeneca, the European Regional Development Fund (ERDF 05567) and a grant from a Wellcome Senior Clinical Fellowship (C.E.B.).

M.B., S.S., S.M., B.H., M.S., C.R., V.M. and D.P. have no financial interests to disclose. M.M., M.J., A.L., P.D., P.R. and P.N. are/were employees of AstraZeneca at the time of data collection and analysis. C.E.B. has received grant support and consultancy fees from AstraZeneca, MedImmune, Roche and GlaxoSmithKline.

The authors would like to thank all of the volunteers that took part in this study and the following people for their assistance in patient characterisation and laboratory support: M. Bourne, H. Pateman, A. Singapuri, S. Terry, M. Pancholi, and W. Monteiro.