Chest
Volume 136, Issue 4, October 2009, Pages 1047-1054
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Original Research
COPD
Leukotriene B4 Contributes to Exhaled Breath Condensate and Sputum Neutrophil Chemotaxis in COPD

https://doi.org/10.1378/chest.08-2782Get rights and content

Background

Neutrophils have been implicated in the pathogenesis of COPD. Several chemoattractants for neutrophils have been measured in samples of exhaled breath condensate (EBC) and induced sputum (IS) from patients with COPD. The aims of this study were to compare EBC and IS supernatant neutrophil chemotactic activity (NCA) from ex-smoking subjects with COPD and healthy ex-smokers, and to assess the contribution of leukotriene B4 (LTB4) to this activity.

Methods

Thirty-four subjects with COPD were compared to 24 control subjects. EBC and IS chemotactic activity for neutrophils was assessed by using Boyden microchambers. The chemotactic index was used to evaluate cell migration. LTB4 was measured by a specific enzyme immunoassay. The contribution of LTB4 to EBC and sputum neutrophil chemotaxis was assessed by an LTB4 receptor antagonist (U-75302; Cayman Chemical Company; Ann Arbor, MI).

Results

EBC and IS samples from both COPD patients and healthy subjects displayed significant NCA, but this activity was raised in COPD patients compared to healthy subjects. The chemotactic activity contained in sputum, however, failed to correlate with that in EBC. In COPD patients, there was a significant correlation between EBC NCA and sputum neutrophil counts. LTB4 levels were raised in EBC samples, but not in sputum samples, from COPD subjects compared to those from healthy subjects. LTB4 receptor antagonist concentrations (2.5 × 10−4 mol/L) reduced by 44.6% and by 44.4%, respectively, the chemotactic activity contained in the EBC and sputum samples.

Conclusions

EBC and IS from COPD patients have a raised NCA to which LTB4 contributes.

Section snippets

Study Design and Subject Characteristics

We recruited 34 stable COPD patients and 24 nonatopic healthy control subjects. Table 1 lists their demographic, functional, and treatment characteristics. All of the COPD patients fulfilled the criteria proposed by the Global Initiative for Chronic Obstructive Lung Disease (GOLD) guidelines17 and were ex-smokers. There were 20 subjects with moderate COPD (GOLD stage 2), 13 subjects with severe COPD (GOLD stage 3), and 1 subject with very severe COPD (GOLD stage 4). All COPD patients were in

Airway Neutrophilic Inflammation in COPD

Table 2 lists the characteristics of the sputum cell counts. COPD was clearly characterized by an intense neutrophilic inflammation, as shown by the raised neutrophil counts in the IS sample.

Effect of EBC and Sputum Dilution on NCA

The optimal concentration of EBC and sputum supernatant that was consecutively used in further chemotaxis assays was first determined by performing serial dilutions (1:1, 1:5, 1:10, and 1:50) of the original sample from 14 subjects with COPD and 7 healthy subjects. Both EBC and IS samples contained

Discussion

In this study, we have clearly shown that samples of EBC, as well as of IS, from ex-smoking COPD patients contained raised NCA compared to those of healthy ex-smokers. The EBC NCA was correlated to sputum neutrophil count. Furthermore, by using a specific LTB4 receptor antagonist, LTB4 was found to contribute significantly to the NCA contained in both EBC and sputum.

Both EBC and IS from COPD patients contain active soluble chemoattractants for neutrophils, as demonstrated by the significant

Acknowledgments

Author contributions: Dr. Corhay conducted and realized this study, and wrote the manuscript. Dr. Louis revised the manuscript. Ms. Henket and Ms. Sele conducted all of the in vitro analyses. Drs. Duysinx and Nguyen recruited COPD patients and helped perform the study.

Financial/nonfinancial disclosures: The authors have reported to the ACCP that no significant conflicts of interest exist with any companies/organizations whose products or services may be discussed in this article.

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Funding/Support: This study was funded by grants from the Fonds d'investissement de recherche scientifique du Centre Hospitalier Universitaire de Liège and from the Belgian Interuniversity Attraction Poles (IAP P6/38).

Reproduction of this article is prohibited without written permission from the American College of Chest Physicians (www.chestjournal.org/site/misc/reprints.xhtml).

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