Mechanisms of allergy and clinical immunology
Identification of a distinct glucocorticosteroid-insensitive pulmonary macrophage phenotype in patients with chronic obstructive pulmonary disease

https://doi.org/10.1016/j.jaci.2013.08.044Get rights and content

Background

In patients with chronic obstructive pulmonary disease (COPD), pulmonary macrophages increase in number, release increased levels of inflammatory mediators, and respond poorly to glucocorticosteroids. Whether this is due to a change in macrophage phenotype or localized activation is unknown.

Objective

We sought to investigate whether macrophages from patients with COPD are a distinct phenotype.

Methods

Macrophage populations were isolated from human lung tissue from nonsmokers, smokers, and patients with COPD by using Percoll density gradients. Five macrophage populations were isolated on the basis of density (1.011-1.023, 1.023-1.036, 1.036-1.048, 1.048-1.061, and 1.061-1.073 g/mL), and cell-surface expression of CD14, CD16, CD163, CD40, and CD206 was assessed by using flow cytometry. Release of active matrix metalloproteinase 9, TNF-α, CXCL8, and IL-10 was measured by using ELISA.

Results

The 2 least dense fractions were more than 90% apoptotic/necrotic, with the remaining fractions greater than 70% viable. Macrophages from nonsmokers and smokers were CD163+, CD206+, CD14+, and CD40, whereas macrophages from patients with COPD were less defined, showing significantly lower expression of all receptors. There were no differences in receptor expression associated with density. Macrophages from patients with COPD of a density of 1.036 to 1.048 g/mL released higher levels of active matrix metalloproteinase 9 compared with cells from nonsmokers, with no difference between the remaining fractions. This population of macrophages from patients with COPD was less responsive to budesonide compared with those from nonsmokers and smokers when stimulated with LPS. Glucocorticosteroid insensitivity was selective for proinflammatory cytokines because budesonide inhibition of LPS-stimulated IL-10 release was similar for all macrophages.

Conclusions

This study identifies a specific macrophage phenotype in the lungs of patients with COPD who are glucocorticosteroid insensitive with a density of 1.036 to 1.048 g/mL but do not correspond to the current concept of macrophage phenotypes.

Section snippets

Subject selection

Lung tissue surplus to diagnostic requirements was obtained from pulmonary resections at the Royal Brompton and Harefield NHS Foundation Trust. Smokers had a smoking history of at least 10 pack years, and patients with COPD were stable and fulfilled the American Thoracic Society criteria.27 All subjects provided written informed consent, as approved by the London-Chelsea Research Ethics Committee. There were significant differences between FEV1 in liters, FEV1 percent predicted, and FEV1/forced

Results

Approximately 90% of cells extracted from the 10% to 20% (vol/vol) and 20% to 30% (vol/vol) Percoll interfaces were nonviable, resulting in their exclusion from further analysis (Table II). Cell viability increased with increasing cell density, with greater than 70% of cells viable in the remaining cell fractions (Table II). Cells from the 3 remaining viable fractions had clearly defined nuclei and cytoplasm reminiscent of macrophages with no clear morphological differences (see Fig E2 in this

Discussion

Macrophages play a pivotal role in the pathophysiology of COPD, in which alveolar macrophages increase in number, release more inflammatory mediators (eg, CXCL8, TNF-α, and MMP-9), and respond less well to glucocorticosteroids.6, 32 This study investigated the possibility that these macrophages might represent a distinct phenotype associated with COPD and showed that within the pulmonary macrophage population, approximately a third of these cells are glucocorticosteroid insensitive and release

References (51)

  • S.V. Culpitt et al.

    Sputum matrix metalloproteases: comparison between chronic obstructive pulmonary disease and asthma

    Respir Med

    (2005)
  • A. Sica et al.

    Tumour-associated macrophages are a distinct M2 polarised population promoting tumour progression: potential targets of anti-cancer therapy

    Eur J Cancer

    (2006)
  • P.J. Barnes

    Macrophages as orchestrators of COPD

    COPD

    (2004)
  • M. Saetta et al.

    Inflammatory cells in the bronchial glands of smokers with chronic bronchitis

    Am J Respir Crit Care Med

    (1997)
  • J.C. Hogg et al.

    The nature of small-airway obstruction in chronic obstructive pulmonary disease

    N Engl J Med

    (2004)
  • S.V. Culpitt et al.

    Effect of high dose inhaled steroid on cells, cytokines, and proteases in induced sputum in chronic obstructive pulmonary disease

    Am J Respir Crit Care Med

    (1999)
  • S.V. Culpitt et al.

    Impaired inhibition by dexamethasone of cytokine release by alveolar macrophages from patients with chronic obstructive pulmonary disease

    Am J Respir Crit Care Med

    (2003)
  • K. Ito et al.

    Cigarette smoking reduces histone deacetylase 2 expression, enhances cytokine expression, and inhibits glucocorticoid actions in alveolar macrophages

    FASEB J

    (2001)
  • S. Gordon

    Alternative activation of macrophages

    Nat Rev Immunol

    (2003)
  • S. Gordon et al.

    Monocyte and macrophage heterogeneity

    Nat Rev Immunol

    (2005)
  • W.S. Walker

    Functional heterogeneity of macrophages: subclasses of peritoneal macrophages with different antigen-binding activities and immune complex receptors

    Immunology

    (1974)
  • T. Mokoena et al.

    Human macrophage activation. Modulation of mannosyl, fucosyl receptor activity in vitro by lymphokines, gamma and alpha interferons, and dexamethasone

    J Clin Invest

    (1985)
  • R. Shaykhiev et al.

    Smoking-dependent reprogramming of alveolar macrophage polarization: implication for pathogenesis of chronic obstructive pulmonary disease

    J Immunol

    (2009)
  • L.I. Kunz et al.

    Smoking status and anti-inflammatory macrophages in bronchoalveolar lavage and induced sputum in COPD

    Respir Res

    (2011)
  • A. Holian et al.

    Separation of bronchoalveolar cells from the guinea pig on continuous gradients of Percoll: functional properties of fractionated lung macrophages

    J Reticuloendothel Soc

    (1983)
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    Supported by a studentship from the BBSRC, United Kingdom. Also supported by the National Institute of Health Research Respiratory Disease Biomedical Research Unit at the Royal Brompton Hospital and Harefield Foundation NHS Trust and Imperial College London.

    Disclosure of potential conflict of interest: P. J. Barnes has been supported by one or more grants from GlaxoSmithKline and AstraZeneca; is a Board member for Boehringer Ingelheim and Pfizer, has consultancy arrangements with Glenmark and Sun Pharma; has provided expert testimony for Boehringer Ingelheim and TEVA; has received one or more grants from or has one or more grants pending with AstraZeneca, Nycomed/Takeda, Novartis, Boehringer Ingelheim, Chiesi, Aquinox, and Pfizer; and has received one or more payments for lecturing from or is on the speakers' bureau for AstraZeneca, Nycomed, Chiesi, Novartis, and Pfizer. L. E. Donnelly has been supported by one or more grants from BBSRC, Pfizer, MRC, Boehringer Ingelheim, Nycomed, and AstraZeneca. The rest of the authors declare that they have no relevant conflicts of interest.

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