SummaryIt has been reported that alveolar macrophages from patients with chronic obstructive pulmonary disease (COPD) display glucocorticoid (Gc) resistance. The Gc sensitivity of inflammatory mediators released by COPD macrophages may vary. The objective of this study was to identify Gc-insensitive inflammatory mediators produced by lipopolysaccharide (LPS)-stimulated alveolar macrophages from COPD patients. LPS-stimulated alveolar macrophages from 15 COPD patients, nine smokers (S) and nine healthy nonsmokers (HNS) were stimulated with LPS with or without dexamethasone (100 and 1000 nM). Luminex and enzyme-linked immunosorbent assay were used to measure 23 inflammatory mediators. After LPS stimulation there were lower levels of inflammatory mediators in COPD patients and S compared to HNS. There was no difference between groups for the effects of dexamethasone at either concentration (P > 0·05 for all comparisons). Tumour necrosis factor (TNF)-a, interleukin (IL)-6 and growth-related oncogene (GRO)-a displayed the greatest sensitivity to dexamethasone in COPD patients, while IL-8, granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) were the least sensitive. COPD macrophages have a reduced response to LPS. Gc sensitivity was similar in COPD macrophages compared to controls. We identify some Gc-insensitive cytokines, including GM-CSF, G-CSF and IL-8, that may be involved in the progression of airway inflammation in COPD patients.

Corticosteroids partially suppress cytokine production by chronic obstructive pulmonary disease (COPD) alveolar macrophages. p38 mitogen-activated protein kinase (MAPK) inhibitors are a novel class of anti-inflammatory drug. We have studied the effects of combined treatment with a corticosteroid and a p38 MAPK inhibitor on cytokine production by COPD alveolar macrophages, with the aim of investigating dose-sparing and efficacy-enhancing effects. Alveolar macrophages from 10 patients with COPD, six smokers, and six nonsmokers were stimulated with lipopolysaccharide (LPS) after preincubation with five concentrations of dexamethasone alone, five concentrations of the p38 MAPK inhibitor 1-(5-tert-butyl-2-p-tolyl-2H-pyrazol-3-yl)-3(4-(2-morpholin-4-ylethoxy)naphthalen-1-yl)urea (BIRB-796) alone, and all combinations of these concentrations. After 24 h, the supernatants were analyzed for interleukin (IL)-8, IL-6, tumor necrosis factor ␣ (TNF␣), granulocyte macrophage-colony-stimulating factor (GM-CSF), IL-1␣, IL-1␤, IL-1ra, IL-10, monocyte chemoattractant protein 3, macrophage-derived chemokine (MDC), and regulated on activation normal T cell expressed and secreted (RANTES). The effect of dexamethasone on p38 MAPK activation was analyzed by Western blotting. Dexamethasone and BIRB-796 both reduced LPSinduced cytokine production in a dose-dependent manner in all subject groups, with no differences between groups. Increasing the concentration of BIRB-796 in combination with dexamethasone produced progressively greater inhibition of cytokine production than dexamethasone alone. There were significant efficacyenhancing benefits and synergistic dose-sparing effects (p Ͻ 0.05) for the combination treatment for IL-8, IL-6, TNF␣, GM-CSF, IL-1ra, IL-10, MDC, and RANTES in one or more subject groups. Dexamethasone had no effect on LPS-induced p38 MAPK activation. We conclude that p38 MAPK activation in alveolar macrophages is corticosteroid-insensitive. Combining a p38 MAPK inhibitor with a corticosteroid synergistically enhances the antiinflammatory effects on LPS-mediated cytokine production by alveolar macrophages from patients with COPD and controls.

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT• Neutrophilic airway inflammation is a feature of chronic obstructive pulmonary disease (COPD). Inhaled lipopolysaccharide (LPS) challenge in healthy non‐smokers has been used to assess the anti‐inflammatory effects of novel drugs on neutrophilic airway inflammation.WHAT THIS STUDY ADDS• We performed inhaled LPS challenge in smokers to study an inflammatory response in subjects who more closely resemble COPD patients. Inhaled LPS caused reproducible pulmonary inflammation in smokers, associated with changes in systemic biomarkers. Inhaled LPS appears to be a suitable model for studying exacerbations of COPD.AIMS Lipopolysaccharide (LPS) is a TLR4 agonist which activates NFκB dependent cytokine production. We investigated LPS inhalation in healthy smokers as a model of COPD bacterial exacerbations. We studied safety, reproducibility, the translocation of the NFκB subunit p65 in sputum cells and changes in systemic biomarkers of inflammation.METHODS Twelve smokers inhaled 5 and 30 µg LPS and safety was monitored over 24 h. IL‐6, CRP, CCl‐18, SP‐D, CC‐16 and β‐defensin 2 were measured in serum samples collected at baseline, 4, 8 and 24 h. Sputum was induced at baseline, 6 and 24 h for cell counts and p65 expression. Repeated challenges were performed after a 2 week interval in 10 smokers.RESULTS LPS inhalation was well tolerated. Significant increases occurred in sputum neutrophil counts with both doses, with a maximum increase of 21.5% at 6 h after 30 µg which was reproducible, ri (intraclass correlation coefficient) = 0.88. LPS increased sputum cell nuclear p65 translocation and phospho‐p65 expression. All of the serum biomarkers increased following challenge but with different temporal patterns.DISCUSSION Inhaled LPS challenge in smokers causes pulmonary and systemic inflammation that involves NFκB activation. This appears to be a suitable model for studying bacterial exacerbations of COPD.