Azithromycin Reduces Exaggerated Cytokine Production by M1 Alveolar Macrophages in Cystic Fibrosis (original) (raw)
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Journal of Cystic Fibrosis, 2009
Background: Beneficial effects of azithromycin in cystic fibrosis (CF) have been reported, however, its mechanism of action remains unclear. The present study aimed at investigating the effect of azithromycin on CF airway epithelial cells. Methods: Primary cultures of purified tracheal epithelial cells from F508del and normal homozygous mice were established. Responses to lipopolysaccharide from Pseudomonas aeruginosa (LPS, 0.1 µg/ml) on mRNA expression of neutrophil-related chemokines, pro-and antiinflammatory cytokines were investigated in the presence or the absence of azithromycin (1 µg/ml). Results: CF airway epithelial cells showed upregulation of MIP-2 and KC responses to LPS, and azithromycin failed to downregulate these responses. In contrast, in CF cells, azithromycin increased KC and TNF-α expression under non-stimulated and LPS-stimulated conditions, respectively. In non-CF cells, the macrolide potentiated the LPS response on MIP-2 and on IL-10. Conclusions: Airway epithelial cells contribute to the dysregulated immune processes in CF. Azithromycin rather stimulates cytokine expression in CF airway epithelial cells.
Azithromycin fails to reduce inflammation in cystic fibrosis airway epithelial cells
European Journal of Pharmacology, 2012
Cystic fibrosis is a hereditary disease caused by a mutation in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene that encodes a chloride (Cl-) channel. Cystic fibrosis pulmonary pathophysiology is characterised by chronic inflammation and bacterial infections. Azithromycin, a macrolide antibiotic, has shown promising anti-inflammatory properties in some inflammatory pulmonary diseases. Moreover, all clinical studies have presented an improvement of the respiratory condition of cystic fibrosis patients, but the molecular and cellular mechanisms remain unknown. The aim of this study was to investigate, in bronchial epithelial cells, the effects of azithromycin on inflammatory pathways involved in cystic fibrosis. We have analysed the effects of azithromycin on cystic fibrosis and non-cystic fibrosis bronchial epithelial cell lines but also in non-immortalized non-cystic fibrosis human glandular cells. To create an inflammatory context, cells were treated with Tumor Necrosis Factor (TNF)-α or Interleukin (IL)1-β. Activation of the NF-κB pathway was investigated by luciferase assay, western blotting, and by Förster Resonance Energy Transfer imaging, allowing the detection of the interaction between the transcription factor and its inhibitor in live cells. In all conditions tested, azithromycin did not have an anti-inflammatory effect on the cystic fibrosis human bronchial epithelial cells and on CFTR-inhibited primary human bronchial glandular cells. More, our data showed no effect of azithromycin on IL-1β-or TNF-α-induced IL-8 secretion and NF-κB pathway activation. Taken together, these data show that azithromycin is unable to decrease in vitro inflammation in cystic fibrosis cells from airways.
Azithromycin increases survival and reduces lung inflammation in cystic fibrosis mice
Inflammation Research, 2009
Objective and Design-Azithromycin (AZM) has been used as an anti-inflammatory agent in the treatment of cystic fibrosis (CF), particularly those with chronic infection with P. aeruginosa (PA). To investigate mechanisms associated with the beneficial effects of AZM in CF, we examined bacterial load, cytokine levels, and clearance of inflammatory cells in CF mice infected with mucoid PA and treated with AZM. Methods-Gut-corrected Cftr tm1Unc-TgN(FABPCFTR)#Jaw CF mice infected with an alginateoverproducing PA CF-isolate were treated with AZM or saline and examined for survival of animals, lung bacterial load, inflammation, cytokine levels and apoptotic cells up to 5 days postinfection. Results-Administration of AZM (20 mg/kg) 24 h after the infection improved 5-day survival to 95% compared to treatment with saline (56%). AZM administration was associated with significant reductions in bacterial load, decreased lung inflammation and increased levels of IFNγ. AZM increased macrophage clearance of apoptotic neutrophils from the lung. Conclusion-AZM enhances bacterial clearance and reduces lung inflammation by improving innate immune defense mechanisms in CF mice.
Anti-inflammatory effects of azithromycin in cystic fibrosis airway epithelial cells
Biochemical and Biophysical Research Communications, 2006
We aimed at identifying molecular mechanisms for anti-inflammatory effects of azithromycin (AZM) suggested by clinical evidences. IL-8 expression and DNA binding activity of two key pro-inflammatory transcription factors (TF), NF-jB and AP-1, were investigated in cystic fibrosis (CF) and isogenic non-CF airway epithelial cell lines. AZM reduced about 40% of IL-8 mRNA and protein expression (n = 9, p = 0.02, and n = 4, p = 0.00011) in CF cells reaching the levels of non-CF cells. In the presence of AZM we found about 50% and 70% reduction of NF-jB and AP-1 DNA binding, respectively (n = 3, p = 0.01, and n = 3, p = 0.0017), leading to levels of non-CF cells. The relevance of NF-jB and AP-1 in regulating IL-8 promoter transcriptional activity was demonstrated by gene reporter assays (n = 4, p = 8.54 · 10 À7 , and n = 4, p = 6.45 · 10 À6 ). Our data support the anti-inflammatory effects of AZM in CF cells, indicating inhibition of transcription of pro-inflammatory genes as possible mechanism, thus providing a rationale for the possible use of specific TF inhibitors for therapy.
Azithromycin alters macrophage phenotype
Journal of Antimicrobial Chemotherapy, 2008
Objectives: To investigate the in vitro effects of azithromycin on macrophage phenotype. Utilizing a mouse macrophage cell line (J774), we examined the effect of azithromycin on the properties that define classical macrophage activation (M1) and alternative macrophage activation (M2).
Antimicrobial Agents and Chemotherapy, 2007
Azithromycin (AZM) ameliorates lung function in cystic fibrosis (CF) patients. This macrolide has been suggested to have anti-inflammatory properties as well as other effects potentially relevant for therapy of CF. In this study, we utilized three CF (IB3-1, 16HBE14o-AS3, and 2CFSMEo-) and two isogenic non-CF (C38 and 16HBE14o-S1) airway epithelial cell lines to investigate whether AZM could reduce tumor necrosis factor alpha (TNF-␣) mRNA and protein levels by real-time quantitative PCR analysis and an enzyme-linked immunosorbent assay (ELISA), respectively. We studied the effects on the DNA binding of NF-B and specificity protein 1 (Sp1) by an ELISA. Non-CF cells express significantly lower TNF-␣ mRNA and protein levels than an isogenic CF cell line. In CF cells, AZM treatment causes a 30% reduction of TNF-␣ mRNA levels (P < 0.05) and a 45% decrease in TNF-␣ secretion (P < 0.05), reaching approximately the levels of the untreated isogenic non-CF cells. In CF cells, NF-B and Sp1 DNA binding activities were also significantly decreased (about 45 and 60%, respectively; P < 0.05) after AZM treatment. Josamycin, a macrolide lacking clinically described anti-inflammatory effects, was ineffective. Finally, AZM did not alter the mRNA expression levels of interleukin-6, a proinflammatory molecule not differentially expressed in CF and isogenic non-CF cells. The results of our study support the anti-inflammatory activities of this macrolide, since we show that AZM reduced the levels of TNF-␣ and propose inhibitions of NF-B and Sp1 DNA binding as possible mechanisms of this effect.
Azithromycin increases phagocytosis of apoptotic bronchial epithelial cells by alveolar macrophages
European Respiratory Journal, 2006
Chronic obstructive pulmonary disease (COPD) is associated with increased apoptosis and defective phagocytosis in the airway. As uncleared cells can undergo secondary necrosis and perpetuate inflammation, strategies to improve clearance would have therapeutic significance. There is evidence that the 15-member macrolide antibiotic azithromycin has antiinflammatory properties. Its effects may be increased in the lung due to its ability to reach high concentrations in alveolar macrophages (AMs).
American Journal of Respiratory Cell and Molecular Biology, 2009
Pulmonary infection with an exaggerated inflammatory response is the major cause of morbidity and mortality in cystic fibrosis (CF). The objective of this study was to determine whether differences in the innate immune system underlie the exaggerated immune response in CF. We established a model that recapitulates the exaggerated immune response in a CF mouse model by exposure to Pseudomonas aeruginosa LPS and assessed the pulmonary cellular and cytokine responses of wild-type (WT) and CF mice. Compared with WT mice, CF mice had increased numbers of neutrophils and increased proinflammatory cytokines in their bronchoalveolar lavage fluid after LPS exposure. Based on the increased levels of IL-1a, IL-6, granulocyte colony-stimulating factor (G-CSF), and keratinocyte chemoattractant, all of which are known to be produced by macrophages, we tested whether two populations of macrophages, bone marrowderived macrophages and alveolar macrophages, directly contribute to the elevated cytokine response of CF mice to LPS. After in vitro stimulation of bone marrow-derived macrophages and alveolar macrophages with LPS, IL-1a, IL-6, G-CSF, and monocyte chemoattractant protein-1 were higher in CF compared with WT cell supernatants. Quantitative analyses for IL-6 and keratinocyte chemoattractant revealed that LPS-stimulated CF macrophages have higher mRNA and intracellular protein levels compared with WT macrophages. Our data support the hypothesis that macrophages play a role in the exuberant cytokine production and secretion that characterizes CF, suggesting that the macrophage response may be an important therapeutic target for decreasing the morbidity of CF lung disease.