JAC Advance Access originally published online on April 5, 2007
Journal of Antimicrobial Chemotherapy 2007 59(6):1096-1101; doi:10.1093/jac/dkm084
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Clarithromycin has an immunomodulatory effect on ERK-mediated inflammation induced by Pseudomonas aeruginosa flagellin
1 Department of Pediatrics, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1081, USA 2 Department of Molecular Medicine, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1081, USA
Received 19 September 2006; returned 28 January 2006; revised 9 January 2007; accepted 22 February 2007
* Corresponding author. Tel: +1-336-716-0262; Fax: +1-336-716-9229; E-mail: brubin{at}wfubmc.edu
Objectives: Pseudomonas aeruginosa exoproducts are potent triggers of immune responses in eukaryotic cells. Clarithromycin initially decreases, then increases and finally produces a sustained suppression of interleukin (IL)-8 secretion from normal human bronchial epithelial (NHBE) cells through inhibition and activation of extracellular signal-regulated kinase (ERK). This polyphasic immune response is referred to as immunomodulation.
Methods: We studied the effects of P. aeruginosa flagellin and alginate on IL-8 secretion from NHBE cells and how this was affected by clarithromycin or dexamethasone. We also assessed the upstream kinase cell signalling intermediates that appear to be responsible for flagellin-induced IL-8 secretion. ELISA was used to measure IL-8 in culture supernatants, and western blots were used to measure kinase and phosphokinase levels.
Results: Flagellin dose-dependently increased IL-8 secretion in NHBE cells at 24 h, whereas alginate had no effect on IL-8. Clarithromycin significantly decreased flagellin-induced IL-8 over the first 9 h but not at 24 h. A 60 min exposure to clarithromycin decreased flagellin-induced ERK phosphorylation, but at 24 h, clarithromycin increased phospho-ERK1/2 beyond the effect of flagellin alone. Pre-treatment with PD98059 (MEK inhibitor) decreased flagellin-induced IL-8 secretion by 47.7% (P < 0.0001) compared with control flagellin exposure and decreased basal IL-8 in the absence of flagellin by 27.9% compared with untreated control cells (P < 0.0001). Dexamethasone and PD98059 together had an additive suppressive effect on flagellin-induced IL-8 secretion.
Conclusions: P. aeruginosa flagellin, but not alginate, stimulates IL-8 secretion in NHBE cells in part through ERK1/2. This effect is modulated by clarithromycin, whereas suppression of IL-8 secretion by dexamethasone probably occurs through different pathways.
Keywords: interleukin-8 , bronchial epithelial cells , extracellular signal-regulated MAP kinases
Present address. Third Department of Internal Medicine, National Defense Medical College, Tokorozawa-city, Japan.
Present address. RI-CEDD, GlaxoSmithKline, Philadelphia, PA 19406, USA.