Clarithromycin has an immunomodulatory effect on ERK-mediated inflammation induced by Pseudomonas aeruginosa flagellin.
OBJECTIVE
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.
Shinkai
Masaharu
M
Department of Pediatrics, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1081, USA.
López-Boado
Yolanda S
YS
Rubin
Bruce K
BK
D000464:Alginates / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D000893:Anti-Inflammatory Agents / Q000494:pharmacology; D017291:Clarithromycin / Q000494:pharmacology*; D003907:Dexamethasone / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000201:enzymology; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D005408:Flagellin / Q000302:isolation & purification; D007155:Immunologic Factors; D007202:Indicators and Reagents; D007249:Inflammation / Q000139:chemically induced* / Q000201:enzymology / Q000473:pathology*; D016209:Interleukin-8 / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D010766:Phosphorylation; D011550:Pseudomonas aeruginosa / Q000737:chemistry*; D015398:Signal Transduction / Q000187:drug effects; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics
D000464:Alginates; D000900:Anti-Bacterial Agents; D000893:Anti-Inflammatory Agents; D007155:Immunologic Factors; D007202:Indicators and Reagents; D016209:Interleukin-8; D008070:Lipopolysaccharides; D005408:Flagellin; D003907:Dexamethasone; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D017291:Clarithromycin