Acetylcholine mediates the release of IL-8 in human bronchial epithelial cells by a NFκB/ERK-dependent mechanism

Acetylcholine may play a role in cell activation and airway inflammation. We evaluated the levels of both mRNA and protein of muscarinic M-1, M-2, M-3 receptors in human bronchial epithelial cell line (16HBE). 16HBE cells were also stimulated with acetylcholine and extracellular signal-regulated kinase 1/2 (ERK1/2) and NF kappa B pathway activation as well as the IL-8 release was assessed in the presence or absence of the inhibitor of Protein-kinase (PKC) (GF109203X), of the inhibitor of mitogenic activated protein-kinase kinase (MAPKK) (PDO9805), of the inhibitor of kinaseB alpha phosphorilation (pIkB alpha) (BAY11-7082), and of muscarinic receptor antagonists tiotropium bromide, 4-Diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP), telenzepine, gallamine. Additionally, we tested the IL-8-mediated neutrophil chemotactic activity of 16HBE supernatants stimulated with acetylcholine in the presence or absence of tiotropium. 16HBE cells expressed both protein and mRNA for muscarinic M-3, M-2 and M-1 receptors with levels of muscarmic M-3 receptor> muscarinic M-1 receptor> muscarinic M-2 receptor. Acetylcholine (10 mu M) significantly stimulated ERK1/2 and NF kappa B activation as well as IL-8 release in 16HBE cells when compared to basal values. Furthermore, while the use of tiotropium, 4-DAMP, GF109203X, PD098059, BAY11-7082 completely abolished these events, the use of telenzepine and gallamine were only partially able to downregulate these effects. Additionally, acetylcholine-mediated IL-8 release from 16HBE cells significantly increased chemotaxis toward neutrophils and this effect was blocked by tiotropium. In conclusion, acetylcholine activates the release of IL-8 from 16HBE involving PKC, ERK 1/2 and NF kappa B pathways via muscarinic receptors, suggesting that it is likely to contribute to IL-8 related neutrophilic inflammatory disorders in the airway. Thus, muscarinic antagonists may contribute to control inflammatory processes in airway diseases. (c) 2008 Elsevier B.V. All rights reserved.