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RESEARCH PRODUCT

Cigarette smoke extract activates human bronchial epithelial cells affecting non-neuronal cholinergic system signalling in vitro.

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Abstract Aims Acetylcholine (ACh) is synthesized by Choline Acetyl-Transferase (ChAT) that exerts its physiological effects in airway epithelial cells via muscarinic receptor (MR) activation. We evaluate the effect of ACh stimulation on human bronchial epithelial cells (16-HBE) and test whether cigarette smoke extract (CSE) can modify the basal cellular response to ACh affecting the non-neuronal cholinergic system signalling. Main methods ACh stimulated 16-HBE were tested for ACh-binding, Leukotriene B 4 (LTB 4 ) release and ERK1/2 and NFkB pathway activation. Additionally, we investigated all the aforementioned parameters as well as ChAT and MR proteins and mRNA expression and endogenous ACh production in CSE-treated 16-HBE. Key findings We showed that ACh induced in 16-HBE, in a concentration-dependent manner, LTB 4 release via the activation of ERK1/2 and NFkB pathways. The addition of Tiotropium (Spiriva®), Gallamine, Telenzepine and 4-DAMP (muscarinic receptor antagonists), as well as of PD 098059 (MAPKK inhibitor) and BAY117082 (inhibitor of IkBα phosphorilation), down-regulated the ACh-induced effects. Additionally, CSE treatment of 16-HBE increased the binding of ACh, and shifted the LTB4 release from the concentration ACh 1 μM to 10 nM. Finally, we observed that the treatment of 16-HBE with CSE increased the expression of ChAT, M 2 and M 3 and of endogenous ACh production in 16-HBE. Tiotropium regulated the LTB4 release and ACh production in CSE treated 16-HBE. Significance CSE increases the pro-inflammatory activity of human bronchial epithelial cells, and promotes the cellular response to lower concentrations of ACh, by affecting the expression of ChAT and MRs. Tiotropium might prevent pro-inflammatory events generated by ACh together with CSE.