Smoke, Choline Acetyltransferase, Muscarinic Receptors, and Fibroblast Proliferation in Chronic Obstructive Pulmonary Disease

Abstract

Acetylcholine (ACh), synthesized by choline acetyltransferase (ChAT), and muscarinic M(1), M(2), and M(3) receptors (MRs) are involved in fibroblast proliferation. We evaluated ChAT, MRs, and extracellular signal-regulated kinase (ERK) 1/2 and nuclear factor (NF) kappaB activation in lung fibroblasts from patients with chronic obstructive pulmonary disease (COPD), control smokers, and controls. Human fetal lung fibroblasts (HFL-1) stimulated with interleukin (IL)-1beta, tumor necrosis factor (TNF)-alpha, and cigarette smoke extracts (CSEs) were evaluated for ChAT and MR expression. We tested the effects of ACh on fibroblast proliferation and its ability to bind fibroblasts from patients with COPD, control smokers, controls, and HFL-1 stimulated with IL-1beta, TNF-alpha, and CSE. ChAT, M(1), and M(3) expression and ERK1/2 and NFkappaB activation were increased, whereas M(2) was reduced, in COPD and smoker subjects compared with controls. IL-1beta increased the ChAT and M(3), TNF-alpha down-regulated M(2), and CSE increased ChAT and M(3) expression while down-regulating the expression of M(2) in HFL-1 cells. ACh stimulation increased fibroblast proliferation in patients with COPD, control smokers, and controls, with higher effect in control smokers and patients with COPD and increased HFL-1 proliferation only in CSE-treated cells. The binding of ACh was higher in patients with COPD and in control smokers than in controls and in CSE-treated than in IL-1beta- and TNF-alpha-stimulated HFL-1 cells. Tiotropium (Spiriva; [1alpha,2beta,4beta,5alpha,7beta-7-hydroxydi-2-thienylacetyl)oxy]-9,9-dimethyl-3-oxa-9-azoniatrcyclo[3.3.1.0(24)], C(19)H(22) NO(4)S(2)Br.H(2)O), gallamine triethiodide (C(19)H(22)N(4)O(2)S.2HCl.H(2)O), telenzepine [4,9-d-dihydro-3-methyl-4-[(4-methyl-1piperazinyl) acetyl]-10H-thieno [3,4-b][1,5]benzodiazepine-10-one dihydrobromide, C(30)H(60)I(3)N(3)O(3)], 4-diphenylacetoxy-N-methylpiperidine, PD098059 [2-(2-amino-3methoxyphenyl)-4H-1benzopyran-4-one, C(16)H(13)NO(3)], and BAY 11-7082 [(E)-3-(4-methylphenylsulfonyl)-2-propenetrile, C(10)H(9)NO(2)C], down-regulated the ACh-induced fibroblast proliferation, promoting the MRs and ERK1/2 and NFkappaB pathways involvement in this phenomenon. These results suggest that cigarette smoke might alter the expression of ChAT and MRs, promoting airway remodeling in COPD and that anticholinergic drugs, including tiotropium, might prevent these events.