Tiotropium suppresses acetylcholine-induced release of chemotactic mediators in vitro

Clinical observations indicate that in chronic obstructive pulmonary disease patients, the long-acting muscarinic antagonist tiotropium delays decline in airway function, suggesting that cholinergic mechanisms contribute to long-term structural changes. Human lung fibroblasts express muscarinic receptors and the present study aimed to explore their role in controlling collagen synthesis.MRC-5, HEL-299 and primary human lung fibroblasts (phLFb) were cultured. Incorporation of [ 3 H]-proline into cellular proteins was determined as measure of collagen synthesis.In MRC-5 cells, the muscarinic agonist carbachol enhanced [3 H]-proline incorporation in a concentration-dependent manner (effective concentration of 50%: 220 nM, increase at 10 mM by 40-55%, in a different series of experiments). Likewise, 10 mM oxotremorine caused an increase of ,65%. For comparison, transforming growth factor-b1 (5 ng?mL In human lung fibroblasts, muscarinic receptors exert stimulatory effects on collagen synthesis. Prolonged blockade of muscarinic-induced collagen synthesis may contribute to reported beneficial long-term effects of anticholinergics in chronic obstructive pulmonary disease.

Section: Discussionsupporting

confidence: 90%

Muscarinic receptors mediate stimulation of collagen synthesis in human lung fibroblasts

Haag¹,

Matthiesen²,

Juergens³

et al. 2008

“…We found a predominance of M2 and M3 muscarinic subtypes, with low amounts of M1 receptors. A similar pattern of expression was described for cultured respiratory epithelial cells [5,26]. In this sense, we found that siRNA specific for siRNA-M3 and the M3 antagonist pFHHSid attenuated the MUC5AC response to carbachol and CSE, while siRNA-M2 and the M2 antagonist methoctramine were without significant effects.…”

Section: Discussionsupporting

confidence: 84%

Aclidinium inhibits cholinergic and tobacco smoke-induced MUC5AC in human airways

Cortijo¹,

Mata²,

Milara³

et al. 2010

Mucus hypersecretion and mucin MUC5AC overexpression are pathological features of chronic obstructive pulmonary disease (COPD). This study examines the inhibitory effect of aclidinium, a new long-acting muscarinic antagonist, on MUC5AC expression in human airway epithelial cells.MUC5AC mRNA (RT-PCR) and protein expression (ELISA and immunohistochemistry) were studied in human bronchial tissue and differentiated human airway epithelial cells activated with carbachol (100 mM) or cigarette smoke extract in the absence or presence of aclidinium.Carbachol increased MUC5AC mRNA and protein expression in human bronchus and cultured epithelial cells. Aclidinium inhibited the carbachol-induced MUC5AC mRNA and protein expression with potency (half maximal inhibitory concentration) ,1 nM in human bronchus and cultured airway epithelial cells. AG1478, a selective inhibitor of epidermal growth factor receptor (EGFR) tyrosine kinase, inhibited carbachol-induced MUC5AC responses, indicating EGFR transactivation. Aclidinium inhibited carbachol-induced phospho-EGFR and phospho-p44/42 MAPK expression. In cultured airway epithelial cells transfected with small interfering (si)RNA against muscarinic receptor subtypes, siRNA-M3 but not siRNA-M2 blocked carbachol-induced MUC5AC expression. Cigarette smoke-induced MUC5AC upregulation in cultured airway epithelial cells was suppressed by aclidinium.In conclusion, aclidinium decreases carbachol and tobacco smoke-induced MUC5AC overexpression in human airway epithelial cells. This effect may contribute to the clinical efficacy of aclidinium in mucus hypersecretory diseases including COPD.

“…A recent study showed that acetylcholine induces the release of neutrophil chemoattractants such as leukotriene B 4 from sputum cells of COPD patients [6]. These results are consistent with studies demonstrating that alveolar macrophages exhibit neutrophil, eosinophil and monocyte chemotactic activities in response to acetylcholine [7,8] and suggest that acetylcholine is a paracrine and/or autocrine mediator that regulates inflammatory processes.…”

supporting

confidence: 80%

Muscarinic M3 receptor stimulation increases cigarette smoke-induced IL-8 secretion by human airway smooth muscle cells

Gosens¹,

Rieks²,

Meurs³

et al. 2009

Acetylcholine is the primary parasympathetic neurotransmitter in the airways and is known to cause bronchoconstriction and mucus secretion. Recent findings suggest that acetylcholine also regulates aspects of remodelling and inflammation through its action on muscarinic receptors.In the present study, we aimed to determine the effects of muscarinic receptor stimulation on cytokine production by human airway smooth muscle cells (primary and immortalised cell lines). The muscarinic receptor agonists carbachol and methacholine both induced modest effects on basal interleukin (IL)-8 and -6 secretion, whereas the secretion of RANTES, eotaxin, vascular endothelial growth factor-A and monocyte chemoattractant protein-1 was not affected. Secretion of IL-8 and -6 was only observed in immortalised airway smooth muscle cells that express muscarinic M3 receptors. In these cells, methacholine also significantly augmented IL-8 secretion in combination with cigarette smoke extract in a synergistic manner, whereas synergistic effects on IL-6 secretion were not significant. Muscarinic M3 receptors were the primary subtype involved in augmenting cigarette smoke extract-induced IL-8 secretion, as only tiotropium bromide and muscarinic M3 receptor subtype selective antagonists abrogated the effects of methacholine.Collectively, these results indicate that muscarinic M3 receptor stimulation augments cigarette smoke extract-induced cytokine production by airway smooth muscle. This interaction could be of importance in patients with chronic obstructive pulmonary disease.