Muscarinic receptors mediate stimulation of collagen synthesis in human lung fibroblasts

Abstract

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 microM by 40-55%, in a different series of experiments). Likewise, 10 microM oxotremorine caused an increase of approximately 65%. For comparison, transforming growth factor-beta1 (5 ng x mL(-1)) caused an increase of approximately 80%. Effects of carbachol on total [(3)H]-proline incorporation and collagenase-sensitive [(3)H]-proline fraction were similar. The effect of 10 microM carbachol was inhibited by tiotropium (inhibitory concentration of 50%: 110 pM), prevented by pertussis toxin and the mitogen-activated protein kinase inhibitor, PD 98059. Muscarinic agonists also enhanced [(3)H]-proline incorporation in a tiotropium-sensitive manner in HEL-299 cells and phLFb. 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.