Expression of muscarinic receptors by human macrophages

Macrophages increase in number and are highly activated in chronic obstructive pulmonary disease (COPD). Muscarinic receptor antagonists inhibit acetylcholine-stimulated release of neutrophilic chemoattractants, suggesting that acetylcholine may regulate macrophage responses. Therefore, expression and function of components of the non-neuronal cholinergic system in monocyte-macrophage cells was investigated. RNA was isolated from monocytes, monocyte-derived macrophages (MDMs), lung and alveolar macrophages from nonsmokers, smokers and COPD patients, and expression of the high-affinity choline transporter, choline acetyltransferase, vesicular acetylcholine transporter and muscarinic receptors (M₁–M₅) ascertained using real-time PCR. M₂ and M₃ receptor expression was confirmed using immunocytochemistry. Release of interleukin (IL)-8, IL-6 and leukotriene (LT)B₄ were measured by ELISA or EIA. All monocyte-macrophage cells expressed mRNA for components of the non-neuronal cholinergic system. Lung macrophages expressed significantly more M₁ mRNA compared with monocytes, and both lung macrophages and alveolar macrophages expressed the highest levels of M₃ mRNA. Expression of M₂ and M₃ protein was confirmed in MDMs and lung macrophages. Carbachol stimulated release of LTB₄ from lung macrophages (buffer 222.3±75.1 versus carbachol 1,118±622.4 pg·mL⁻¹; n=15, p<0.05) but not IL-6 or IL-8. LTB₄ release was attenuated by the M₃ antagonist, 1,1-dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP; half maximal effective concentration 5.2±2.2 nM; n=9). Stimulation of macrophage M₃ receptors promotes release of LTB₄, suggesting that anti-muscarinic agents may be anti-inflammatory.