Although nicotine is thought to be one of the major immunomodulatory components of cigarette smoking, how nicotine alters the host defense of the lung and, in particular, immune responses of alveolar macrophages, which are critical effector cells in the lung defense to infection, is poorly understood. Nicotinic acetylcholine receptors (nAChRs) are the receptor for nicotine and may be involved in the modulation of macrophage function by nicotine. In this study, therefore, nicotine-induced suppression of antimicrobial activity and cytokine responses of alveolar macrophages mediated by nAChRs to Legionella pneumophila, a causative agent for pneumonia, were examined. The murine MH-S alveolar macrophage cell line cells expressed the messages for α4 and β2 subunits of nAChRs, but not α7 subunits, determined by RT-PCR. The nicotine treatment of MH-S alveolar macrophages after infection with L. pneumophila significantly enhanced the replication of bacteria in the macrophages and selectively down-regulated the production of IL-6, IL-12, and TNF-α, but not IL-10, induced by infection. These effects were completely blocked by a nonselective antagonist, d-tubocurarine, for nAChRs, but not by a selective antagonist, α-bungarotoxin, for α7-nAChRs. Furthermore, the stimulation of nAChRs with another agonist, 1,1-dimethyl-4-phenylpiperazinium iodide, showed the same effects, which were blocked by the antagonist d-tubocurarine, on the bacterial replication and cytokine regulation with that of nicotine. Thus, the results revealed that nAChRs, the major exogenous ligands of which are nicotine, are involved in the regulation of macrophage immune function by nicotine and may contribute to the cigarette-induced risk factors for respiratory infections in smokers.

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Virus infections are a major cause of chronic obstructive pulmonary disease (COPD) exacerbations. Recently, Toll-like receptor 3 (TLR3) has been demonstrated to react to double-stranded RNA (dsRNA) and to be involved in the immune responses after viral infections. In the present study, we examined whether oxidative stress, which is involved in the pathogenesis of COPD, enhances the responses of TLR3 in airway epithelial cells. The effect of hydrogen peroxide (H 2 O 2 ) on the release of IL-8 from BEAS-2B cells and primary human bronchial epithelial cells after stimulation with polyinosinepolycytidylic acid [poly(I:C)], a synthetic analog of viral dsRNA and a ligand for TLR3, and the signal transduction were examined. One hundred to 150 mM H 2 O 2 significantly potentiated the release of IL-8 from the epithelial cells after stimulation with 10 mg/ml poly(I:C). The H 2 O 2 -augmented IL-8 release was inhibited by treatment with N-acetylcysteine. One hundred micromoles of H 2 O 2 enhanced the translocation of nuclear factor (NF)-kB p65, but not that of interferon regulatory factor-3 (IRF-3), into the nucleus and the NF-kB DNA binding activity after poly(I:C) stimulation, which effect was inhibited not by the silencing of IRF-3 but by MG132, a proteasome inhibitor, or dexamethasone. One hundred micromoles of H 2 O 2 potentiated the TLR3 expression on the airway epithelial cells treated with poly(I:C). These data suggest that oxidative stress augments the response of TLR3 in airway epithelial cells via NF-kB and that this effect might be partly mediated by the enhancement of TLR3 expression. Modulation of this pathway may be a therapeutic target for viral-induced exacerbations of COPD.

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