Fig. 4.

Identification of nicotinic acetylcholine receptors on neutrophils

(A) Staining of human neutrophils with Alexa Fluor 647-labelled α-bungarotoxin and anti-α7-specfic nAchR antibody, merged with nuclear stain DAPI(4’,6-diamidino-2-phenylindole)), shows a pattern of surface staining (scale bar: 50 μm). (B) Magnified view of neutrophil stained with α-bungarotoxin and an anti-α7-specfic nAchR antibody, demonstrating co-localization on the plasma membrane (scale bar: 10 μm). (C) Stimulation of neutrophils with an α7-nAChR agonist, GTS-21 (0.8 mM), results in nicotine-induced NETosis (*P < 0.05, **P < 0.01). (D) Nicotine-induced NETosis is attenuated by pretreating neutrophils with a non-specific nAChR antagonist (50 μM) or an α7-specfic nAChR antagonist (10 μM) before exposure to nicotine (*P < 0.05). Bars represent the mean (s.e.m.). nAChR: nicotinic acetylcholine receptor.