title: Nicotinic acetylcholine receptor subunits and receptor activity in the epithelial cell line HT29.
abstract: In the present study we have used RT-PCR to investigate nicotinic acetylcholine receptor (nAChR) subunit expression, and studied the effect of nicotine on TNFalpha-induced cytokine (IL-8) release in the epithelial cell line HT29. RNA was extracted using a commercial kit and amplified by RT-PCR. RT-PCR products were separated by electrophoresis and visualised using ethidium bromide. IL-8 release was measured by ELISA from cells activated for 6 h with TNFalpha (50 ng ml(-1)) in the absence and presence of nicotine (10(-11)-10(-6) M). HT29 cells contained mRNA for beta1, alpha4, alpha5, and alpha7 nAChR subunits. Activation of HT29 cells increased IL-8 release from undetectable amounts to 3.92 +/- 0.51 ng ml(-1) (n = 5). Nicotine significantly inhibited TNFalpha-induced IL-8 release in a concentration related manner with peak inhibition occurring at 10(-7) M (2.39 +/- 0.78 ng ml(-1), n = 5). Our data suggests that, while HT29 cells express mRNA for nAChR subunits, the only nAChR subunits that could form functional receptors and inhibit IL-8 release are alpha7.
authors.affiliation: Department of Biosciences, CP Snow Building, University of Hertfordshire, College Lane, Hatfield, Hertfordshire AL10 9AB, UK.
authors.forename: Andrea E
authors.initials: AE
authors.lastname: Summers
authors.forename: Clifford J
authors.initials: CJ
authors.lastname: Whelan
authors.forename: Mike E
authors.initials: ME
authors.lastname: Parsons
mesh_terms: D003093:Colitis, Ulcerative / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000378:metabolism*; D019073:HT29 Cells; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D009538:Nicotine / Q000494:pharmacology; D018722:Nicotinic Agonists / Q000494:pharmacology; D011978:Receptors, Nicotinic / Q000096:biosynthesis / Q000187:drug effects / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D016209:Interleukin-8; D018722:Nicotinic Agonists; D011978:Receptors, Nicotinic; D014409:Tumor Necrosis Factor-alpha; D009538:Nicotine