Recruitment of α7 nicotinic acetylcholine receptor to caveolin-1-enriched lipid rafts is required for nicotine-enhanced Escherichia coli K1 entry into brain endothelial cells

Aim: We investigate how the alpha 7 nicotinic acetylcholine receptor (alpha 7 nAChR), an essential regulator of inflammation, contributes to the alpha 7 agonist nicotine-enhanced Escherichia coli K1 invasion of human brain microvascular endothelial cells (HBMECs) through lipid rafts/caveolae-mediated signaling. Materials & methods: alpha 7 nAChR-mediated signaling and bacterial invasion were defined by lipid raft fractionation, immunofluorescence microscopy and siRNA knockdown. Results: Nicotine-enhanced bacterial invasion was dose-dependently inhibited by two raft-disrupting agents, nystatin and filipin. Significant accumulation of the lipid raft marker GM3 was observed in HBMEC induced by E. coli K1 and nicotine. The recruitment of alpha 7 nAChR and related signaling molecules, including vimentin, and Erk1/2, to caveolin-1 enriched lipid rafts was increased upon treatment with E44 or E44 plus nicotine. Erk1/2 activation (phosphorylation), which is required for alpha 7 nAChR-mediated signaling and E44 invasion, was associated with lipid rafts and nicotine-enhanced bacterial infection. Furthermore, E44 invasion, E44/nicotine-induced activation of Erk1/2 and clustering of alpha 7 nAChR and caveolin-1 was specifically blocked by both siRNAs. Conclusion: alpha 7 nAChR-mediated signaling through lipid rafts/caveolae is required for nicotine-enhanced E. coli K1 invasion of HBMEC.