Conclusions
Nicotine/cotinine exposure differentially alters bacterial metabolism and vesicle physiology, ultimately impacting the inflammatory response of tolerogenic macrophages.
Methods
In vitro cultures of the gastrointestinal bacteria (Bacteroides fragilis 25285, Prevotella bryantii B14, and Acetoanaerobium sticklandii SR) were continuously exposed to nicotine or cotinine. Supernatant samples were collected for fermentation acid analysis. Vesicles were collected and analyzed for physiological changes in number, size, and total protein cargo. Cultured macrophages were stimulated to a tolerogenic phenotype, exposed to control or altered (nicotine or cotinine - exposed) vesicles, and inflammatory plasticity assessed via inflammatory cytokine production.
Results
Nicotine/cotinine exposure differentially affected metabolism of all bacteria tested in a Gram (nicotine) and concentration-dependent (cotinine) manner. Physiological studies demonstrated changes in vesiculation number and protein cargo following nicotine/cotinine exposures. Continuous exposure to 1 μM nicotine and 10 μM cotinine concentrations reduced total protein cargo of Gram (-) - 25285 and B14 vesicles, while cotinine generally increased total protein in Gram (+) - SR vesicles. We found that theses physiological changes to the vesicles of 25285 and SR formed under nicotine and cotinine, respectively, challenged the plasticity of tolerogenic macrophages. Tolerogenic macrophages exposed to vesicles from 1 μM nicotine, and 5 or 10 μΜ cotinine cultures produced significantly less IL-12p70, TNFα, or KC/GRO, regardless of macrophage exposure to nicotine/cotinine. Conclusions: Nicotine/cotinine exposure differentially alters bacterial metabolism and vesicle physiology, ultimately impacting the inflammatory response of tolerogenic macrophages.