Analytical Cellular Pathology / 2015 / Article / Fig 2

Research Article

Ex Vivo Nicotine Stimulation Augments the Efficacy of Human Peripheral Blood Mononuclear Cell-Derived Dendritic Cell Vaccination via Activating Akt-S6 Pathway

Figure 2

Ex vivo nicotine stimulation augments semimature DCs-mediated PBMC proliferation, promotes the release of IL-12, and increases the percentage of IFN-gamma/CD8 cells in the supernatant of cocultured DCs-PBMC. DCs derived from human PBMC with 100 ng/mL recombinant human GM-CSF and IL-4 were conferred with α-bungarotoxin (2 μg/mL) or tubocurarine chloride (4 × 10−5 mol/L) 2 h prior to nicotine (10−7 mol/L) 12~15 h stimulation. Then, the DCs were pulsed with endotoxin-free EndoGrade-ovalbumin (50 µg/mL) for 3 h, followed by 1 h LPS (1 ng/mL) stimulation. After that, the DCs were coincubated with PBMC of the same donor at a ratio of 1 : 1 for 5 d and the effect of nicotine on DCs-mediated PBMC proliferation, the IL-12 release in the supernatant, and IFN-gamma/CD8 cell percentage of cocultured DCs-PBMC were determined by BrdU cell proliferation assay (a), ELISA (b), and flow cytometry (c), respectively. Data were given as mean ± SEM, , one-way ANOVA with post-Newman-Keuls test. , and . One representative from 3 independent experiments is shown. Ni: nicotine; BTX: α-bungarotoxin; and TC: tubocurarine chloride.

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