Fig. 2

Activated pre-naïve B cells show IL-10-mediated ineffective promotion of CD4⁺ T-cell proliferation. a Representative contour plot showing the frequencies of CD4⁺INF-γ⁺ T cells and CD4⁺TNF-α⁺ T cells measured by intracellular cytokine staining after co-culture with B cells and CD154-L cells for 72 hours. b The frequencies of CD4⁺INF-γ⁺ T cells and CD4⁺TNF-α⁺ T cells measured as described in (a) (n = 4, cumulative data from three independent experiments). c Effect of total B cells and each B-cell subset on CD4⁺ T-cell proliferation in co-culture with B cells and CD154-L cells for 5 days (n = 10, cumulative data from eight independent experiments). d IFN-γ and TNF-α secretion was measured by enzyme-linked immunosorbent assay in the culture supernatants from experiments described in (c). e The effect of each B-cell subset on T-cell proliferation cells in co-culture of CD4⁺ T cells, B cells, and CD154-L cells for 5 days cultured in medium alone or in the presence of anti-IL-10 mAb (n = 6, cumulative data from five independent experiments). f IFN-γ and TNF-α secretion tested in supernatants from cultures as described in (e). g Representative histogram showing frequencies of CD4⁺FoxP3⁺ T cells in co-culture with CD4⁺ T cells, B cells, and CD154-L cells for 5 days. h The effect of each B-cell subset on frequencies of CD4⁺FoxP3⁺ T cells from experiments described in (g) (n = 3, cumulative data from three independent experiments). i The effect of pre-naïve B cells on frequencies of CD4⁺FoxP3⁺ T cells from experiments described in (g) cultured in medium alone or in the presence of anti-IL-10 mAb (n = 3, cumulative data from three independent experiments). *P < 0.05, **P < 0.01. IFN-γ interferon-gamma, IL interleukin, mAb monoclonal antibody, TNF-α tumor necrosis factor-alpha