Figure 7.

Cytokine production by stimulated B cells. (A) The number of cytokine positive B cells per 1000 total CD45⁺ cells for each stimulation at 24 h with the background (unstimulated) cytokine-production removed. (B) Cluster analysis of IL-4 secreting cells with cluster 6 and 12 corresponding to B cells. The pie-chart indicates cluster-specific and percent of total contribution to all IL-4 secreting cells. (C) Evaluation of the effect of LAM (orange circles), PIM (green boxes), or PPD (black triangles) stimulation on IL-4 secreting B cell clusters. (D) Overlay of concatenated IL-4 secreting B cells for cluster 6 (green) and cluster 12 (blue) assessing IgD and CD27 or CD11c and CD27 surface expression. (E, G) A similar analysis for IL-10 secreting cells and (H–J) GM-CSF secreting cells. (K) IL-4 and IL-10 co-expression among GM-CSF⁺ B cells (green) and total B cells (grey). (L) Overlay scatter plot of GM-CSF⁺IL-4⁺IL-10⁺ triple-secreting B cells (red) and total B cells (grey) assessing IgD and CD27 or CD11c and CD27 surface expression, with frequency of cells included in the gates indicated. (M) Overlay histogram indicating HLA-DR expression for GM-CSF⁺IL-4⁺IL-10⁺ triple-secreting B cells (red) and total B cells (grey). Numbers in dot plots indicate the percentage within the cluster. Statistical differences between stimulations in individual groups (A, F, I) were evaluated by Friedman’s test with Dunnet’s posttest, while comparisons within multiple clusters (C) were evaluated by a matched-pair two-way ANOVA with Geissner-Greenhouse correction followed by Tukey’s posttest (n=15/stimulation). *p<0.05, **p<0.01, ***p<0.001. n=15 for each group. Scatter and overlay plots show data concatenated from all samples and donors (n=60).