Activated tonsillar B cells slow down the differentiation of monocytes into iDCs and delay the complete maturation of iDCs into mDCs. Tonsillar B cells were purified and stimulated for 5 days on CD40L-transfected NIH3T3 murine fibroblasts in the presence of 0.25μM CpG-ODN 2006. Peripheral blood monocytes (A-C) were purified from healthy donors (day 0), cultured alone (1:0), or with activated B cells added with their supernatant (ratio 1 monocyte/4 B cells) in the presence of GM-CSF (1000 U/mL) and IL-4 (500 U/mL) for 6 days. (A) Expression of HLA-DR, CD80, and CD86 were evaluated by flow cytometry on CD19-negative cells at days 2, 4, and 6. A representative experiment is shown with dotted lines corresponding to isotype controls, open histograms to monocytes cultured alone, and gray histograms to monocytes cocultured with B cells. (B) MFI of HLA-DR, CD80, and CD86. *P < .05. (C) Representative example of the expression of CD40, CD45RO, CD54, CD58, CD11b, CD11c, CD25, and CD83 evaluated by flow cytometry. In other experiments, monocytes were purified from the peripheral blood of healthy donors and stimulated with GM-CSF and IL-4 for 6 days. (D-F) Differentiated iDCs were then cultured alone (1:0) or with activated B cells added with their supernatant (ratio 1 iDC/4 B cells), in the presence of LPS (100 ng/mL) and IFN-γ (1000 U/mL) for 48 hours. (D) Expression of HLA-DR, CD80, CD86, and IL-12 were evaluated by flow cytometry on CD19-negative cells. A representative experiment is shown with dotted lines corresponding to isotype controls, open histograms to iDCs cultured alone, and gray histograms to iDCs cocultured with B cells. (E) MFI of HLA-DR, percentages of CD80^low and CD86^low iDCs, and MFI of IL-12 are shown. *P < .05; ns, nonsignificant. (F) Representative example of the expression of CD45RO, CD11b, CD11c, and CD83 evaluated by flow cytometry.