Rapid disruption of the splenic MZ area upon in vivo treatment with α-NRR2 antibody. (A) irf4^−/− mice were injected with α-NRR2 (12 and 24 h) or IgG (24 h) antibodies, and NOTCH2 and B220 expression in spleen sections was assessed by IF. Nuclei were stained with DAPI. Dashed lines demarcate the border between the FO and MZ areas. Arrows indicate B220⁺ B cells expressing nuclear NOTCH2. One representative mouse out of two per group is shown (two independent experiments). Boxes demarcate the area shown below. (B) Wild-type mice were injected with α-NRR2 antibody (or IgG control), and CD21 and CD23 expression on CD19⁺ splenic (top) or peripheral blood (bottom) B cells was analyzed by flow cytometry at the indicated time points after antibody injection. Numbers above gates indicate percentage of CD23⁻CD21^hi/+ B cells. Graphs show mean value ± SD of CD23⁻CD21^hi/+ B cells in the spleen and in the peripheral blood of wild-type mice at the indicated time points (n = 4, four independent experiments). Statistical significance was analyzed by Student’s t test (*, P < 0.05; **, P < 0.01; ***, P < 0.001). (C) Wild-type mice were treated as in B, and IgM expression in the spleen was assessed by IHC. One representative mouse out of four per group is shown (four independent experiments). Bars: (A, top) 100 µm; (A, bottom) 50 µm; (C, top) 1,000 µm; (C, middle) 400 µm; (C, bottom) 200 µm.