Lack of p100 results in the induction of ectopic HEV-like structures in the splenic red pulp. (A) QRT-PCR analysis of GlyCAM-1, β3GlcNAcT-3 (core 1), Core2GlcNAcT-I (core 2), GlcNAc6ST-1, GlcNAc6ST-2, and FucT-VII in spleens from wild-type and p100^−/− mice. Note the log scale of relative mRNA expression. Error bars are SD. Significant differences are indicated (Student t test, ***P < .002). (B) Immunohistochemical staining of wild-type and p100^−/− spleen sections for MZMs (anti-MARCO; DAB brown) and MECA-79 (Vector Blue). (C) 2-color immunofluorescence of p100^−/− spleen sections stained with MECA-367 (M367; FITC green) and MECA-79 mAbs (M79; Texas red). Most MECA-367⁺ structures exhibited coexpression of PNAd (merge; yellow fluorescence). Insets, mesenteric LN HEVs from p100^−/− mice. (D) 2-color immunofluorescence of p100^−/− spleen sections stained with MECA-79 (M79; FITC green) and anti-CCL21 (SLC; Texas red). Most MECA-79⁺ structures exhibited coexpression of CCL21 (merge; yellow fluorescence). (E) Spleen sections from p100^−/− mice (middle panel) and control littermates (left panel) that were intravenously injected with FITC-labeled E. coli were stained with the IBL-11 mAb (red) to indicate the white pulp. The right panel shows FITC-E. coli in p100^−/− spleen sections stained with MECA-367 (Vector Blue) to indicate ectopic HEV-like structures. Dotted lines mark the boundary of the white pulp. Images were acquired as in Figure 2E, with 20×/0.70 objective (B), 40×/0.75 objective (C,D); 20×/0.70 objective (panel E left and middle), 40×/0.75 objective (panel E right).