Figure S2.
Most cDC2s are located in follicles in CXCR5-deficient mice, whereas Cxcr5−/− MZ B cells in Cd19−/− mice block FTY720-induced cDC2 relocalization.(A) Frequencies of in vivo anti-CD45-PE–labeled CD8+ cDC1s in Cd19+/+ and Cd19−/− chimeras. (B) Frequencies of in vivo anti-CD45-PE–labeled CD8+ cDC1s in MD4/B6 and MD4/ML5 chimeras. Each symbol represents one mouse, and lines denote means. Data are pooled from two independent experiments. (C) Gel image of Cd11c-Cre–mediated recombination of S1pr1loxP using cDC2 and cDC1 DNA samples. S1pr1ΔEx2, S1pr1 allele after Cre-mediated recombination. (D and E) Frequencies of in vivo anti-CD45-PE–labeled MZ B cells (D) and cDC1s (E) in spleens of indicated genotyped BM chimeras. Each symbol represents one mouse, and lines denote means. Data are pooled from two independent experiments. (F–H) Frequencies of CD21hi CD23lo MZ B cells in total B220+ B cells (F), of in vivo anti-CD45-PE–labeled cDC2s (H), and representative distribution patterns of DCIR2+ cDC2s (red) relative to IgD+ B cells (blue) in spleens (G) of mice of the indicated types. Scale bar, 200 µm. (I–L)Cxcr5−/− MZ B cells in Cd19−/− mice restrict cDC2 access. (I) Representative profile of Cxcr5−/− MZ B cell reconstitution 2 mo after transfer. (J) Representative distribution patterns of IgMhi MZ B cells (red) and CD169+ MMMs relative to IgD+ follicular B cells (blue) in spleens of 4-h saline- or FTY720-treated mice. Scale bar, 200 µm. (K and L) Frequencies of in vivo anti-CD45-PE–labeled cDC2s (K) and representative distribution patterns of DCIR2+ cDC2s (red) relative to IgD+ B cells (blue) in spleens (L) of 4-h saline- or FTY720-treated mice. Scale bar, 200 µm. (M) Frequencies of PKH26-dye acquisition in DCIR2+ cDC2s of indicated genotypes 3 h after immunization with PKH26-labeled SRBCs. (N) Mean fluorescence intensity (MFI) and representative histogram of surface CD86 expression on cDC2s from mice treated as in M. (O and P) Representative FACS plot of proliferation of transferred OT-II T cells in mice of the indicated genotype at day 3 after SRBCs-OVA (O) or OVA plus LPS (P) immunization. Each symbol represents one mouse, and lines denote means. One of two independent experiments with similar results is shown, in which each group had at least three mice. Sections are representative of multiple spleen cross sections from five mice of each type. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Most cDC2s are located in follicles in CXCR5-deficient mice, whereas Cxcr5−/− MZ B cells in Cd19−/− mice block FTY720-induced cDC2 relocalization. (A) Frequencies of in vivo anti-CD45-PE–labeled CD8+ cDC1s in Cd19+/+ and Cd19−/− chimeras. (B) Frequencies of in vivo anti-CD45-PE–labeled CD8+ cDC1s in MD4/B6 and MD4/ML5 chimeras. Each symbol represents one mouse, and lines denote means. Data are pooled from two independent experiments. (C) Gel image of Cd11c-Cre–mediated recombination of S1pr1loxP using cDC2 and cDC1 DNA samples. S1pr1ΔEx2, S1pr1 allele after Cre-mediated recombination. (D and E) Frequencies of in vivo anti-CD45-PE–labeled MZ B cells (D) and cDC1s (E) in spleens of indicated genotyped BM chimeras. Each symbol represents one mouse, and lines denote means. Data are pooled from two independent experiments. (F–H) Frequencies of CD21hi CD23lo MZ B cells in total B220+ B cells (F), of in vivo anti-CD45-PE–labeled cDC2s (H), and representative distribution patterns of DCIR2+ cDC2s (red) relative to IgD+ B cells (blue) in spleens (G) of mice of the indicated types. Scale bar, 200 µm. (I–L)Cxcr5−/− MZ B cells in Cd19−/− mice restrict cDC2 access. (I) Representative profile of Cxcr5−/− MZ B cell reconstitution 2 mo after transfer. (J) Representative distribution patterns of IgMhi MZ B cells (red) and CD169+ MMMs relative to IgD+ follicular B cells (blue) in spleens of 4-h saline- or FTY720-treated mice. Scale bar, 200 µm. (K and L) Frequencies of in vivo anti-CD45-PE–labeled cDC2s (K) and representative distribution patterns of DCIR2+ cDC2s (red) relative to IgD+ B cells (blue) in spleens (L) of 4-h saline- or FTY720-treated mice. Scale bar, 200 µm. (M) Frequencies of PKH26-dye acquisition in DCIR2+ cDC2s of indicated genotypes 3 h after immunization with PKH26-labeled SRBCs. (N) Mean fluorescence intensity (MFI) and representative histogram of surface CD86 expression on cDC2s from mice treated as in M. (O and P) Representative FACS plot of proliferation of transferred OT-II T cells in mice of the indicated genotype at day 3 after SRBCs-OVA (O) or OVA plus LPS (P) immunization. Each symbol represents one mouse, and lines denote means. One of two independent experiments with similar results is shown, in which each group had at least three mice. Sections are representative of multiple spleen cross sections from five mice of each type. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

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