Figure 1
Figure 1. Maintenance of spleen microarchitecture is primarily dependent on TNF expressed by B cells with distinct contribution of T-cell–derived TNF. (A) Mice were immunized with i.p. SRBCs and spleen sections analyzed on day 8. B-TNF KO mice display reduced GCs, lack of polarized B-cell follicles, and disruption of FDC networks. Note that some FDC- and PNA-positive cells are present in the spleens of B-TNF KO mice (shown by arrows), while no FDCs and no polarized B-cell follicles are observed in spleen of T,B-TNF KO mice. Original magnification is ×200. Representative images from 1 of 3 independent experiments (n = 5 mice per group) are shown. (B) Quantification of GC and FDC areas from panel A. Data represent means ± SEM; *P < .05, **P < .01, ***P < .001. (C) Analysis of germinal center B cells. Mice were immunized with i.p. SRBCs, and GL7+ GC B cells were analyzed by flow cytometry on day 8 (represents 1 of 2 independent experiments, 3 mice per group). Data represent means ± SD; *P < .05. (D) MZ structure is dependent on TNF produced by B, T, and other cell types. Mice were immunized with i.p. SRBCs, and spleen sections were analyzed on day 8. Frozen spleen sections were labeled with anti-ER-TR7, anti-CD169, and anti-MAdCAM-1 antibodies. Note the increased breadth of the MZ (ER-TR7 labeling), reduced and disorganized MAdCAM-1, and CD169 staining in B-TNF, T,B-TNF, and TNF KO mice (marked by black arrows). Microphotographs were taken with an Olympus BX41 microscope with a Zeiss Axiocam digital camera; images were captured with Zeiss Imaging AxioVision (Version 4.7) and processed using Adobe Photoshop CS3 (Version 10.0). Original magnification is ×200. Representative images from 1 of 3 independent experiments (n = 5 mice per group) are shown. (E) Quantification of MZ size on panel D (ER-TR7 labeling). MZ size (shown by arrows) was quantified using NIH ImageJ software. Data represent means ± SEM. (F) Expression of MAdCAM-1 in spleens of indicated mutant mice measured by real-time PCR. Data represent means ± SD; n = 3, *P < .05.

Maintenance of spleen microarchitecture is primarily dependent on TNF expressed by B cells with distinct contribution of T-cell–derived TNF. (A) Mice were immunized with i.p. SRBCs and spleen sections analyzed on day 8. B-TNF KO mice display reduced GCs, lack of polarized B-cell follicles, and disruption of FDC networks. Note that some FDC- and PNA-positive cells are present in the spleens of B-TNF KO mice (shown by arrows), while no FDCs and no polarized B-cell follicles are observed in spleen of T,B-TNF KO mice. Original magnification is ×200. Representative images from 1 of 3 independent experiments (n = 5 mice per group) are shown. (B) Quantification of GC and FDC areas from panel A. Data represent means ± SEM; *P < .05, **P < .01, ***P < .001. (C) Analysis of germinal center B cells. Mice were immunized with i.p. SRBCs, and GL7+ GC B cells were analyzed by flow cytometry on day 8 (represents 1 of 2 independent experiments, 3 mice per group). Data represent means ± SD; *P < .05. (D) MZ structure is dependent on TNF produced by B, T, and other cell types. Mice were immunized with i.p. SRBCs, and spleen sections were analyzed on day 8. Frozen spleen sections were labeled with anti-ER-TR7, anti-CD169, and anti-MAdCAM-1 antibodies. Note the increased breadth of the MZ (ER-TR7 labeling), reduced and disorganized MAdCAM-1, and CD169 staining in B-TNF, T,B-TNF, and TNF KO mice (marked by black arrows). Microphotographs were taken with an Olympus BX41 microscope with a Zeiss Axiocam digital camera; images were captured with Zeiss Imaging AxioVision (Version 4.7) and processed using Adobe Photoshop CS3 (Version 10.0). Original magnification is ×200. Representative images from 1 of 3 independent experiments (n = 5 mice per group) are shown. (E) Quantification of MZ size on panel D (ER-TR7 labeling). MZ size (shown by arrows) was quantified using NIH ImageJ software. Data represent means ± SEM. (F) Expression of MAdCAM-1 in spleens of indicated mutant mice measured by real-time PCR. Data represent means ± SD; n = 3, *P < .05.

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