Volume 120, Issue 5, Pages 1173-1182 (April 2001) Effect of mature lymphocytes and lymphotoxin on the development of the follicle- associated epithelium and M cells in mouse Peyer's patches  Nathalie Debard, Frédéric Sierro, Jeffrey Browning, Jean–Pierre Kraehenbuhl  Gastroenterology  Volume 120, Issue 5, Pages 1173-1182 (April 2001) DOI: 10.1053/gast.2001.22476 Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 1 Distribution of Peyer's patches along the entire small intestine in Rag-1−/− and wild-type mice. The number of Peyer's patches in both mouse strains is larger in the duodenum (3–5) and ileum (3–5) than in the jejunum (2–3). The length of the small intestine varies from 30 to 43 cm, irrespective of the mouse strain. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 2 Peyer's patches from wild-type, nude, μMT, Rag-1−/− mice, and a wild-type bone marrow–reconstituted μMT mice. B-cell–containing follicles (F) are absent in the Peyer's patches of μMT and Rag-1−/− mice, whereas interfollicular regions (I) and rudimentary subepithelial dome (S) regions are maintained. Note the reduced size of the dome epithelium of μMT and Rag-1−/− Peyer's patches compared with that of nude and wild-type Peyer's patches. Adoptive transfer of wild-type bone marrow cells into μMT mice restores B-cell follicles and the FAE size. Bars = 100 μm. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 3 FAE's differentiation markers in wild-type mice and in Rag-1−/− mice, treated or not with LTβR-Ig fusion protein. Paraffin sections were stained with anti-SC and anti–α-smooth muscle actin antibodies to visualize the poly-Ig receptor and the myofibroblasts, respectively. Note the down-regulation of SC expression in the dome epithelium of Rag-1−/− Peyer's patches as well as the absence of underlying myofibroblasts as in wild-type Peyer's patches. Treatment of Rag-1−/− mice with the LTβR-Ig fusion protein did not affect the expression pattern of SC and the distribution of myofibroblasts. Bars = 50 μm. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 4 ALP activity in whole-mount Peyer's patches from wild-type and Rag-1−/− mice with or without LTβR-Ig fusion protein. Enterocytes expressing high ALP activity are stained in red, and mature M cells lacking ALP activity appear white. Goblet cells that also lack ALP activity are selectively labeled in blue using alcian blue. The pink cells observed with this technique are considered to represent cells with a phenotype intermediate between mature enterocytes and M cells. Insets represent a high-power view of part of the domes. Bars = 50 μm. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 5 Scanning and transmission electron micrographs of the FAE from Rag-1−/− mice. The FAE of Rag-1−/− mice contains M cells recognized by their depressed surface with (A) short irregular microvilli and (B) the presence of infiltrating mononuclear cells. (Original magnifications 2500×.) Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 6 Quantitation of intestinal M cells on whole-mount preparation Peyer's patches stained for ALP. Whole mounts were stained as in Figure 4, and ALP-negative cells were counted. (A) The abundance of M cells (% of the total epithelial cells within FAE) was significantly enhanced (P < 0.05) in Rag-1−/− mice compared with wild-type mice. (B) Six weeks of treatment with the LTβR-Ig fusion protein resulted in a significant reduction (P < 0.05) in frequency of M cells. Two to 5 mice per group were used to score M cells. Values are given as means ± SD. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions

Fig. 7 Immunohistochemical analysis of frozen sections of Peyer's patches from Rag-1−/− mice treated or not with LTβR-Ig fusion protein. Tissue sections were stained for DCs (CD11c), FDCs (FDC-M1), and HEVs (MadCAM-1). FDC-like clusters were present in Rag-1−/− mice treated with polyclonal human IgG, whereas they were lost in mice that received the LTβR-Ig fusion protein. The number of DCs in SED (arrows) was reduced in LTβR-Ig–treated mice compared with Ig-injected control mice. Bars = 50 μm. Gastroenterology 2001 120, 1173-1182DOI: (10.1053/gast.2001.22476) Copyright © 2001 American Gastroenterological Association Terms and Conditions