In vivo demonstration of T lymphocyte migration and amelioration of ileitis in intestinal mucosa of SAMP1/Yit mice by the inhibition of MAdCAM-1

Abstract

The aetiology of Crohn's disease (CD) remains unknown. Since SAMP1/Yit mice have been reported to develop CD-like spontaneous enteric inflammation, such mice have been studied as an animal model of CD. In this study, using this model we examined T lymphocyte migration in microvessels of intestinal mucosa in vivo and the expression of adhesion molecules by immunohistochemistry. Fluorescence-labelled T lymphocytes isolated from AKR/J (control) mice were injected into the tail veins of recipient mice, and T lymphocyte migration in the postcapillary venules of Peyer's patches, submucosal microvessels, and villus capillaries of the terminal ileum was monitored using an intravital microscope. Adhesion of T lymphocytes was significantly increased in 35 week old SAMP1/Yit mice compared with that in AKR/J or 15 week old SAMP1/Yit mice. Immunohistochemical study showed increased infiltration of CD4, CD8 and beta7-integrin-positive cells and increased expression of MAdCAM-1 and VCAM-1 in the terminal ileum of SAMP1/Yit mice. Antibodies against MAdCAM-1 and VCAM-1 significantly inhibited adhesion of T lymphocytes to microvessels of the terminal ileum, and anti-MAdCAM-1 antibody showed stronger suppressive effect than the anti-VCAM-1 antibody. Periodical administration of anti-MAdCAM-1 antibody twice a week for 7 weeks significantly ameliorated ileitis of SAMP1/Yit mice, but submucosal hypertrophy was not significantly suppressed. Anti-VCAM-1 antibody treatment failed to show significant resolution of ileitis. In addition, anti-MAdCAM-1 antibody treatment also attenuated established ileitis. The results demonstrate that, although MAdCAM-1 and VCAM-1 play an important role in T lymphocyte-endothelial cell interactions in SAMP1/Yit mice, MAdCAM-1 may be a more appropriate target for therapeutic modulation of chronic ileitis.

Fig. 1

Immunohistochemical study of MAdCAM-1 and VCAM-1 expression in the intestinal mucosa. (a) MAdCAM-1 expression of AKR/J mice on the vessel near Peyer's patches (b) MAdCAM-1 expression of SAMP1/Yit mice demonstrating the increased MAdCAM-1 expression (c) VCAM-1 expression of AKR/J mice was weak in the lamina propria. (d) VCAM-1 positive vessels of SAMP-1/Yit mice were increased mainly in the submucosal area compared with those in AKR/J mice. Bar: 100 µm (objective lens × 20).

Fig. 2

Representative photographs of adherence of CFDSE-labelled T lymphocytes in PCVs of Peyer's patches of AKR/J mice or SAMP1/Yit mice observed from the serosal side under an intravital microscope at 50–60 min after infusion of labelled T lymphocytes. (a) In AKR/J mice, there were a few adhered lymphocytes in PCVs of Peyer's patches. (b) On the other hand, in 35 week old SAMP1/Yit mice, there were a significant number of adhered lymphocytes. (c) Anti-MAdCAM-1 antibody significantly decreased the number of adhered T lymphocytes. (d) Anti-VCAM-1 antibody also decreased the number of adhered T lymphocytes (objective lens × 10).

Fig. 3

Time courses of adhesion of T lymphocytes in the intestinal mucosa (a) in PCV of Peyer's patches, (b) in submucosal venules and (c) in villus microvessels. *P < 0·01 versus AKR/J mice and 15 weeks old SAMP1/Yit mice. n = 5 in each group.

Fig. 4

Effects of pretreatment with anti- MAdCAM-1 MAb or anti-VCAM-1 MAb on adherence of T lymphocytes to PCVs of Peyer's patches, submucosal venules, and microvessels of villus mucosa at 60 min after infusion of CFSE-labelled T lymphocytes. All results are expressed as percentages of those in SAMP1/Yit mice with control rat IgG. Values are means ± SE of 5 animal experiments. *P < 0·05 compared with values of SAMP/Yit mice with control rat IgG. #P < 0·05 compared with anti-VCAM-1 MAb.

Fig. 5

Representative H&E-staining of terminal ileum of SAMP-1/Yit mice after the treatment with (a) control IgG and (b) anti-MAdCAM-1 MAb. Anti-MAdCAM-1 MAb treatment suppressed villus atrophy and infiltration of inflammatory cells in the ileal mucosa. Bar 100 µm (objective lens × 20).