Decreased cell infiltrates and IFN-γ production of liver mononuclear cells in 2-OA-BSA immunized CD1d^-/- mice.

<p>(A-C) CD1d^-/- and wild type mice were immunized with 2-OA-BSA at 0, 2, 4, 6 and 8 and sacrificed at week 12. (A) Liver total mononuclear cells (MNCs) were measured. (B) The numbers of T (CD3⁺ NK1.1^-), NKT (CD3⁺NK1.1⁺), NK (CD3^-NK1.1⁺) and B (CD19⁺) cells were measured. (C) The numbers of CD4⁺ and CD8⁺ T cells were detected. n = 16–20 per group. *, p<0.05. (D) CD1d^-/- or wild type mice were immunized with 2-OA-BSA at weeks 0, 2, and 4 and sacrificed 3 days after last immunization. IFN-γ production of liver mononuclear cells stimulated with anti-CD3 and anti-CD28 Abs for 2 days was measured by ELISA. n = 8 mice for PBS treated group, n = 13–17 mice for 2-OA-BSA immunized group. *, p<0.05; **, p<0.01.</p

Lower serum levels of IFN-γ after OCH injection.

<p>C57BL/6 mice were intravenously injected with α-GalCer, OCH, or PBS. Serum samples were collected at 2 and 18 hours after α-GalCer, OCH, or PBS injection. IFN-<i>γ</i> (A) and IL-4 (B) were measured by ELISA. n = 10 mice per group. ***, p<0.001.</p

Increased serum AMAs in mice injected with 2-OA-BSA/OCH.

<p>Wild type mice were immunized with 2-OA-BSA and α-GalCer (group name: 2-OA/a-GC), OCH (group name: 2-OA/OCH) or PBS (group name: 2-OA/PBS) at weeks 0, 2, 4, 6 and 8. At week 12, serum levels of autoantibodies to mPDC-E2 were measured by ELISA. n = 9–10 mice per group. *, p < 0.05 in 2-OA/a-GC to 2-OA/PBS; #, p < 0.05 in 2-OA/OCH to 2-OA/PBS.</p

Decreased AMAs in 2-OA-BSA immunized CD1d^-/- mice.

<p>(A) iNKT cell deficiency in CD1d^-/- mice was confirmed by staining with CD3 and CD1d tetramer. (B) CD1d^-/- and wild type mice were immunized with 2-OA-BSA at 0, 2, 4, 6 and 8 and sacrificed at week 12. Serum levels of IgM and IgG to mPDC-E2 were measured by ELISA. n = 10 mice for each group. *, p<0.05.</p

OCH administration increased cell infiltrates and activation of T cells in mice.

<p>(A) C57BL/6 mice were intravenously injected with α-GalCer, OCH, or PBS. Liver total mononuclear cells (MNC) were counted 3 days after α-GalCer, OCH, or PBS injection. n = 10–13 mice per group. ***, p<0.001. (B-E) Wild type mice were immunized with 2-OA-BSA and α-GalCer (group name: 2-OA/a-GC), OCH (group name:2-OA/OCH) or PBS (group name: 2-OA/PBS) at weeks 0, 2, 4, 6 and 8 and sacrificed at week 12. (B) Liver total mononuclear cells (MNC) were measured. (C) The numbers of T (CD3⁺ NK1.1^-), NKT (CD3⁺NK1.1⁺), NK (CD3^-NK1.1⁺) and B (CD19⁺) cells were measured. (D) The numbers of CD4⁺ and CD8⁺ T cells were detected. (E) The expression of CD69 and CD44 in CD4⁺ and CD8⁺ T cells was measured by flowcytometry. n = 9–10 mice per group. *, p<0.05; **, p<0.01; ***, p<0.001.</p

The increase of portal inflammation and fibrosis in mice injected with 2-OA-BSA/OCH.

<p>Mice were immunized with 2-OA-BSA and α-GalCer (group name: 2-OA/a-GC), OCH (group name: 2-OA/OCH) or PBS (group name: 2-OA/PBS) at weeks 0, 2, 4, 6 and 8 and sacrificed at week 12. (A) Representative stained liver sections of haematoxylin and eosin (H&E) and Masson’s trichrome stain. (B) Histopathological scores of individual livers on portal inflammation and fibrosis. 0 = no significant change, 1 = minimal, 2 = mild, 3 = moderate, and 4 = severe pathology. Individual symbols each represent a single mouse.</p

Cytokine profiles of CD4+ and CD8+ T cells in the liver and spleen of IL-2Rα^−/− mice and IL-2Rα^+/− mice.

<p>Hepatic and splenic MNC were analyzed by flow cytometry. Flow cytometry profiles are shown in (A) and (C), and quantification of frequency of IFN-γ+ and IL-17A+ cells in the CD4+ and CD8+ T cell populations in (B) and (D). Data are representative with four mice in IL-2Rα^−/− group and five mice in IL-2Rα^+/− group. * P<0.05, ** P<0.01.</p

presentation_1_The CXC Chemokine Receptor 3 Inhibits Autoimmune Cholangitis via CD8+ T Cells but Promotes Colitis via CD4+ T Cells.PDF

<p>CXC chemokine receptor 3 (CXCR3), a receptor for the C-X-C motif chemokines (CXCL) CXCL9, CXCL10, and CXCL11, which not only plays a role in chemotaxis but also regulates differentiation and development of memory and effector T cell populations. Herein, we explored the function of CXCR3 in the modulation of different organ-specific autoimmune diseases in interleukin (IL)-2 receptor deficiency (CD25^−/−) mice, a murine model for both cholangitis and colitis. We observed higher levels of CXCL9 and CXCL10 in the liver and colon and higher expression of CXCR3 on T cells of the CD25^−/− mice compared with control animals. Deletion of CXCR3 resulted in enhanced liver inflammation but alleviated colitis. These changes in liver and colon pathology after CXCR3 deletion were associated with increased numbers of hepatic CD4⁺ and CD8⁺ T cells, in particular effector memory CD8⁺ T cells, as well as decreased T cells in mesenteric lymph nodes and colon lamina propria. In addition, increased interferon-γ response and decreased IL-17A response was observed in both liver and colon after CXCR3 deletion. CXCR3 modulated the functions of T cells involved in different autoimmune diseases, whereas the consequence of such modulation was organ-specific regarding to their effects on disease severity. Our findings emphasize the importance of extra caution in immunotherapy for organ-specific autoimmune diseases, as therapeutic interventions aiming at a target such as CXCR3 for certain disease could result in adverse effects in an unrelated organ.</p

Lymphoma-like T cell infiltration is transplantable into Rag1^−/− mice.

<p>A. Flow cytometric analysis of HMNCs from donor mouse showing a TCRβ⁺NK1.1⁺CD4⁻CD8⁻ phenotype. B. Representative flow cytometric analysis of splenic and hepatic MNCs from recipient mice 6 weeks post-transfer. C, Intracellular IFN-γ and IL-2 production. D. H&amp;E stained spleen and liver sections from Rag1^−/− recipient mice 6 weeks post-transfer of 2×10⁴ or 2×10⁵ HMNCs from inbred dnTGFβRII mice with lymphomatous lesions. E. Total HMNCs in Ly5.1Rag1^−/− recipient mice six weeks post-transfer. Ly5.1Rag1^−/− mice were adoptively transferred with 2×10⁴ or 2×10⁵ hepatic mononuclear cells from inbred dnTGFβRII mice with (n = 4) or without (n = 3) lymphomatous lesions, respectively.</p

Comparison of CDR3 region of the TCR β family between dnTGFβRII and dnTGFβRII IL-6^−/− mice.

<p>The arrows indicate clonal expansion of specific Vβ. C57B6 mice were used as negative control. With this technique, if there is no detectable T cell expansion within a Vβ spectrum, a Gaussian distribution of CDR3 lengths is observed. In contrast, clonal expansions are observed as a perturbation of this Gaussian distribution.</p