Activation of beta-catenin in dendritic cells regulates immunity versus tolerance in the intestine

Abstract

Dendritic cells (DCs) play a vital role in initiating robust immunity against pathogens as well as maintaining immunological tolerance to self antigens. However, the intracellular signaling networks that program DCs to become tolerogenic remain unknown. We report here that the Wnt-beta-catenin signaling in intestinal dendritic cells regulates the balance between inflammatory versus regulatory responses in the gut. beta-catenin in intestinal dendritic cells was required for the expression of anti-inflammatory mediators such as retinoic acid-metabolizing enzymes, interleukin-10, and transforming growth factor-beta, and the stimulation of regulatory T cell induction while suppressing inflammatory effector T cells. Furthermore, ablation of beta-catenin expression in DCs enhanced inflammatory responses and disease in a mouse model of inflammatory bowel disease. Thus, beta-catenin signaling programs DCs to a tolerogenic state, limiting the inflammatory response.

Fig. 1

β-catenin signaling is constitutively active in intestinal APCs and regulates the induction of T_reg cells and effector T cells. (A) Expression of β-galactosidase (representing β-catenin activity) by intestinal DCs and macrophages from SI-LP and LI-LP of TCF-reporter mice. (B and C) Fluorescence-activated cell sorting (FACS) plots representing percentages of CD4⁺ T cells positive for FoxP3 (T_reg cells), IL-17 (TH17), and IFN-γ (TH1) isolated from SI-LP, LI-LP, ceacum, and spleen of β-cat^fl/fl and β-cat^DC−/− mice. (D and E) Intracellular expression of FoxP3, IL-17, and IFN-γ in naïve CD4⁺OT-II T cells stimulated to differentiate in vitro by intestinal APCs (CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻ CD11b⁺ macrophages) isolated from β-cat^fl/fl or β-cat^DC−/− mice, in the presence of TGF-β (1 ng/ml). Numbers in FACS plots represent percentage of cells positive for the indicated protein. Data are from one experiment representative of three.

Fig. 2

β-catenin signaling in intestinal DCs promotes the expression of Raldh and suppresses the expression of proinflammatory cytokines. (A) Expression of Aldh1a1 and Aldh1a2 mRNA in CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl or β-cat^DC−/− mice. (B) Expression of Raldh protein by CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻ CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl or β-cat^DC−/− mice, as assessed by in-tracellular staining and flow cytometry. (C) Expression of Il-10, Tgf-β1, Il-23a, and Il-6 mRNAs in CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl or β-cat^DC−/− mice. (D) Cytokine concentrations in the supernatants of CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl or β-cat^DC−/− mice after 24 hours. *P < 0.05; **P < 0.005. Error bars indicate mean ± SEM. Data are representative of three experiments.

Fig. 3

β-catenin activation in intestinal DCs is independent of commensals and induced by Wnt-signaling. (A and B) FACS plots representing the percentage of CD4⁺ T cells that express (A) FoxP3, IL-17, and (B) IFN-γ isolated from SI-LP of β-cat^fl/fl or β-cat^DC−/− mice treated with (Atx) or without (None) antibiotics. (C) FACS plots showing the intracellular expression levels of β-galactosidase or β-catenin protein in CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻CD11b⁺ macrophages isolated from SI-LP of TCF-reporter mice treated with or without antibiotics. (D and E) mRNA expression of (D) Fzd receptors and (E) Wnt ligands in CD11c⁺CD11b and CD11c⁺CD11b⁺ DCs and CD11c CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl mice. (F) mRNA expression levels of Wnt–β-catenin target genes Wisp1, Wisp2, and Axin1 in CD11c⁺CD11b⁻ and CD11c⁺CD11b⁺ DCs and CD11c⁻CD11b⁺ macrophages isolated from SI-LP of β-cat^fl/fl mice. Error bars indicate mean ± SEM. Data are from one experiment representative of three.

Fig. 4

β-catenin signaling in LP-DCs regulates intestinal homeostasis. (A) Percent body weight of β-cat^fl/fl or β-cat^DC−/− mice treated with 2% DSS for 7 days. Error bars indicate mean ± SD. (B) FACS plot representing percentage of CD4⁺ cells positive for IL-17 and IFN-γ isolated on day 8, from Ceacum and LI-LP of β-cat^fl/fl or β-cat^DC−/− mice treated with DSS for 6 days. Data are from one experiment representative of three. (C) Histopathological changes in colon tissue from β-cat^fl/fl or β-cat^DC−/− mice treated with or without 2% DSS treatment for 7 days. Areas of interest are infiltrations, edema (yellow arrows), globlet cells (black arrows), and basement membrane (green arrows). Images are 10× original magnification.