Interleukin-33 amplifies IgE synthesis and triggers mast cell degranulation via interleukin-4 in naïve mice

Abstract

Background: The regulation and function of IgE in healthy individuals and in antigen-naïve animals is not well understood. IL-33 administration increases serum IgE in mice with unknown mechanism. We tested the hypothesis that IL-33 provides an antigen-independent stimulus for IgE production and mast cell degranulation.

Methods: IL-33 was administered to naïve wild-type (WT), nude and ST2(-/-) , IL-4(-/-) , IL4Rα(-/-) and T-or B-cell-specific IL-4Rα(-/-) mice. IgE and cytokines were quantified by ELISA. T- and B-lymphocyte numbers and CD40L expression were determined by flow cytometry. Anaphylaxis was measured by temperature, mast cell degranulation and histamine release.

Results: IL-33 enhanced IgE production in naïve WT, T-IL-4Rα(-/-) but not in ST2(-/-) , IL-4(-/-) , IL-4Rα(-/-) or B-cell-specific IL-4Rα(-/-) mice, demonstrating IL-33 specificity and IL-4 dependency. Moreover, IL-4 was required for IL-33-induced B-cell proliferation and T-cell CD40L expression, which promotes IgE production. IL-33-induced IL-4 production was mainly from innate cells including mast cells and eosinophils. IL-33 increased mast cell surface IgE and triggered degranulation and systemic anaphylaxis in allergen-naïve WT but not in IL-4Rα(-/-) mice.

Conclusion: IL-33 amplifies IgE synthesis and triggers anaphylaxis in naïve mice via IL-4, independent of allergen. IL-33 may play an important role in nonatopic allergy and idiopathic anaphylaxis.

Figure 1

IL-33 induces IgE but not type-II cytokine production in an IL-4-dependent manner. Groups of wild-type (A), ST2^−/− (B), IL-4Rα^−/− (C) and IL-4^−/− (D) mice were injected i.p. with IL-33 (2 μg) or PBS daily for 7 days. Total serum IgE, IL-4 and IL-13 concentrations were measured by ELISA. Data are representative of three experiments, n = 5–7 mice per group, *P < 0.05, **P < 0.01 compared to PBS controls.

Figure 2

IL-33 induces type-II cytokines mainly via innate cells in naïve mice. Mice were treated with IL-33 for 7 days. Serum IL-4, IL-13 and IgE concentrations in (A) nude and wild-type (WT) mice were measured by ELISA. (B) The frequency of IL-4⁺ cells and (C) total numbers of different cell populations in the peritoneum of WT mice treated with IL-33 or PBS were determined by intracellular staining using FACScan and quantified. Data are the representative of two experiments, n = 5 mice per group, *P < 0.05, **P < 0.01 compared to the PBS control or as indicated.

Figure 3

B cells expressing IL-4Rα are required for IL-33-induced B-cell expansion and IgE production in vivo. Wild-type, IL-4Rα and B-IL-4Rα mice were injected with IL-33. The levels of serum IgE (A) and IL-4/IL-13 (B) were measured by ELISA. (C) Total cell number, T cells and B cells in the spleen were determined by differential cell count. Data are the representative of two experiments, n = 5 mice per group, *P < 0.05, **P < 0.01 compared to PBS group.

Figure 4

IL-4Rα T cells contribute to IL-33-induced IgE synthesis. Wild-type (WT), IL-4^−/− or T-IL-4Rα^−/− mice were treated with IL-33. (A) Serum IgE and (B) IL-4 and IL-13 concentrations were measured by ELISA. (C) The levels of CD40L and CD25 on CD4⁺ T cells and the number of CD4⁺ CD40L⁺, CD4⁺ CD25⁺ T cells in the spleen of WT and IL-4^−/− mice were determined by FACScan and differential counting. (D) Total CD4⁺ T and CD19⁺ B cells in the spleen of WT and IL-4^−/− mice were determined by differential cell count. Data are from two experiments, n = 6 mice per group, *P < 0.05, **P < 0.01 compared to PBS control.

Figure 5

IL-33 triggers anaphylaxis in naïve mice in the absence of allergen. Wild-type (WT), ST2^−/− or IL-4Rα^−/− mice were injected with IL-33 for 3 consecutive days. (A) The levels of peritoneal c-Kit⁺ mast cell surface IgE and FcεR1 were determined by flow cytometry using specific antibodies. (B) Skin biopsies were stained with toluidine blue to determine the number of degranulated mast cells (indicated by arrows). (C) WT, ST2^−/− or IL-4Rα^−/− mice were injected i.p. for 3 consecutive days with IL-33 or PBS, and their rectal temperature was determined for up to 50 min postinjection. (D) Mice were killed at the end of 50 min after the third IL-33 injection, and the histamine levels were determined by ELISA. Data are pooled from 3 experiments, n = 15 mice per group, *P < 0.05, **P < 0.01 compared to PBS-treated mice.

Figure 6

Schematic representation of the mechanism of IL-33-induced IgE synthesis and anaphylaxis in naive mice. IL-33 binding to ST2⁺ innate cells, such as mast cells and eosinophils, leads to mast cell proliferation and increased IL-4 synthesis. IL-4 would then activate B cells to proliferate and to produce IgE. IL-4 produced by the innate cells could also activate T cells to express CD40L that interacts with CD40 on B cells to further enhance IgE production. IgE, together with IL-33 and IL-4, would stimulate mast cells to degranulate, resulting in anaphylaxis.