IgE-mediated enhancement of CD4+ T cell responses in mice requires antigen presentation by CD11c+ cells and not by B cells

Abstract

IgE antibodies, administered to mice together with their specific antigen, enhance antibody and CD4(+) T cell responses to this antigen. The effect is dependent on the low affinity receptor for IgE, CD23, and the receptor must be expressed on B cells. In vitro, IgE-antigen complexes are endocytosed via CD23 on B cells, which subsequently present the antigen to CD4(+) T cells. This mechanism has been suggested to explain also IgE-mediated enhancement of immune responses in vivo. We recently found that CD23(+) B cells capture IgE-antigen complexes in peripheral blood and rapidly transport them to B cell follicles in the spleen. This provides an alternative explanation for the requirement for CD23(+) B cells. The aim of the present study was to determine whether B-cell mediated antigen presentation of IgE-antigen complexes explains the enhancing effect of IgE on immune responses in vivo. The ability of spleen cells, taken from mice 1-4 h after immunization with IgE-antigen, to present antigen to specific CD4(+) T cells was analyzed. Antigen presentation was intact when spleens were depleted of CD19(+) cells (i.e., primarily B cells) but was severely impaired after depletion of CD11c(+) cells (i.e., primarily dendritic cells). In agreement with this, the ability of IgE to enhance proliferation of CD4(+) T cells was abolished in CD11c-DTR mice conditionally depleted of CD11c(+) cells. Finally, the lack of IgE-mediated enhancemen of CD4(+) T cell responses in CD23(-/-) mice could be rescued by transfer of MHC-II-compatible as well as by MHC-II-incompatible CD23(+) B cells. These findings argue against the idea that IgE-mediated enhancement of specific CD4(+) T cell responses in vivo is caused by increased antigen presentation by B cells. A model where CD23(+) B cells act as antigen transporting cells, delivering antigen to CD11c(+) cells for presentation to T cells is consistent with available experimental data.

Figure 1. CD11c⁺ cells present IgE-antigen to CD4⁺ T cells ex vivo.

A & B) BALB/c or CD23^-/- mice were immunized with 100 µg OVA-TNP alone or together with 250 µg of IgE anti-TNP. After 4 h, spleens were removed, fractionated, and tested for ability to activate CD4⁺ OVA-specific DO11.10 T cells ex vivo. C) CD11c-DTR mice were treated with 100 ng diphtheria toxin or left untreated. After 24 hours, the mice were immunized as in A and 4 hours later spleen cells were removed and tested for ability to activate DO11.10 T cells ex vivo. A) is representative of two identical and 4 similar experiments, B) of three similar experiments and C) of 5 identical and 2 similar experiments.

Figure 2. Diphtheria toxin depletes CD11c-DTR mice of CD11c⁺ cells.

CD11c-DTR mice were treated with 100 ng diphtheria toxin or left untreated. Twenty-four hours later total spleen cells were analyzed in flow cytometry and dendritic cells were defined as CD11c⁺, MHC-II^hi cells. The numbers show % of live cells in a representative experiment with 97% depletion of dendritic cells.

Figure 3. IgE-mediated enhancement of T cell proliferation in vivo is dependent on CD11c⁺ cells.

CD11c-DTR mice, and in B) also wild type littermates, were treated with 100 ng diphtheria toxin or left untreated. The same day, the mice were transferred i.v. with 2.3−3.0×10⁶ CD4⁺ DO11.10 spleen cells. Twenty-four hours after diphtheria toxin treatment, mice were immunized i.v. with 20 µg OVA-TNP with or without 50 µg IgE anti-TNP. Three days later, spleens were analyzed for CD4⁺ KJ1-26⁺ T cells by flow cytometry (A, B) or for KJ1-26⁺ cells in confocal microscopy (C, D). A) Representative dot-plot of flow cytometry data; values represent % CD4⁺KJ1-26⁺ cells of gated lymphocytes. B) Mean values ± SEM of % CD4⁺KJ1-26⁺ of CD4⁺ cells analyzed in flow cytometry (3 mice/group) C) Mean values ± SEM of KJ1-26⁺ cells in non-consecutive sections of T cell zones analyzed by confocal microscopy (3 mice/group; 3–12 T cell zones/mouse). D) Visualization of representative T cell zones from each group: KJ1-26⁺ cells (red) and B220⁺ cells (blue). B) is representative of 4 experiments and C) of 1 experiment.

Figure 4. MHC-II incompatible B cells rescue IgE-mediated enhancement of T cell proliferation in CD23^-/- mice.

Mice were transferred with 5×10⁶ CD4⁺ DO11.10 T cells. The next day, indicated mice were transfused with B cells (spleen cells depleted of CD43⁺ cells on a MACS column; 98% CD19⁺) and subsequently immunized with 20 µg OVA-TNP, with or without 50 µg IgE anti-TNP. Three days later spleens were removed and the number of KJ1-26⁺ T cells in non-consecutive sections of T cell zones were analyzed by confocal microscopy (2–4 mice/group; 4–16 T cell zones/mouse; mean values ± SEM). Representative of two identical and one similar experiment.