Ovalbumin-specific IgE modulates ovalbumin-specific T-cell response after repetitive oral antigen administration Nemuko Omata, MD, Yusei Ohshima, MD,

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Ovalbumin-specific IgE modulates ovalbumin-specific T-cell response after repetitive oral antigen administration Nemuko Omata, MD, Yusei Ohshima, MD, PhD, Motoko Yasutomi, MD, Akiko Yamada, MD, Hajime Karasuyama, MD, PhD, Mitsufumi Mayumi, MD, PhD Journal of Allergy and Clinical Immunology Volume 115, Issue 4, Pages 822-827 (April 2005) DOI: 10.1016/j.jaci.2004.12.1121 Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 1 Effects of oral OVA administration on OVA-specific cell proliferation. Mononuclear cells purified from the spleen (A), Peyer's patches (B), and mesenteric lymph nodes (C) of OVA-fed or PBS-fed OVA-TCR/IgE-Tg mice and OVA-TCR-Tg littermates were restimulated with the indicated concentrations of OVA. Shown are the means ± SDs of 12 independent experiments. ∗P < .05; ∗∗P < .01. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 2 Effects of oral antigen administration on the number of OVA-specific TCR-bearing T cells in the spleen. The percentage (A) and the absolute number (B) of KJ1-26+ T cells in the spleen of OVA-fed or PBS-fed OVA-TCR/IgE-Tg mice and OVA-TCR-Tg littermates were determined. The bars indicate the median of 5 pooled independent experiments. ∗P < .05; ∗∗P < .01. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 3 Oral OVA administration induces T-cell anergy. TCR expression levels of splenic T cells were determined by gating KJ1-26+ T cells (A). Splenic CD4+ T cells were restimulated with OVA and irradiated splenocytes of nontreated wild-type mice (B). Splenocytes were restimulated with OVA in the presence of recombinant IL-2 (C). Shown are the means ± SDs of 7 experiments. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 4 Oral OVA administration modulated cytokine-producing profiles of splenocytes. The splenocytes of OVA-fed or PBS-fed OVA-TCR/IgE-Tg mice and OVA-TCR-Tg littermates were restimulated with 1000 μg/mL OVA. Supernatants were tested by ELISA to determine the production of IL-4 (A), IFN-γ (B), IL-10 (C), and TGF-β1 (D). The bars indicate the median of 7 pooled independent experiments. ∗P < .05; P < .01. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 5 Roles of TGF-β1 and IL-10 in the hyporesponsiveness of orally tolerated splenocytes. Splenocytes of the fed mice were restimulated with 200 μg/mL OVA in the presence or absence of antibodies as shown. The data indicate the means ± SDs of triplicate cultures. Similar results were observed in 3 independent experiments. ∗P < .05; ∗∗P < .01. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions

Fig 6 Effects of oral OVA administration on the frequency of CD25+CD4+ T cells in the spleen. OVA-TCR/IgE-Tg mice and OVA-TCR-Tg littermates were fed OVA or control PBS. The percentage of CD25+CD4+ T cells in the splenocytes was determined by flow cytometry. Values represent the percentages of CD25+CD4+ cells. This experiment represents 3 independent experiments with similar results. Journal of Allergy and Clinical Immunology 2005 115, 822-827DOI: (10.1016/j.jaci.2004.12.1121) Copyright © 2005 American Academy of Allergy, Asthma and Immunology Terms and Conditions