Supplementary Material for: Regulatory T Cells Control Type I Food Allergy to Beta-Lactoglobulin in Mice

<i>Background:</i> Regulatory T cells contribute to peripheral immune tolerance, yet their ability to control immediate-type hypersensitivity (ITH) reactions involved in IgE-mediated food allergy is still poorly documented. <i>Objectives:</i> We investigated in mice whether CD4+CD25+ regulatory T cells could control ITH to β-lactoglobulin (BLG), a major allergen in cow’s milk. <i>Methods:</i> C3H/HeOuJ mice were sensitized by repeated oral gavage with BLG plus cholera toxin as adjuvant and orally challenged with BLG alone to elicit allergic symptoms. Mice were treated with the anti-CD25 mAb (PC61) before sensitization. Oral sensitization (afferent phase of ITH) was assessed by production of BLG-specific serum antibodies and Th1/Th2-type cytokines by specific CD4+ T cells in mesenteric lymph nodes. ITH was elicited by oral BLG challenge (efferent phase of ITH) and we monitored symptom scores, numbers and function of intestinal mast cells and serum level of the mucosal mast cell protease mMCP-1. <i>Results:</i> Upon oral BLG challenge, orally sensitized mice developed only mild clinical signs. Anti-CD25 mAb-treated mice exhibited enhancement of both BLG-specific CD4+ T cell priming with IL-4, IL-5, IL-13 and IFN-γ production and total IgE, and BLG-specific IgE, IgG1 and IgG2a in serum. Anti-CD25 mAb treatment caused more severe symptoms upon BLG challenge, which correlated with enhanced serum levels of the mucosal mast cell protease mMCP-1. <i>Conclusions:</i> These data document that constitutive CD4+CD25+ regulatory T cells alleviate clinical signs of ITH to dietary BLG by modulating the priming of BLG-specific T and B cell responses during oral sensitization and enhancing mast cell degranulation.