Contribution of Classic and Alternative Effector Pathways in Peanut-Induced Anaphylactic Responses Joost J. Smit1,2*, Karina Willemsen1, Ine Hassing 1 , Danielle Fiechter1, Gert Storm3, Louis van Bloois3, Jeanette H. W. Leusen 4 , Maarten Pennings2,4, Dietmar Zaiss5, Raymond H. H. Pieters1 1 Immunotoxicology, Institute for Risk Assessment Sciences, Utrecht University, Utrecht, The Netherlands, 2 Utrecht Centre for Food Allergy, Utrecht, The Netherlands, 3 Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Utrecht, The Netherlands, 4 Utrecht University Medical Center, Utrecht, The Netherlands, 5 Faculty of Veterinary Sciences, Utrecht University, Utrecht, The Netherlands Abstract Food allergy affects approximately 5% of children and is the leading cause of hospitalization for anaphylactic reactions in westernized countries. However, the pathways of anaphylaxis in food allergy are still relatively unknown. We investigated the effector pathways of allergic and anaphylactic responses of different strains of mice in a clinical relevant model of peanut allergy. C3H/HeOuJ, C57BL/6 and BALB/c mice were sensitized by intragastric peanut extract and challenged by intragastric or intraperitoneal injection of peanut. Peanut-specific T cell responses, IgE, IgG1 and IgG2a and mucosal mast cell degranulation were induced to different extent in C3H/HeOuJ, C57BL/6 and BALB/c mice. Interestingly, anaphylactic symptoms after systemic challenge were highest in C3H/HeOuJ followed by C57BL/6 but were absent in BALB/c mice. Mechanistic studies showed that the food allergic systemic anaphylaxis was dependent on platelets, FcRc and mast cells, and partially dependent on platelet activating factor and monocytes/macrophages, depending on mouse strain. These data demonstrate that in three mouse strains, components of the classic and alternative anaphylactic cascade are differently expressed, leading to differential outcomes in parameters of allergic disease and food induced systemic anaphylaxis. Citation: Smit JJ, Willemsen K, Hassing I, Fiechter D, Storm G, et al. (2011) Contribution of Classic and Alternative Effector Pathways in Peanut-Induced Anaphylactic Responses. PLoS ONE 6(12): e28917. doi:10.1371/journal.pone.0028917 Editor: Jacques Zimmer, Centre de Recherche Public de la Sante´ (CRP-Sante´), Luxembourg Received August 24, 2011; Accepted November 17, 2011; Published December 14, 2011 Copyright: 2011 Smit et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: The authors have no support or funding to report. Competing Interests: The authors have declared that no competing interests exist. * E-mail: j.j.smit@uu.nl Introduction Food allergy is generally defined as an adverse health effect arising from a specific immune response that occurs reproducibly on exposure to a given food. Although other food allergies such as milk allergy are generally outgrown in children, peanut allergy is persistent to adulthood [1,2]. Even more concerning is that peanut and tree nut allergy account for more than half of all hospitalizations for anaphylaxis [3]. Despite that diagnosis of food allergy relies strongly on the detection of specific IgE and mast cell-mediated skin-prick testing, it is argued that allergic reactions may occur independently of antigen-specific IgE and mast cells [4].Already in the 70’s it was demonstrated that human anaphylaxis could be mediated by IgG antibodies [5]. More recently, two pathways of systemic anaphylaxis have been demonstrated in mice: a classical pathway involving IgE, FceRI, mast cells and histamine versus an alternative pathway mediated by IgG, FccRIII, neutrophils, macrophages, basophils and platelet activating factor (PAF) [6,7]. Interestingly, PAF was associated with the severity of anaphylaxis in humans [8], which was confirmed in a mouse study [9]. Nonetheless, most mouse studies looking at the role of the alternative pathway of anaphylaxis did not use relevant food allergens, or used mice preconditioned for responsiveness by using vasoactive mediators [10,11,12]. Therefore, until now no evidence exists for a role of alternative pathway in food allergy and food-induced anaphy- lactic responses. Without doubt, animal models have contributed to the insight in the mechanisms of oral sensitization to food proteins. Generally, C3H/HeJ mice or C3H/HeOuJ mice are used [13,14,15,16,17], but other mouse strains, including BALB/c or C57BL/6 have been used in food allergy models as well [18,19]. To assess the relevance of both pathways of anaphylaxis, we compared food allergic responses in C3H/HeOuJ, C57BL/6 and BALB/c mice. Marked differences were already observed in the level of oral sensitization to peanut, but most pronounced differences were observed with regard to anaphylactic responses in these mouse strains. These responses involved a variable, genetically determined combination of components of the classical and the alternative pathway of systemic anaphylaxis. This finding is of relevance to the human situation where inter-individual differences may be the cause of sometimes inconclusive diagnosis of and limited success of therapeutic approaches for food allergy. Results Specific antibody and T cell responses and mucosal mast cell degranulation differ considerably in different mouse strains First, we compared antibody responses in C3H/HeOuJ, C57BL/6 and BALB/c mice intragastrically exposed to peanut extract (PE) with cholera toxin (CT). Oral PE oral sensitization led to the induction of PE-specific IgA, IgG1, IgG2a and IgE in all mouse strains (figure 1). However, the levels of these antibodies PLoS ONE | www.plosone.org 1 December 2011 | Volume 6 | Issue 12 | e28917