Definition of a pool of epitopes that recapitulates the T cell reactivity against major house dust mite allergens.

House dust mites belong to the most frequent indoor allergen sources worldwide and are potent inducers of perennial asthma and rhinitis [1-3]. Major allergens in house dust are associated with mites of the genus Dermatophagoides [4]. To date, 17 groups of allergens from Dermatophogoides sp. with diverse biological functions have been described (www.allergen.org) [5, 6]. Especially group 1 and 2 allergens from Dermatophagoides pteronyssinus (Der p) and D. farina (Der f), secreted from mite bodies or excreted with their feces, are among the strongest immunogens in humans [7, 8]. Der p 1 and Der f 1 are proteins with cysteine protease activity while Der p 2 and Der f 2 contribute to allergenicity by interacting with the innate immune system [9, 10]. Recently, a new HDM allergen, Der p 23, a perithrophin-like protein was identified. It was shown to induce Immunoglobulin E (IgE)-levels comparable to Der p 1 and 2 and hence represents another major HDM allergen [11]. IgE is of particular importance for the pathogenesis of an allergic immune response. Cross-linking of IgE molecules bound to Fce-receptors on the surface of mast cells and basophil granulocytes by allergens leads to the release of histamine and other proinflammatory mediators and triggers immediate type clinical symptoms of allergic disease [12, 13]. T cells are also major contributors to allergic disease. T helper type 2 (Th2)-derived cytokines such as IL-4 and IL-13 induce immune class switching from IgG4 to IgE in B cells [14-16]. In addition to promoting the production of IgE, Th2 cells may also contribute to the immune response directly by releasing proinflammatory cytokines such as IL-5 that trigger eosinophilia. While several studies have reported several T cell epitopes derived from HDM [17-23], a comprehensive characterization of the epitopes recognized by human T cells in the context of house dust mite is still lacking. The precise definition and exact mapping of the epitopes involved, antigens of origin, and patterns of associated T helper cell responses would allow the identification of immunodominant epitopes and thereby contribute to a better understanding of pathogenic immune responses. A number of studies have shown that the frequency of allergen-specific T cells is extremely low, in the range of 1 to 6/10000 cells [22, 24]. In particular, epitope sets that allow ex vivo characterization of HDM-specific T cell responses are lacking, and most if not all data is derived from protocols that utilize in vitro re-stimulation. This issue is of relevance, since it is well known that in vitro re-stimulation can alter the phenotype of the responding cells. In this study, we analyzed patterns of immune recognition of Der p- and Der f-proteins, which are considered most dominant within mite allergens [7, 8]. We comprehensively defined T cell responses and characterized the Th1/Th2 balance in HDM-specific immune responses directed against different house dust mite allergens and epitopes. To examine whether the group 1 and 2 allergens also dominate at the T cell level, over other antigens that are subdominant in terms of IgE titers, we also included the recently described Der p 23 [11] into our studies. Based on these results we define a pool of immunodominant epitopes that allows the detection of HDM-specific T cells directly ex vivo.