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Background In the house dust mite (HDM) Dermatophagoides pteronyssinus, Der p 1, 2, 5, 7, 21, and 23 have been identified as the most important allergens. The aim of this study was to define hypoallergenic peptides derived from the sequences of the six allergens and to use the peptides and the complete allergens to study antibody, T cell, and cytokine responses in sensitized and nonsensitized subjects. Methods IgE reactivity of HDM‐allergic and non‐HDM‐sensitized individuals to 15 HDM allergens was established using ImmunoCAP ISAC technology. Thirty‐three peptides covering the sequences of the six HDM allergens were synthesized. Allergens and peptides were tested for IgE and IgG reactivity by ELISA and ImmunoCAP, respectively. Allergenic activity was determined by basophil activation. CD4+ T cell and cytokine responses were determined in PBMC cultures by CFSE dilution and Luminex technology, respectively. Results House dust mite allergics showed IgE reactivity only to complete allergens, whereas 31 of the 33 peptides lacked relevant IgE reactivity and allergenic activity. IgG antibodies of HDM‐allergic and nonsensitized subjects were directed against peptide epitopes and higher allergen‐specific IgG levels were found in HDM allergics. PBMC from HDM‐allergics produced higher levels of IL‐5 whereas non‐HDM‐sensitized individuals mounted higher levels of IFN‐gamma, IL‐17, pro‐inflammatory cytokines, and IL‐10. Conclusion IgG antibodies in HDM‐allergic patients recognize peptide epitopes which are different from the epitopes recognized by IgE. This may explain why naturally occurring allergen‐specific IgG antibodies do not protect against IgE‐mediated allergic inflammation. A mix of hypoallergenic peptides containing T cell epitopes of the most important HDM allergens was identified., A mix of thirty‐one hypoallergenic peptides containing T cell epitopes of the six most important house dust mite (HDM) allergens was identified. Natural IgG antibodies of HDM‐allergic patients recognize different epitopes than IgE antibodies. Hypoallergenic T cell epitope‐containing peptides comprising the important HDM allergens may be used for therapy and prevention of HDM allergy.

IgE-mediated allergy to birch pollen affects more than 100 million patients world-wide. Bet v 1, a 17 kDa protein is the major allergen in birch pollen responsible for allergic rhinoconjunctivitis and asthma in birch pollen allergic patients. Allergen-specific immunotherapy (AIT) based on therapeutic administration of Bet v 1-containing vaccines is an effective treatment for birch pollen allergy but no allergen-specific forms of prevention are available. We developed a mouse model for IgE sensitization to Bet v 1 based on subcutaneous injection of aluminum-hydroxide adsorbed recombinant Bet v 1 and performed a detailed characterization of the specificities of the IgE, IgG and CD4+ T cell responses in sensitized mice using seven synthetic peptides of 31-42 amino acids length which comprised the Bet v 1 sequence and the epitopes recognized by human CD4+ T cells. We then demonstrate that preventive systemic administration of a mix of synthetic non-allergenic Bet v 1 peptides to 3-4 week old mice significantly reduced allergic immune responses, including IgE, IgG, IgE-mediated basophil activation, CD4+ T cell and IL-4 responses to the complete Bet v 1 allergen but not to the unrelated major grass pollen allergen Phl p 5, without inducing Bet v 1-specific allergic sensitization or adaptive immunity. Our results thus demonstrate that early preventive administration of non-allergenic synthetic T cell epitope-containing allergen peptides could be a safe strategy for the prevention of allergen-specific IgE sensitization.

Additional file 1: Table S1. Clinical and demographic data of 150 mugwort pollen-allergic individuals sIgE against mugwort extract (w6), Art v 1 (w231) and Art v 3(w233) determined by ImmunoCAP, ND, not determined. AS asthma; AR allergic rhinitis; C, conjunctivitis; E, eczema. I-1, I-2, I-3, I-4 indicates the patients serum used in ImmunoCAP inhibition assay belonging to four groups of different sensitization patterns (1, Art v 1 and Art v 3 positive; 2, Art v 1 positive, Art v 3 negative; 3, Art v 1 negative, Art v 3 positive; 4, Art v 1 and Art v 3 negative). The 82 patients reported in previously studies17, 20, 28 are indicated by an asterisk. Table S2. GenBank accession numbers for three allergen groups in seven Artemisia species.Table S3. Productivity of the three group allergens purified by specific mAb. Table S4. ImmunoCAP IgE characterization of serum pools from five areas. Figure S1. Six Artemisia species collected from China. Figure S2. SDS-PAGE of natural purified Art v 1, Art v 2 and Art v 3 homologous allergens from six Chinese Artemisia species. a, natural Art v 1 homologues purified by specific mAb A7-G4-E6; b, natural Art v 2 homologues purified by specific mAb C9-C1 shown in six different gels; c, natural Art v 3 homologues purified by specific mAb A2-B8. Figure S3. Mass spectra of natural purified Art v 1(a), Art v 2(b) and Art v 3(c) homologues. The peptides verified by LC–MS/MS are shown in red and highlighted. Figure S4. ELISA quantification of three allergen components in Artemisia spp. pollen. a, Chinese silver mugwort (A. argyi) pollen extract (ArE) in SDS gel and reaction to polyclonal antibodies (pAb) by Western blot; b, ELISA standard curve for Art v 1 allergen (mAb A7-G4-E6 and rabbit pAbs); c, ELISA standard curve for Art v 2 homologous allergen (mAb C9-C1 with rabbit pAbs); d, ELISA standard curve for Art v 3 homologous allergen with two mAbs (mAbs A2-B8 and A9-G10) with representative different isoforms. Figure S5..Inhibition of IgE binding to Art v 1 and Art v 3 with seven pollen extracts using two serum pools. a: inhibition ELISA coated with rArt v 1.0101, serum pool from Datong, Shanxi (nArt v 1: 11 kUA/l); b: inhibition ELISA coated with rArt v 1.0101, serum pool from Yantai, Shandong (nArt v 1: 6.51 kUA/l); c: inhibition ELISA coated with rArt v 3.0201, serum pool from Datong, Shanxi (CAP nArt v 3: 11.27 kUA/l); d: inhibition ELISA coated with rArt v 3.0201, serum pool from Yantai, Shandong (CAP nArt v 3: 9.65 kUA/l). Figure S6. Inhibition of sera pool from Shanxi with extract from different species at 100 μg/ml in ELISA coated with 10 μg/ml of different pollen extracts and mixture. 6-mix, mixture of six Artemisia spp. extract in the same proportions.

Background CD23 mediates IgE-facilitated allergen presentation and subsequent allergen-specific T-cell activation in allergic patients. Objective We sought to investigate key factors regulating IgE-facilitated allergen presentation through CD23 and subsequent T-cell activation. Methods To study T-cell activation by free allergens and different types of IgE–Bet v 1 complexes, we used a molecular model based on monoclonal human Bet v 1–specific IgE, monomeric and oligomeric Bet v 1 allergen, an MHC-matched CD23-expressing B-cell line, and a T-cell line expressing a human Bet v 1–specific T-cell receptor. The ability to cross-link Fcε receptors of complexes consisting of either IgE and monomeric Bet v 1 or IgE and oligomeric Bet v 1 was studied in human FcεRI-expressing basophils. T-cell proliferation by monomeric or oligomeric Bet v 1, which cross-links Fcε receptors to a different extent, was studied in allergic patients’ PBMCs with and without CD23-expressing B cells. Results In our model non–cross-linking IgE–Bet v 1 monomer complexes, as well as cross-linking IgE–Bet v 1 oligomer complexes, induced T-cell activation, which was dependent on the concentration of specific IgE. However, T-cell activation by cross-linking IgE–Bet v 1 oligomer complexes was approximately 125-fold more efficient. Relevant T-cell proliferation occurred in allergic patients’ PBMCs only in the presence of B cells, and its magnitude depended on the ability of IgE–Bet v 1 complexes to cross-link CD23. Conclusion The extent of CD23-mediated T-cell activation depends on the concentration of allergen-specific IgE and the cross-linking ability of IgE-allergen complexes.