Birgit Nagl

Human blood basophils do not act as antigen-presenting cells for the major birch pollen allergen Bet v 1

Several studies in mice have recently shown that basophils can act as antigen‐presenting cells (APC) inducing Th2‐mediated immune responses against parasites or protease allergens. The aim of this study was to investigate whether human basophils function as APC for the major birch pollen allergen Bet v 1.

Pru p 3, the nonspecific lipid transfer protein from peach, dominates the immune response to its homolog in hazelnut

To cite this article: Schulten V, Nagl B, Scala E, Bernardi ML, Mari A, Ciardiello MA, Lauer I, Scheurer S, Briza P, Jürets A, Ferreira F, Jahn‐Schmid B, Fischer GF, Bohle B. Pru p 3, the nonspecific lipid transfer protein from peach, dominates the immune response to its homolog in hazelnut. Allergy 2011; 66: 1005–1013.

Oral exposure to Mal d 1 affects the immune response in patients with birch pollen allergy

BACKGROUND Antibodies and T cells specific for the major birch pollen allergen Bet v 1 cross-react with structurally related food allergens, such as Mal d 1 in apple. OBJECTIVE We sought to evaluate the effects of oral uptake of Mal d 1 on the allergen-specific immune response in patients with birch pollen allergy. METHODS Patients received 50 μg of rBet v 1 sublingually on 2 consecutive days outside of the birch pollen season. One year later, equal amounts of rMal d 1 were administered. Blood samples were collected before and after oral exposure, as well as before and after the intermediate birch pollen season. Allergen-specific IgE levels were determined by using ImmunoCAP. Proliferation of allergen-stimulated PBMCs was assessed, as well as the expression of IL-5, IL-13, IL-10, IFN-γ, and forkhead box protein 3 (Foxp3) in isolated T cells (real-time PCR). Allergen-specific T-cell lines were analyzed for epitope recognition. RESULTS Orally administered Bet v 1 transiently reduced Bet v 1-specific serum IgE levels, as well as Bet v 1- and Mal d 1-induced T-cell proliferation, and enhanced the expression of IL-5, IL-10, and Foxp3. Orally applied Mal d 1 significantly decreased Bet v 1- and Mal d 1-specific IgE levels and induced IL-5 and IL-10 but no Foxp3 expression. In contrast to Bet v 1, Mal d 1 triggered IFN-γ production and T cells with a different epitope repertoire. Inhalation of birch pollen significantly enhanced allergen-specific IgE levels, T-cell proliferation, and IL-5, IL-10, IL-13, and Foxp3 expression. CONCLUSION Two sublingual administrations of 50 μg of Mal d 1 were well tolerated and induced transient immune responses seen during peripheral tolerance development. Thus recombinant Mal d 1 might be suitable and relevant for sublingual treatment of birch pollen-related apple allergy.

Glutathione-S-Transferase: A Minor Allergen in Birch Pollen due to Limited Release from Hydrated Pollen

Background Recently, a protein homologous to glutathione-S-transferases (GST) was detected in prominent amounts in birch pollen by proteomic profiling. As members of the GST family are relevant allergens in mites, cockroach and fungi we investigated the allergenic relevance of GST from birch (bGST). Methodology bGST was expressed in Escherichia coli, purified and characterized by mass spectrometry. Sera from 217 birch pollen-allergic patients were tested for IgE-reactivity to bGST by ELISA. The mediator-releasing activity of bGST was analysed with IgE-loaded rat basophil leukaemia cells (RBL) expressing human FcεRI. BALB/c mice were immunized with bGST or Bet v 1. Antibody and T cell responses to either protein were assessed. IgE-cross-reactivity between bGST with GST from house dust mite, Der p 8, was studied with murine and human sera in ELISA. The release kinetics of bGST and Bet v 1 from birch pollen were assessed in water, simulated lung fluid, 0.9% NaCl and PBS. Eluted proteins were quantified by ELISA and analysed by immunoblotting. Principle findings Only 13% of 217 birch pollen-allergic patients showed IgE-reactivity to bGST. In RBL assays bGST induced mediator release. Immunization of mice with bGST induced specific IgE and a Th2-dominated cellular immune response comparably to immunization with Bet v 1. bGST did not cross-react with Der p 8. In contrast to Bet v 1, only low amounts of bGST were released from pollen grains upon incubation in water and the different physiological solutions. Conclusion/Significance Although bGST is abundant in birch pollen, immunogenic in mice and able to induce mediator release from effector cells passively loaded with specific IgE, it is a minor allergen for birch pollen-allergic patients. We refer this discrepancy to its limited release from hydrated pollen. Hence, bGST is an example demonstrating that allergenicity depends mainly on rapid elution from airborne particles.

Recombinant Mal d 1 facilitates sublingual challenge tests of birch pollen‐allergic patients with apple allergy

It is still unclear whether allergen‐specific immunotherapy (AIT) with birch pollen improves birch pollen‐related food allergy. One reason for this may be the lack of standardized tests to assess clinical reactions to birch pollen‐related foods, for example apple. We tested the applicability of recombinant (r) Mal d 1, the Bet v 1‐homolog in apple, for oral challenge tests. Increasing concentrations of rMal d 1 in 0.9% NaCl were sublingually administered to 72 birch pollen‐allergic patients with apple allergy. The dose of 1.6 μg induced oral allergy syndromes in 26.4%, 3.2 μg in 15.3%, 6.3 μg in 27.8%, 12.5 μg in 8.3%, 25 μg in 11.1%, and 50 μg in 4.2% of the patients. No severe reactions occurred. None of the patients reacted to 0.9% NaCl alone. Sublingual administration of 50 μg of rMal d 1 induced no reactions in three nonallergic individuals. Our approach allows straight forward, dose‐defined sublingual challenge tests in a high number of birch pollen‐allergic patients that inter alia can be applied to evaluate the therapeutic efficacy of birch pollen AIT on birch pollen‐related food allergy.

Efficacy and safety of 4 months of sublingual immunotherapy with recombinant Mal d 1 and Bet v 1 in patients with birch pollen–related apple allergy

Background: Birch pollen–related apple allergy is among the most prevalent food allergies in adolescent/adult subjects and mainly results from sensitization to the major birch pollen allergen Bet v 1 and subsequent cross‐reaction with the apple protein Mal d 1. However, specific immunotherapy with birch pollen has inconsistent effects on apple allergy. Objective: We sought to compare the safety and efficacy of sublingual immunotherapy (SLIT) with 2 formulations containing either rMal d 1 or rBet v 1 on birch pollen–related apple allergy. Methods: Sixty participants with birch pollen–related apple allergy were randomized to daily sublingual application of placebo (n = 20) or 25 &mgr;g of rMal d 1 (n = 20) or rBet v 1 (n = 20) for 16 weeks. Adverse events were regularly recorded. Sublingual challenges with standardized doses of rMal d 1, skin prick tests with recombinant allergens, and measurements of allergen‐specific IgE and IgG4 antibodies were performed before and after treatment. Results: Both formulations caused comparable, mainly local adverse events. No systemic reactions occurred. Compared with the placebo and rBet v 1–treated groups, SLIT with rMal d 1 reduced rMal d 1–induced oral symptoms (P = .001 and P = .038) accompanied by longitudinally reduced rMal d 1–specific cutaneous reactions (P = .022) and enhanced IgG4/IgE ratios (P = .012). SLIT with rBet v 1 neither improved the clinical reactivity to rMal d 1 nor enhanced rMal d 1–specific IgG4/IgE ratios. Participants receiving placebo showed no allergen‐specific changes. Conclusion: Sublingual treatment with a recombinant food allergen was safe and clinically effective, as determined by using standardized challenges. We present a promising approach for the effective treatment of birch pollen–related apple allergy.

Tackling Bet v 1 and associated food allergies with a single hybrid protein

Background Allergy vaccines should be easily applicable, safe, and efficacious. For Bet v 1–mediated birch pollen and associated food allergies, a single wild‐type allergen does not provide a complete solution. Objective We aimed to combine immunologically relevant epitopes of Bet v 1 and the 2 clinically most important related food allergens from apple and hazelnut to a single hybrid protein, termed MBC4. Methods After identification of T cell epitope–containing parts on each of the 3 parental allergens, the hybrid molecule was designed to cover relevant epitopes and evaluated in silico. Thereby a mutation was introduced into the hybrid sequence, which should alter the secondary structure without compromising the immunogenic properties of the molecule. Results MBC4 and the parental allergens were purified to homogeneity. Analyses of secondary structure elements revealed substantial changes rendering the hybrid de facto nonreactive with patients’ serum IgE. Nevertheless, the protein was monomeric in solution. MBC4 was able to activate T‐cell lines from donors with birch pollen allergy and from mice immunized with the parental allergens. Moreover, on immunization of mice and rabbits, MBC4 induced cross‐reactive IgG antibodies, which were able to block the binding of human serum IgE. Conclusion Directed epitope rearrangements combined with a knowledge‐based structural modification resulted in a protein unable to bind IgE from allergic patients. Still, properties to activate specific T cells or induce blocking antibodies were conserved. This suggests that MBC4 is a suitable vaccine candidate for the simultaneous treatment of Bet v 1 and associated food allergies. Graphical abstract Figure. No Caption available.

The quantity and quality of α‐gal‐specific antibodies differ in individuals with and without delayed red meat allergy

IgG to galactose‐α‐1,3‐galactose (α‐gal) are highly abundant natural antibodies (Ab) in humans. α‐Gal‐specific IgE Ab cause a special form of meat allergy characterized by severe systemic reactions 3–7 h after consumption of red meat. We investigated 20 patients who experienced such reactions and characterized their α‐gal‐specific IgE and IgG responses in more detail.

Blocking antibodies induced by allergen-specific immunotherapy ameliorate allergic airway disease in a human/mouse chimeric model

Allergen‐specific immunotherapy (AIT) induces specific blocking antibodies (Ab), which are claimed to prevent IgE‐mediated reactions to allergens. Additionally, AIT modulates cellular responses to allergens, for example, by desensitizing effector cells, inducing regulatory T and B lymphocytes and immune deviation. It is still enigmatic which of these mechanisms mediate(s) clinical tolerance. We sought to address the role of AIT‐induced blocking Ab separately from cellular responses in a chimeric human/mouse model of respiratory allergy.

Characterization of the T cell response to Dau c 1, the Bet v 1-homolog in carrot

In contrast to other Bet v 1‐related food allergens, the major carrot allergen, Dau c 1, has been suggested to induce food allergy independently from Bet v 1. As T cells are crucial in the sensitization process, we sought to characterize the T‐cell response to Dau c 1 and its cross‐reactivity with Bet v 1.