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Background: Growing up on a cattle farm and consuming raw cow's milk protects against asthma and allergies. We expect a cattle‐specific protein as active component in this farm effect. Methods: Dust was collected from cattle and poultry stables and from mattresses of households. Urine was obtained from cattle, and ambient aerosols were sampled. Samples were analysed for BLG by SDS PAGE/immunoblot and mass spectrometry, and for association with metals by SEC‐ICP‐MS. PBMC of healthy donors were incubated with BLG +/− zinc, and proliferation and cytokines determined. BALB/c mice were pre‐treated intranasally with stable dust extract containing BLG or depleted of BLG, and subsequent allergy response after sensitization was evaluated on antibody and symptom level. Results: A major protein in dust from cattle farms and ambient air was identified as BLG. Urine from female and male cattle is a major source of BLG. In dust samples, BLG was associated with zinc. In vitro, zinc‐BLG provoked significantly lower proliferation of CD4+ and CD8+ cells while inducing significantly higher levels of IFN‐γ and IL‐6 than the apo‐BLG devoid of zinc. In vivo, pre‐treatment of mice with dust extract containing BLG resulted in lower allergy symptom scores to BLG and unrelated Bet v 1 than pre‐treatment with extract depleted of BLG. These in vitro and in vivo effects were independent of endotoxin. Conclusion: The lipocalin BLG is found in large amounts in cattle urine, accumulates in bovine dust samples and is aerosolized around farms. Its association with zinc favorably shapes the human cellular immune response towards Th1‐cytokines in vitro. BLG together with zinc in stable dust protects mice from allergic sensitization. BLG with its associated ligands may in an innate manner contribute to the allergy‐protective farm effect. [ABSTRACT FROM AUTHOR]

Background Methods Results Conclusion We investigated the biological function of the mould allergen Alt a 1 as a carrier of micronutrients, such as the vitamin A metabolite retinoic acid (RA) and the influence of RA binding on its allergenicity in vitro and in vivo.Alt a 1‐RA complex formation was analyzed in silico and in vitro. PBMCs from Alternaria‐allergic donors were stimulated with Alt a 1 complexed with RA (holo‐Alt a 1) or empty apo‐Alt a 1 and analyzed for cytokine production and CD marker expression. Serum IgE‐binding and crosslinking assays to apo‐ and holo‐protein were correlated to B‐cell epitope analysis. Female BALB/c mice already sensitized to Alt a 1 were intranasally treated with apo‐Alt a 1, holo‐Alt a 1 or RA alone before measuring anaphylactic response, serum antibody levels, splenic cytokines and CD marker expression.In silico docking calculations and in vitro assays showed that the extent of RA binding depended on the higher quaternary state of Alt a 1. Holo‐Alt a 1 loaded with RA reduced IL‐13 released from PBMCs and CD3+CD4+CRTh2 cells. Complexing Alt a 1 to RA masked its IgE B‐cell epitopes and reduced its IgE‐binding capacity. In a therapeutic mouse model of Alternaria allergy nasal application of holo‐Alt a 1, but not of apo‐Alt a 1, significantly impeded the anaphylactic response, impaired splenic antigen‐presenting cells and induced IL‐10 production.Holo‐Alt a 1 binding to RA was able to alleviate Th2 immunity in vitro, modulate an ongoing Th2 response and prevent anaphylactic symptoms in vivo, presenting a novel option for improving allergen‐specific immunotherapy in Alternaria allergy. [ABSTRACT FROM AUTHOR]

Background: Previously, the protective farm effect was imitated using the whey protein beta‐lactoglobulin (BLG) that is spiked with iron‐flavonoid complexes. Here, we formulated for clinical translation a lozenge as food for special medical purposes (FSMP) using catechin‐iron complexes as ligands for BLG. The lozenge was tested in vitro and in a therapeutical BALB/c mice model. Methods: Binding of iron‐catechin into BLG was confirmed by spectroscopy and docking calculations. Serum IgE binding of children allergic or tolerating milk was assessed to loaded (holo‐) versus empty (apo‐) BLG and for human mast cell degranulation. BLG and Bet v 1 double‐sensitized mice were orally treated with the holoBLG or placebo lozenge, and immunologically analysed after systemic allergen challenge. Human PBMCs of pollen allergic subjects were flow cytometrically assessed after stimulation with apoBLG or holoBLG using catechin‐iron complexes as ligands. Results: One major IgE and T cell epitope were masked by catechin‐iron complexes, which impaired IgE binding of milk‐allergic children and degranulation of mast cells. In mice, only supplementation with the holoBLG lozenge reduced clinical reactivity to BLG and Bet v 1, promoted Tregs, and suppressed antigen presentation. In allergic subjects, stimulation of PBMCs with holoBLG led to a significant increase of intracellular iron in circulating CD14+ cells with significantly lower expression of HLADR and CD86 compared to their stimulation with apoBLG. Conclusion: The FSMP lozenge targeted antigen presenting cells and dampened immune activation in human immune cells and allergic mice in an antigen‐non‐specific manner, thereby conferring immune resilience against allergic symptoms. The whey protein beta‐lactoglobulin BLG binding to micronutrients such as iron‐catechin complexes (holoBLG) was recognized less by IgE antibodies of children allergic to milk, leading to reduced mast cell degranulation. BLG also facilitated catechin‐dependent activation of the anti‐inflammatory arylhydrocarbon receptor. In mice, oral application of a holoBLG‐based lozenge which also contained zinc and vitamin A, improved the allergic symptoms of double‐sensitized mice independent of the allergens they were challenged with. Double‐sensitized mice supplemented with the holoBLG lozenge also had reduced levels of allergen‐specific antibodies and a lower frequency of activated antigen presenting cells. [ABSTRACT FROM AUTHOR]