Novel insights into the allergenic relationship between red meat and bovine milk

Novel insights into the allergenic relationship between red meat and bovine milk Marija Perusko1, Danijela Apostolovic2, Maria Starkhammar3, Tanja Cirkovic Velickovic4,5,6,7, Marianne van Hage2 1Innovation Center of the Faculty of Chemistry, Belgrade, Serbia 2Department of Medicine, Solna, Immunology and Allergy Unit, Karolinska Institutet and University Hospital, Stockholm, Sweden 3Department of Internal Medicine, Sodersjukhuset, Stockholm, Sweden 4Serbian Academy of Sciences and Arts, Belgrade, Serbia 5Center of Excellence for Molecular Food Sciences & Department of Biochemistry, University of Belgrade - Faculty of Chemistry, Belgrade, Serbia 6Ghent University Global Campus, Yeonsu-gu, Incheon, South Korea 7Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium Background Red meat allergy is a severe form of food allergy with delayed symptoms including anaphylaxis where the IgE antibodies are directed against a carbohydrate epitope, galactose-α-1,3-galactose (α-Gal). Many red meat allergic patients report allergic symptoms upon consumption of milk or dairy products. The aim of the project was to investigate the allergenic relationship between bovine milk and red meat at a molecular level. Methods Adults with diagnosed red meat allergy (n = 27) were recruited and their specific IgE levels to α-Gal, beef and milk were analyzed by ImmunoCAP. Milk proteins were assayed by immunoblot and inhibition ELISA for the presence of the α-Gal epitope and for the binding to red meat allergic patients’ IgE. The involvement of the carbohydrate epitope in the IgE binding to milk proteins was assessed by an inhibition assay with thyroglobulin. Basophil activation test was performed with milk and milk proteins in samples from 11 red meat allergic patients and 2 controls. Results All patients were IgE positive to milk, but the IgE levels to milk were lower than those to α-Gal or beef. Significant correlations between IgE levels to milk and α-Gal (rs=0.64, P < 0.01), as well as between milk and beef (rs=0.90, P < 0.01) were observed. Immunoblot analysis of milk proteins revealed bovine γ-globulin (BGG) as α-Gal carrier. Other milk proteins, α-lactalbumin, β-lactoglobulin, α-casein, β-casein and κ-casein were negative for the presence of α-Gal epitope. BGG was also shown to bind IgE antibodies of red meat allergic patients. Inhibition immunoblot with thyroglobulin resulted in the loss of IgE binding to BGG. Additionally, ELISA experiments showed that BGG, as well as whey proteins exert a dose-dependent inhibition of red meat allergic patients’ IgE binding to -Gal. Inhibition with raw milk and commercially available milk preparations showed that raw milk exerted a slightly higher inhibition of the IgE binding to the α-Gal epitope than the commercially available milks. Importantly, activation of red meat allergic patient’s basophils by BGG and milk was demonstrated. Conclusion BGG was identified as a major milk carrier of the -Gal epitope that bound IgE antibodies and furthermore activated basophils of red meat allergic patients. This study highlights the importance of milk as allergenic food source among the meat allergic population