Rice induced anaphylaxis
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  • These allergens are glycoproteins of about 30 kDa that are quickly and profusely released by grass pollen upon hydration; in humans they bind to IgE antibodies to initiate the allergic response
  • To discriminate between IgE specific for peptide or carbohydrate epitopes, antibody binding to CCDs was inhibited by preincubating sera with 5 µg/mL of MUXF-BSA as done in previous studies  MUXF-BSA is a synthetic glycoprotein obtained by coupling purified N-glycans from pineapple stem bromelain to BSA [MM means that it has no glucose of fucose at the surface

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  • 1. RICE-INDUCED ANAPHYLAXIS: IGE- MEDIATED ALLERGY AGAINST A 56-KDA GLYCOPROTEIN Int Arch Allergy Immunol 2012;158:9–17 DOI: 10.1159/000330641
  • 2. Food for thought! “Rice is great if you're really hungry and want to eat two thousand of something.” Mitch Hedberg
  • 3. Characters of the Study (Drama of Allergy)  Actors: Immunoglobulin E (IgE) is the main actor. ( Hired Assassin) Mr. Rice. (Villain)  Hero to be followed in another story :P
  • 4. Introduction  Rice is the seed of the monocot plant Oryza sativa.  Cereals such as wheat, barley, rye, oats, maize, and rice are reported to elicit allergic reactions. Rice is the main and most important food taken every day in Eastern Asia. The prevalence of IgE- mediated rice allergy is about 10% in atopic subjects in Japan. Rice allergy is more prominent in adults than in children.  In contrast to its wide consumption, rice allergy seems to be exceedingly rare, and only few cases of IgE-mediated hypersensitivity are published.
  • 5. Cont.  Symptoms frequently associated with rice allergy are atopic dermatitis, eczema, and asthma. Anaphylactic reactions have been reported in severe cases.  The development of food allergy is influenced by genetic factors, cultural and dietary habits, and exposure to allergens early in life  A multigene family of 14-16 kDa proteins presents the major allergens from rice seeds/grain, which show significant homology to the alpha- amylase / trypsin inhibitor family from wheat and barley. Other rice seed/grain allergens have been identified as a 33-kDa major allergen and a 60-kDa minor allergen. Most recently IgE binding cross-reactivity of rice proteins to a lipid-transfer protein from maize has been demonstrated, indicating the presence of this plant-pan allergen in rice. In addition to ingestive allergens rice presents an important aero- allergen (Ory s 1) belonging to the grass pollen group I allergens.
  • 6. Aims and objectives  Determine the high-molecular-weight allergens from rice seeds, predominantly a 56-kDa glycoprotein?
  • 7. Materials and Methods  Rice-Allergic Patient and Patient Sera  Skin Tests Skin prick tests (SPTs) were performed with commercially available allergens (pollen from grass, rye, birch, alder, hazel, beech, mugwort, cat and dog danders, house dust mites, egg, milk, herring and hazel nut. standardized food prick-to-prick test with celeriac, carrot, tomato, apple, orange, banana, fish, milk, egg yolk, egg white, rye flour, wheat flour, soya flour, guar flour, hazel, peanut, walnut, almond, mustard seed, pepper, red pepper, curry, oat, buckwheat, millet and maize. Rice seeds from basmati rice and long-grain rice were tested raw as ground powder and after cooking for 20 min. In addition, prepared extracts from basmati rice and long-grain rice, and a commercially available basmati rice extract (Squarix, Marl,Germany)
  • 8. Materials and Methods-Cont.  Determination of Specific IgE to Rice and Potentially Cross-Reacting Cereals Sera were analyzed for specific IgE antibodies to rice (f9) and glycopeptide MUXF (Ro214)  Basophil Activation Test Blood (5 ml) from the rice-allergic patient and a non-allergic control was used within 6 h of blood sampling for the basophil activation test (BAT)  SDS-PAGE and IgE Immunoblotting bound IgE antibodies were detected with alkalinephosphatase-conjugated monoclonal mouse anti-human IgE Visualization was performed by NBT/BCIP substrate  IgE Inhibition Assay (To further characterize the allergens nature)
  • 9. Results  Table(1) SPT with raw and cooked rice seeds (basmati rice and long- grain rice) and with rice extracts at 100 g/ml, prepared from basmati rice, long-grain rice and the commercially available extract from basmati rice (Squarix) ;revealed positive reactions, respectively
  • 10. Results Fig. 1. BAT: blood cells stimulated in vitro with allergens (rice extract/control antigens) were double stained with anti-IgEmand anti-CD63 monoclonal antibodies (mAb) and analyzed by flow cytometry. Histograms show gated IgE-positive cells analyzed for the activation marker CD63. Basophil activation after stimulation with basmati rice extract (Squarix) was positive in the patient (61%) and negative in the control person (1%).
  • 11. Cont.
  • 12. Subsequently fractions displaying purified proteins with an apparent molecular weight of 49 kDa (fractions 61– 67), 52 kDa (fractions 72–77), 56 kDa (fractions 83–90) and 98 kDa (fractions 170–175) were pooled. Purified proteins were subjected to IgE immunoblotting and IgE inhibition assays, and transferred to (Polyvinylidene difluoride) PVDF membranes for further specificity characterization.
  • 13. Results
  • 14. Discussion  This study, reported on a German patient who exhibited several episodes of anaphylaxis after ingestion of cooked rice. Symptoms occurred within 20 min and comprised generalized urticaria, pruritus, angioedema, dysphagia and bronchospasm. Activation of basophils was still induced at very low concentrations of rice extract (10 –5 g/ml), indicating rather high allergenic potency.  Furthermore, this study investigated IgE binding to rice protein extracts from 2 cooked rice varieties and 1 native basmati rice extract - prepared without cooking.
  • 15. Cont  Noteworthy, in all extracts the patient’s serum constantly detected a 56-kDa band. For further characterization, this 56- kDa protein and 3 additional IgE-reactive proteins of approximately 49, 52 and 98 kDa have been purified and shown to be recognized by the serum from our rice-allergic patient as well as by 3 additional sera from patients who reported hypersensitivity symptoms after rice ingestion. IgE inhibition assays showed that all 4 purified putative allergenic proteins, i.e. 49, 52, 56 and 98 kDa, express IgE-reactive glycan structures, indicating that CCD contributed to IgE binding. Interestingly, only IgE reactivity to the 56- kDa protein was not completely inhibited by pre-incubation with the MUXF- glycopeptide. Therefore, the heatstable 56-kDa protein may be the allergen responsible for the anaphylactic reactions after
  • 16. Conclusion  In summary, the authors of this study have identified 4 glycosylated proteins as potential food allergens in rice seed. Results of inhibition studies indicate that the 56-kDa glycoprotein may be the allergen responsible for the anaphylaxis after rice consumption in this patient.
  • 17. Recommendation  Since this research showed that the IgE epitopes of the 56-kDa protein are determined by both the heat-stable CCD and the protein structure.  Unfortunately, the structure of this glycosylated 56-kDa allergen has not been elucidated yet. So this is something you can consider for future studies and validation!
  • 18. Reference  Trcka J. · Schäd S.G. · Scheurer S. · Conti A. · Vieths S. · Gross G. · Trautman n A. Rice-Induced Anaphylaxis: IgE-Mediated Allergy against a 56-kDa Glycoprotein, Int Arch Allergy Immunol 2012;158:9–17