Investigations of immunogenic, allergenic and adjuvant properties of Cry1Ab protein after intragastric exposure in a food allergy model in mice.

BACKGROUND: In genetically modified (GM) crops there is a risk that the inserted genes may introduce new allergens and/or adjuvants into the food and feed chain. The MON810 maize, expressing the insecticidal Cry1Ab toxin, is grown in many countries worldwide. In animal models, intranasal and intraperitoneal immunisations with the purified Cry1Ab proteins have induced immune responses, and feeding trials with Cry1Ab-containing feed have revealed some altered immune responses. Previous investigations have primarily measured antibody responses to the protein, while investigations of clinical food allergy symptoms, or allergy promotion (adjuvant effect) associated with the Cry1Ab protein are largely missing. We aimed to investigate immunogenic, allergenic and adjuvant properties of purified Cry1Ab toxin (trypCry1Ab, i.e., trypsin activated Cry1Ab) in a mouse model of food allergy. METHOD: Female C3H/HeJ mice were immunized by intragastric gavage of 10 µg purified, trypsin activated Cry1Ab toxin (trypCry1Ab) alone or together with the food allergen lupin. Cholera toxin was added as a positive control for adjuvant effect to break oral tolerance. Clinical symptoms (anaphylaxis) as well as humoral and cellular responses were assessed. RESULTS: In contrast to results from previous airway investigations, we observed no indication of immunogenic properties of trypCry1Ab protein after repeated intragastric exposures to one dose, with or without CT as adjuvant. Moreover, the results indicated that trypCry1Ab given by the intragastric route was not able to promote allergic responses or anaphylactic reactions against the co-administered allergen lupin at the given dose. CONCLUSION: The study suggests no immunogenic, allergenic or adjuvant capacity of the given dose of trypCry1Ab protein after intragastric exposure of prime aged mice.

Cross-allergic reactions to legumes in lupin and fenugreek-sensitized mice.

Several legumes may induce allergy, and there is extensive serological cross-reactivity among legumes. This cross-reactivity has traditionally been regarded to have limited clinical relevance. However, the introduction of novel legumes to Western countries may have changed this pattern, and in some studies cross-allergy to lupin has been reported in more than 60% of peanut-allergic patients. We wanted to explore cross-reactions among legumes using two newly established mouse models of food allergy. Mice were immunized perorally with fenugreek or lupin with cholera toxin as adjuvant. The mice were challenged with high doses of fenugreek, lupin, peanut or soy, and signs of anaphylactic reactions were observed. Cross-allergic mechanisms were investigated using serum mouse mast cell protease-1 (MMCP-1), antibody responses, immunoblotting and ex vivo production of cytokines by spleen cells. Signs of cross-allergy were observed for all the tested legumes in both models. The cross-allergic symptoms were milder and affected fewer mice than the primary allergic responses. The cross-allergy was reflected to a certain extent in the antibody and T-cell responses, but not in serum MMCP-1 levels. Cross-allergy to peanut, soy, fenugreek and lupin was observed in lupin-sensitized and fenugreek-sensitized mice. Differences in serological responses between primary allergy and cross-allergy might be due to mediation through different immune mechanisms or reflect different epitope affinity to IgE. These differences need to be further investigated.

The prevalence of sensitization to lupin flour in France and Belgium: a prospective study in 5,366 patients, by the Allergy Vigilance Network.

OBJECTIVES: To determine the prevalence of sensitization to lupin flour in patients consulting allergists, in order to evaluate the risk of primary and secondary allergies to lupin. METHODS: A prospective study carried out by members of the Allergy Vigilance Network, using prick-tests with a commercial lupin flour extract in patients with various allergic symptoms. The study design classified patients into four groups: peanut allergy, current atopic disease, latent atopy, no atopy. Data were collected and analysed by Network coordinators. RESULTS: Over a two-month period, 88 French and Belgian allergists tested 5,366 patients: 2,680 children and 2,686 adults aged over 16 years. Of the 2,680 children, 11.15% presented with peanut allergy. The frequency of cross-reactivity with lupin was 17.1% for patients with peanut allergy, 2.5% for children with current atopic disease and 1.7% for healthy children with latent atopy. In the 2,686 adults, peanut allergy was diagnosed in 1.86% of patients with cross-reactivity to lupin in 14.6%. Sensitization to lupin was detected in 3.7% of patients with current atopic disease and in 1.8% of those with latent atopy. CONCLUSION: The relative frequency of latent sensitisation to lupin in patients of all ages presenting with atopic disease is a new factor indicating the likelihood of an increase in primary food allergies to lupin flour. This justifies the recent decision requiring mandatory labelling of lupin, and shows the need to inform consumers who may be unaware that this ingredient is being used increasingly. Sensitization to lupin should be searched by prick-tests in any case of peanut allergy. Prick-test to lupin may be valuable whenever a food allergy is suspected when no current food allergens have been identified.

Lupine allergy: not simply cross-reactivity with peanut or soy.

BACKGROUND: Reports of lupine allergy are increasing as its use in food products increases. Lupine allergy might be the consequence of cross-reactivity after sensitization to peanut or other legumes or de novo sensitization. Lupine allergens have not been completely characterized. OBJECTIVES: We sought to identify allergens associated with lupine allergy, evaluate potential cross-reactivity with peanut, and determine eliciting doses (EDs) for lupine allergy by using double-blind, placebo-controlled food challenges. METHODS: Six patients with a history of allergic reactions to lupine flour were evaluated by using skin prick tests, CAP tests, and double-blind, placebo-controlled food challenges. Three of these patients were also allergic to peanut. Lupine allergens were characterized by means of IgE immunoblotting and peptide sequencing. RESULTS: In all 6 patients the ED for lupine flour was 3 mg or less for subjective symptoms and 300 mg or more for objective symptoms. The low ED and moderate-to-severe historical symptoms indicate significant allergenicity of lupine flour. Two patients allergic to lupine but not to peanut displayed IgE binding predominantly to approximately 66-kd proteins and weak binding to 14- and 24-kd proteins, whereas patients with peanut allergy and lupine allergy showed weak binding to lupine proteins of about 14 to 21 or 66 kd. Inhibition of binding was primarily species specific. CONCLUSION: Lupine allergy can occur either separately or together with peanut allergy, as demonstrated by 3 patients who are cosensitized to peanut and lupine. CLINICAL IMPLICATIONS: Lupine flour is allergenic and potentially cross-reactive with peanut allergen, thus posing some risk if used as a replacement for soy flour.

Unsuspected lupin allergens hidden in food.

BACKGROUND: Lupin is a herbaceous plant from the legume family whose seed allergens usually have cross-reaction with peanut. Lupin flour is used in human nutrition because of its high nutritional and functional qualities. AIMS: The aim of this work was to detect non-specified lupin proteins contained in several manufactured foods. METHODS: Serum from a patient suffering anaphylactic episodes after ingestion of a certain brand of cookies and with oral allergy syndrome after eating chicken bouillon was used as a tracer. Lupin seeds and commercial food extracts were analyzed by SDS-PAGE, immunoblotting and immunoblotting inhibition. Lupin extract allergenicity after thermal processing was also analyzed. RESULTS: A lupin allergen with a molecular weight close to 14 kDa was detected in extracts from cookies, a chicken bouillon cube and a chicken dehydrated soup. CONCLUSIONS: The presence of unsuspected, hidden non-specified lupin sources in food labeling was demonstrated. According to the results of this study, it is important for food-allergic patients that food labels should declare all the components irrespective of their quantity.

Assessment of residual immunoreactivity in red or white wines clarified with pea or lupin extracts.

Vegetable proteins could be a suitable alternative to animal proteins in the clarification of wine, but their residues could represent a risk for subjects with food allergy or intolerance. The aim of this study was to investigate the presence of specific immunoreactivity in red and white wines treated, as must or wine, with vegetable proteins in the clarification process. The proteins considered were prepared from lupins and peas, which are not included among the allergens listed in annex Illbis of Directive 2003/89/EC. The presence of residual immunoreactivity to specific rabbit anti-lupin and anti-pea polyclonal antibodies in treated wines was assessed by electrophoresis (SDS-PAGE) and immunoblotting. Residual protein was not detectable in red wines clarified with lupin, pea or a mixture of pea and lupin proteins or in white wines clarified with pea proteins. A small number of musts treated with lupin or pea proteins and white wines treated with lupin proteins yielded equivocal results, probably because of the presence of interfering material (e.g., sugar-rich proteins from grape and yeast). The use of bentonite as a secondary clarifying agent is therefore recommended since its combination with vegetable proteins is particularly effective in removing overall protein immunoreactivity.