Faculty Bibliography

PURPOSE OF REVIEW/OBJECTIVE:To review recent advances in the area of food allergen processing and the effect on protein allergenicity. RECENT FINDINGS/RESULTS:Heating generally decreases protein allergenicity by destroying conformational epitopes. In peanut and shrimp, heat-induced Maillard reaction (glycation) may increase allergenicity. The majority of milk and egg-allergic children tolerate extensively heated (baked with wheat matrix) milk and egg. Introduction of extensively heated milk and egg proteins is associated with decreasing sizes of skin prick test wheals and increasing serum food-specific IgG4 levels. SUMMARY/CONCLUSIONS:Heating and other methods of food processing have different effects on food allergens, even those contained in the same complex food. Structural homology does not reliably predict the effect of processing on allergenicity, and individual food allergens have to be tested. Interactions with other proteins, fat, and carbohydrates in the food matrix are complex and poorly understood. Introduction of extensively heated milk and egg proteins into the diet of allergic children may represent an alternative approach to oral tolerance induction. Better characterization of these aspects of food allergy is critical for elucidation of food protein interactions with the gut-associated lymphoid tissue, the ability to induce IgE sensitization, the potential to trigger hypersensitivity reactions, and different clinical phenotypes of food allergy with regard to severity and persistence.

Oral food challenges are procedures conducted by allergists/immunologists to make an accurate diagnosis of immediate, and occasionally delayed, adverse reactions to foods. The timing of the challenge is carefully chosen based on the individual patient history and the results of skin prick tests and food specific serum IgE values. The type of the challenge is determined by the history, the age of the patient, and the likelihood of encountering subjective reactions. The food challenge requires preparation of the patient for the procedure and preparation of the office for the organized conduct of the challenge, for a careful assessment of the symptoms and signs and the treatment of reactions. The starting dose, the escalation of the dosing, and the intervals between doses are determined based on experience and the patient's history. The interpretation of the results of the challenge and arrangements for follow-up after a challenge are important. A negative oral food challenge result allows introduction of the food into the diet, whereas a positive oral food challenge result provides a sound basis for continued avoidance of the food.

BACKGROUND:Cockroach is an important allergen in inner-city asthma. The diagnosis and treatment of cockroach allergy has been impeded by the lack of standardized cockroach extracts. OBJECTIVE:We investigated the utility of a mediator release assay based on rat basophil leukemia (RBL) cells for comparing the potency of German cockroach extracts. METHODS:RBL cells (line 2H3) transfected with human FcepsilonRI were passively sensitized with sera from subjects with cockroach allergy and stimulated with serial dilutions of 3 commercial cockroach extracts (1:10 weight/volume). In addition, the in-house prepared extract was tested in separate experiments with pooled sera that produced optimal performance in the RBL assay. N-hexosaminidase release (NHR) was used as a marker of RBL cell degranulation and was examined in relation to the intradermal skin test (ID(50)EAL) and serum cockroach-specific and total IgE levels. RESULTS:The median cockroach-specific IgE concentration in 60 subjects was 0.72 kU(A)/L (interquartile range, 0.35-2.97 kU(A)/L); 19 sera (responders) produced a minimum 10% NHR to more than 1 extract. Responders had higher median cockroach-specific IgE (7.4 vs 1.0 kU(A)/L) and total IgE (429 vs 300 kU/L) levels than nonresponders. Ranking of extract potency was consistent between the mediator release assay and the ID(50)EAL. For the in-house prepared cockroach extract, the dose-response curves were shifted according to the concentration of the extract. NHR was reproducible between different experiments by using pooled sera. CONCLUSION/CONCLUSIONS:The mediator release assay measures biologic potency and correlates with the ID(50)EAL. It should be further evaluated to determine whether it could be used to replace intradermal skin test titration for assessing the potency of cockroach extract.

BACKGROUND:Children with milk allergy who tolerate heat-denatured milk (HM) have less severe reactions and outgrow the condition earlier than those who react to HM, which might be related to differences in IgE-dependent effector cell function. OBJECTIVE:We sought to apply a novel assay to test the hypothesis that HM-tolerant children have suppressed IgE-mediated basophil responses. METHODS:Allergic, HM-tolerant, outgrown, or control subjects were defined based on oral food challenges. Whole blood cells were stimulated in vitro with a range of milk allergen doses in the presence or absence of autologous serum or with dilutions of autologous serum. Activated basophils were identified by means of flow cytometry as CD63(bright)CD123+CD203c+HLA-DR(-)CD41a(-). RESULTS:HM-tolerant subjects' basophils were significantly less responsive to milk allergen stimulation at all doses than were basophils from HM-reactive (allergic) individuals. In the absence of autologous serum, HM-tolerant subjects' basophils were significantly more reactive at low allergen concentrations. To a lesser extent, autologous serum also inhibited IL-3- and anti-IgE-induced, but not N-formyl-methionyl-leucyl-phenylalanine-induced, responses. The allergen-specific responsiveness of HM-tolerant subjects' basophils increased with dilution of autologous serum with normal pooled serum. CONCLUSION/CONCLUSIONS:Children with milk allergy with a favorable prognosis have evidence of extrinsically suppressed allergen-specific effector cell reactivity.

BACKGROUND:About 70% of children with milk allergy tolerate extensively heated milk (HM) products and outgrow their allergy earlier than those who react to HM. OBJECTIVE:To test the hypothesis that HM-tolerant children have a higher precursor frequency of adaptive allergen-specific regulatory T (Treg) cells. METHODS:Allergic, HM-tolerant, outgrown, or control subjects were defined by oral food challenge. PBMCs were cultured with purified caseins and controls for 7 days, and proliferating CD25(+)CD27(+) Treg cells were identified by flow cytometry. Proliferating cells were also characterized for their expression of FoxP3, CTLA 4, CD45RO, and CD127. Allergen-specific Treg cell origin and function were assessed by depletion of CD25(hi) cells before culture. RESULTS:There was a higher percentage (median [25th% to 75th%], 16.85% [7.1-31.7]) of proliferating allergen-specific CD25(+)CD27(+) T cells from cultures of HM-tolerant subjects (n = 18) than subjects with allergy (n = 8; 4.91% [2.6-7.5]; P < .01). Control subjects with no history of milk allergy (n = 7) also had low percentages of these cells (2.9% [2.4-6.0]), whereas outgrown subjects (n = 7) had intermediate percentages (9.0% [2.7-16.4]). There were no significant differences between the patient groups in the frequency of polyclonal Treg cells or allergen-specific effector T cells. Allergen-specific Treg cells were found to be FoxP3(+)CD25(hi)CD27(+), cytotoxic T lymphocyte-associated antigen 4(+), CD45RO(+)CD127(-) and were derived from circulating CD25(hi) T cells. Depletion of the CD25(hi) cells before in vitro culture significantly enhanced allergen-specific effector T-cell expansion. CONCLUSION/CONCLUSIONS:A higher frequency of milk allergen-specific Treg cells correlates with a phenotype of mild clinical disease and favorable prognosis.

BACKGROUND:Prior studies have suggested that heated egg might be tolerated by some children with egg allergy. OBJECTIVE:We sought to confirm tolerance of heated egg in a subset of children with egg allergy, to evaluate clinical and immunologic predictors of heated egg tolerance, to characterize immunologic changes associated with continued ingestion of heated egg, and to determine whether a diet incorporating heated egg is well tolerated. METHODS:Subjects with documented IgE-mediated egg allergy underwent physician-supervised oral food challenges to extensively heated egg (in the form of a muffin and a waffle), with tolerant subjects also undergoing regular egg challenges (in a form of scrambled egg or French toast). Heated egg-tolerant subjects incorporated heated egg into their diets. Skin prick test wheal diameters and egg white, ovalbumin, and ovomucoid IgE levels, as well as ovalbumin and ovomucoid IgG4 levels, were measured at baseline for all subjects and at 3, 6, and 12 months for those tolerant of heated egg. RESULTS:Sixty-four of 117 subjects tolerated heated egg, 23 tolerated regular egg, and 27 reacted to heated egg. Heated egg-reactive subjects had larger skin test wheals and greater egg white-specific, ovalbumin-specific, and ovomucoid-specific IgE levels compared with heated egg- and egg-tolerant subjects. Continued ingestion of heated egg was associated with decreased skin test wheal diameters and ovalbumin-specific IgE levels and increased ovalbumin-specific and ovomucoid-specific IgG4 levels. CONCLUSIONS:The majority of subjects with egg allergy were tolerant of heated egg. Continued ingestion of heated egg was well tolerated and associated with immunologic changes that paralleled the changes observed with the development of clinical tolerance to regular egg.

BACKGROUND:Cow's milk allergy is the most common childhood food allergy. Previously we noted that children who outgrew their milk allergy had milk-specific IgE antibodies primarily directed against conformational epitopes; those with persistent milk allergy also had IgE antibodies directed against specific sequential epitopes. OBJECTIVE:Because high temperature largely destroys conformational epitopes, we hypothesized that some children with milk allergy would tolerate extensively heated (baked) milk products. METHODS:Children with milk allergy were challenged with heated milk products; heated milk-tolerant subjects were subsequently challenged with unheated milk. Heated milk-tolerant, unheated milk-reactive subjects ingested heated milk products for 3 months and were then re-evaluated. Immune responses were assessed in all subjects; growth and intestinal permeability were followed in heated milk-tolerant subjects. RESULTS:One hundred children (mean age, 7.5 years; range, 2.1-17.3 years) underwent heated milk challenges. Sixty-eight subjects tolerated extensively heated milk only, 23 reacted to heated milk, and 9 tolerated both heated and unheated milk. Heated milk-reactive subjects had significantly larger skin prick test wheals and higher milk-specific and casein-specific IgE levels than other groups. At 3 months, subjects ingesting heated milk products had significantly smaller skin prick test wheals and higher casein-IgG(4) compared with baseline; other immunologic parameters, growth, and intestinal permeability were not significantly different. Heated milk-reactive subjects had more severe symptoms during heated milk challenge than heated milk-tolerant subjects experienced during their unheated milk challenge. CONCLUSION/CONCLUSIONS:The majority (75%) of children with milk allergy tolerate heated milk.

BACKGROUND:Food allergy is the most common cause of anaphylaxis outside the hospital setting. OBJECTIVE:We sought to determine the rate, circumstances, and risk factors for repeated doses of epinephrine in the treatment of food-induced anaphylaxis in children. METHODS:Anonymous questionnaires were distributed to families of children with food allergies during allergy outpatient visits to a food allergy referral center. Demographic information, allergy and reaction history, and details regarding the last 2 anaphylactic reactions requiring epinephrine were collected. RESULTS:A total of 413 questionnaires were analyzed. Seventy-eight children (median, 4.5 years of age; range, 0.5-17.5 years) reported 95 reactions for which epinephrine was administered. Two doses were administered in 12 (13%) and 3 doses in an additional 6 (6%) reactions treated with epinephrine. Peanut, tree nuts, and cow's milk were responsible for >75% of reactions requiring epinephrine. Patients receiving multiple doses of epinephrine more often had asthma (P = .027) than children receiving a single dose. The amount of food ingested or a delay in the initial administration of epinephrine were not risk factors for receiving multiple doses. The second dose of epinephrine was administered by a health care professional in 94% of reactions. CONCLUSION/CONCLUSIONS:In this referral population of children and adolescents with multiple food allergies, 19% of food-induced anaphylactic reactions were treated with more than 1 dose of epinephrine. Prospective studies are necessary to identify risk factors for severe anaphylaxis and to establish rational guidelines for prescribing multiple epinephrine autoinjectors for children with food allergy.

Food-induced anaphylaxis is a steadily increasing problem in westernized countries and now represents the leading cause of anaphylaxis in the outpatient setting, particularly in children. Much of our knowledge of the pathophysiology of food-induced anaphylaxis comes from animal studies. Food anaphylaxis in humans is thought to be entirely IgE mediated. Several features appear to be unique to these reactions; factors such as exercise can lower the "threshold" for anaphylaxis in certain susceptible individuals. Different methods of thermal processing can modify the allergenicity of food proteins. Alteration of stomach pH can allow for incomplete digestion of food proteins, leading to increased absorption of intact food allergens. Low serum platelet-activating factor acetylhydrolase may predispose to fatal food-induced anaphylaxis. With a greater understanding of the pathophysiology of food-induced anaphylaxis, novel approaches not only to identify those at risk, but to treat and ultimately prevent food-induced anaphylaxis, are on the horizon.