Sub-class of IgG in allergic disease I. IgG sub-class antibodies in immediate and non-immediate food allergy

Abstract. Previous studies have suggested that, apart from IgE-mediated reactions, some of the symptoms of food allergy may be caused by IgG antibodies to food proteins. This study was carried out to see if there were any distinctive features of the IgG sub-class antibody response to dietary antigens which occurs in food allergic patients. IgG subclass antibodies were measured using a quantitative enzyme-linked immuno-sorbent assay (ELISA) to wheat gliadin, ovalbumin and bovine casein in twenty patients who had coeliac disease and in twenty-eight egg allergic patients. These were compared with twenty-one atopic dermatitis patients who did not have food allergy and twenty-six healthy control subjects. Coeliac disease patients tended to have raised IgG antibody levels (especially IgG1) to all three antigens but these overlapped considerably with that seen in egg allergic and atopic dermatitis patients. Coeliacs who avoided gluten had anti-gliadin antibody levels which did not differ from those seen in healthy subjects but nevertheless had raised anti-ovalbumin and casein-specific antibodies. The IgG antibody was largely restricted to IgG1 and IgG4 sub-class although the relative amount of each varied with the antigen. Although gliadin-specific antibodies were mainly IgG1, ovalbumin-specific antibodies were mainly IgG4. The increased antibody levels to all three antigens in coeliacs were caused by a raised IgG1 response, IgG4 antibodies were usually normal. Egg allergic patients also had raised IgG1 but not IgG4 antibodies to ovalbumin. These data show that the response to different dietary antigens can vary with the antigen. The fact that IgG1 and not IgG4antibodies were raised to all three antigens in patients with coeliac disease suggests that they are a secondary consequence of the disease, perhaps reflecting increased transport of antigens across a damaged gut mucosa rather than a specific immunopathological reaction. However, the observation that antibodies to gliadin, and not ovalbumin or casein, fell following gluten avoidance shows that the response to gliadin, at least, is dependent upon continued exposure to antigen.     

The use of aluminium hydroxide as adjuvant modulates the specific antibody response—A Brown Norway rat study with native and denatured cow's milk allergens

There is a need for efficient methods to treat food allergy; however, no immunotherapeutic method has yet been satisfactory due to the high rate of unpredictable severe reactions and the limited efficacy. Therefore, modified versions of food allergens have been suggested as alternatives to the parent proteins for immunotherapy. The aim of the study was to compare the inherent allergenicity of the native and denatured version of the cow's milk proteins β-lactoglobulin and α-lactalbumin, and to study the impact of the use of Al(OH)₃ as an adjuvant. Brown Norway rats were immunized intraperitoneally with either native or denatured β-lactoglobulin or α-lactalbumin, with or without the use of Al(OH)₃ as adjuvant. Antibody responses were analysed in various ways by means of different ELISAs. Both the immunogenicity and the sensitizing capacity of the cow's milk allergens were influenced by their globular folding, with the native version being more allergenic than the denatured counterpart. The native folded proteins mainly raised antibodies against conformational epitope, whereas the denatured versions predominantly raised antibodies against linear epitopes. Most interestingly, the study showed that the use of Al(OH)₃, besides increasing immunogenicity and sensitizing capacity of the cow's milk allergens, caused a modification of the specificity of the antibodies raised against the native version of the proteins. Adsorption of the native forms of the allergens to Al(OH)₃ caused a significant greater proportion of antibodies raised against linear epitopes, stressing that the adsorption induced a partly unfolding of the proteins. This may have implications for IT safety and efficacy.     

Animal models to study gluten sensitivity

The initial development and maintenance of tolerance to dietary antigens is a complex process that, when prevented or interrupted, can lead to human disease. Understanding the mechanisms by which tolerance to specific dietary antigens is attained and maintained is crucial to our understanding of the pathogenesis of diseases related to intolerance of specific dietary antigens. Two diseases that are the result of intolerance to a dietary antigen are celiac disease (CD) and dermatitis herpetiformis (DH). Both of these diseases are dependent upon the ingestion of gluten (the protein fraction of wheat, rye, and barley) and manifest in the gastrointestinal tract and skin, respectively. These gluten-sensitive diseases are two examples of how devastating abnormal immune responses to a ubiquitous food can be. The well-recognized risk genotype for both is conferred by either of the HLA class II molecules DQ2 or DQ8. However, only a minority of individuals who carry these molecules will develop either disease. Also of interest is that the age at diagnosis can range from infancy to 70-80 years of age. This would indicate that intolerance to gluten may potentially be the result of two different phenomena. The first would be that, for various reasons, tolerance to gluten never developed in certain individuals, but that for other individuals, prior tolerance to gluten was lost at some point after childhood. Of recent interest is the concept of non-celiac gluten sensitivity, which manifests as chronic digestive or neurologic symptoms due to gluten, but through mechanisms that remain to be elucidated. This review will address how animal models of gluten-sensitive disorders have substantially contributed to a better understanding of how gluten intolerance can arise and cause disease.     

Species-specific tissue antigens

Rabbit antisera to human tracheal mucosa, liver and gastric mucosa, after complete absorption with human serum, reacted with extracts of all human tissues tested. Monkey tissue extracts also reacted, but tissues of other species did not. The reactions were attributed to `species-specific tissue antigens'. Two of the most prominent antigens were partially isolated and characterized by chromatography, gel filtration, and electrophoresis. Antigen I migrated as β₂-globulin, Antigen E as a β₁-globulin. The antigens persisted in malignant and in long-term cultured cells.     

Quantitative estimation of cellular infiltration of the small intestinal mucosa in children with cow's milk and gluten intolerance

Quantitative estimation of the infiltration by intraepithelial lymphocytes and eosinophils of the mucosa was carried out in 21 children with cow's milk and 35 children with gluten intolerance. Before dietary treatment, a statistically significant increase in the infiltration by LIE in children with milk intolerance to the mean value of 34.1 cells and in children with gluten intolerance to 39.0 cells was found, what statistically significantly differed from the mean value of LIE for the control group (19.0 cells/100 epithelial cells). The eosinophilic infiltration in this phase of the disease was noted in 38% of children with cow's milk intolerance (16.9 cells/mm2) and in 27% of children with gluten intolerance (28.6 cells/mm2). After 8-24 months of elimination diets--a decrease in the mean value of the LIE infiltration in the mucosa was revealed in both treated groups.     

C1 inactivator protein complexed with albumin in plasma from a patient with angioneurotic edema

An abnormal protein, antigenically identical with C1 inactivator and belonging electrophoretically to the α₁-globulins was found in plasma and serum from a patient with angioneurotic edema. Treatment of the pathological protein with 2-mercaptoethanol or with neuraminidase followed by analysis with antigen-antibody crossed electrophoresis showed that the protein was a complex of C1 inactivator and albumin. The patient's serum and plasma was found to contain also a C1 inactivator protein migrating with α₂-globulin but somewhat faster than normal C1 inactivator. Neither the α₂- nor the α₁-C1 inactivator inhibited the hydrolyzing effect of C1 on N-acethyl-L-tyrosine ethyl ester.     

Studies of intestinal lymphoid tissue. IV--The predictive value of raised mitotic indices among jejunal epithelial lymphocytes in the diagnosis of gluten-sensitive enteropathy.

It has been established that considerable blast-transformation and mitotic activity occurs among epithelial lymphocytes of untreated coeliac mucosa. This paper is concerned solely with the proliferative activity of epithelial lymphocytes (expressed as percentage "mitotic index") in the prospective diagnosis of coeliac disease, in comparison with other conditions such as lymphoma. Crohn's disease and immunodeficiency which are often associated with malabsorption and flattening of jejunal mucosa. The results demonstrate that a high mitotic index (greater than 0.2%) clearly distinguishes, and hence predicts, gluten-associated enteropathies (including dermatitis herpetiformis and malignant histiocytosis) from others in which gluten plays no aetiological role and where the mitotic index differs insignificantly from normal control mucosae (much less than 0.2%). Furthermore, it has been demonstrated that the mitotic index is raised in so-called "non-responsive coeliacs," thus suggesting that such patients may also be gluten-sensitive despite their subsequent failure to respond morphologically to dietary gluten restriction.     

Prevention

The development of food allergy depends on several factors, including genetic factors and early exposure to allergenic proteins in the diet, food protein uptake and handling, and the development of tolerance. Many hypotheses, as regards the possible causal relationships, have been raised during the past few years, including the hygiene theory, the role of bacterial gut flora, and the potential effect of different cytokines in breast milk. Although interesting, these are mainly speculations based on non-interventional and often retrospective/cross-sectional studies including small study populations. These theories remain to be documented in proper, controlled and prospective studies. Breastfeeding and the late introduction of solid foods (>4 months) is associated with a reduced risk of food allergy, atopic dermatitis, and recurrent wheezing and asthma in early childhood. In all infants, breastfeeding should be encouraged for 4-6 months. In high-risk infants a documented extensively hydrolysed formula is recommended if exclusive breastfeeding is not possible for the first 4 months of life. There is no evidence for preventive dietary intervention neither during pregnancy nor lactation. Preventive dietary restrictions after the age of 4-6 months are not scientifically documented.     

Immune response patterns in coeliac disease. Serum antibodies to dietary antigens measured by an enzyme linked immunosorbent assay (ELISA)

Serum IgG, IgA and IgM activities to wheat, egg and cow's milk antigens were measured by an ELISA method in children and adults with coeliac disease (CD). In untreated patients, the IgA activity was characteristically raised to gluten antigens but often also to proteins from egg or cow's milk. Setting the upper reference range for gluten antibodies as the highest IgA reading obtained in healthy controls and patients with other intestinal disorders, IgA measurements afforded virtually 100% diagnostic sensitivity and specificity and detected 94% of children and 80% of adults with untreated CD. Such measurements, therefore, represent a valuable adjunct in the diagnosis of this disease. IgA activity to beta-lactoglobulin, casein or ovalbumin higher than the normal 95 percentile was found in 44-89% of untreated patients. Reduction of these antibody titres seemed to reflect relatively well the response to treatment with a gluten free diet, particularly the activity to beta-lactoglobulin. Monitoring of IgA antibodies to dietary antigens other than gluten may therefore be of particular importance in the follow-up of CD patients.     

Passive sensitization of skin and lung by guinea-pig immunoglobulins

Guinea-pig γ₁- and γ₂-globulins have been purified by preparative electrophoresis followed by chromatography. No γ₁-globulin was detectable in purified γ₂-globulin, but purified γ₁-globulin always contained fast γ₂-globulin. Normal guinea-pig serum contained much less γ₁-globulin than immune serum. Antisera prepared against normal guinea-pig serum did not contain useful amounts of antibody specific for γ₁-globulin.

Guinea-pig lung tissue was sensitized by very low concentrations of guinea-pig γ₁-globulin (of the order of 6×10^-10 molar) but γ₂-globulin antibodies were almost inactive. No evidence was found that the trace of activity in γ₂-globulin was not due to very slight contamination with γ₁-globulin antibodies.

The finding that γ₁-globulin antibodies are far more potent than γ₂-globulin antibodies in sensitizing skin has been confirmed, but several lines of evidence suggest that γ₂-globulin antibodies may also have weak activity. Thus quantitative passive cutaneous anaphylaxis (PCA) tests showed that whenever the γ₂-globulin fraction contained antibody it appeared far more potent relative to γ₁-globulin than when the same proteins were tested on lung tissue. The PCA activity of moderate amounts of purified γ₂-globulin antibodies disappeared faster than the skin sensitization produced by small amounts of γ₁-globulin antibodies, and the γ₂-globulin preparations did not contain enough γ₁-globulin impurity to account for their PCA activity. No inhibition of skin responses was observed with the largest doses of antigen tested.

The most plausible explanation of these results is that, under the conditions of our experiments, γ₂-globulin antibody had weak PCA activity. Objections to this hypothesis are discussed. The PCA activity of γ₂-globulin antibody probably involves a mechanism different from that of the sensitization produced by the highly potent γ₁-globulin antibody.

     

Changes of serum antibody activities to various dietary antigens related to gluten withdrawal or challenge in children with coeliac disease

IgG, IgA, and IgM serum antibody activities to gluten, a gluten fraction called glyc-gli, and antigens from egg and cow's milk were monitored by an enzyme-linked immunosorbent assay (ELISA) in children with coeliac disease during treatment and gluten challenge. The IgA activity to gluten antigens showed in most patients a rapid and significant reduction after gluten withdrawal, whereas the IgG activity decreased more slowly. During gluten challenge, both these activities rose significantly, and the increases could usually be detected several months before overt clinical relapse. Such determinations, therefore, represent a valuable adjunct in the follow-up of children with coeliac disease. IgA activities to egg and cow's milk antigens likewise tended to decrease after gluten withdrawal and increase during challenge, but the changes were less consistent for individual antigens. Nevertheless, monitoring of IgA activities to a selection of dietary antigens other than gluten may be particularly valuable when it comes to evaluation of intestinal responses in patients on a gluten-free diet.     

Repeatability of Dietary Patterns Derived Using α-Priori and α-Posterior Methods

We aimed to examine the repeatability of dietary patterns derived using α-priori and α-posterior techniques. During 2008, 500 participants were enrolled and asked to fill-in a food frequency questionnaire twice. The Mediterranean dietary pattern was α-priori assessed by the MedDietScore, while principal components analysis (PCA) and cluster analysis (CA) were used as the α-posterior techniques. Results revealed that the overall MedDietScore was similar between the two recordings (M = 28, SD = 3.7 vs. M = 28, SD = 3.8). Although PCA revealed 13 patterns in each record (60% of explained variability), the food items characterizing each pattern were varying through the two recordings. According to CA, three clusters were revealed from both records. The α-posterior methods should be used with caution, while the α-priori approach leads to more robust results.     

Small-intestinal TG2-specific plasma cells at different stages of coeliac disease

Background

In coeliac disease, ingestion of gluten induces the production of transglutaminase 2 (TG2)-targeted autoantibodies by TG2-specific plasma cells present at high frequency in the small intestinal mucosa in untreated disease. During treatment with a gluten-free diet (GFD), the number of these cells decreases considerably. It has not been previously investigated whether the cells are also present prior to development of villous atrophy, or in non-responsive patients and those with dietary lapses. We aimed to define the frequency of small bowel mucosal TG2-specific plasma cells in coeliac disease patients with varying disease activity, and to investigate whether the frequency correlates with serum and small intestinal TG2-targeting antibodies as well as mucosal morphology and the number of intraepithelial lymphocytes.

Results

Mucosal TG2-specific plasma cells were found in 79% of patients prior to development of mucosal damage, in all patients with villous atrophy, and in 63% of the patients after 1 year on GFD. In these disease stages, TG2-specific plasma cells accounted for median of 2.3, 4.3, and 0.7% of all mucosal plasma cells, respectively. After long-term treatment, the cells were present in 20% of the patients in clinical remission (median 0%) and in 60% of the patients with poor dietary adherence (median 5.8%). In patients with non-responsive coeliac disease despite strict GFD, the cells were found in only one (9%) subject; the cells accounted for 2.4% of all plasma cells. A positive correlation between the percentage of TG2-specific plasma cells and serum TG2 antibody levels (r_S?=?0.69, P?<?0.001) and the intensity of mucosal TG2-targeting IgA deposits (r_S?=?0.43, P?<?0.001) was observed.

Conclusions

Our results show that TG2-specific plasma cells are already detectable prior to villous atrophy, and that generally their frequency increases during overt disease. By contrast, on GFD, the percentage of these cells decreases. Overall, the presence of TG2-specific plasma cells in the small bowel mucosa mirrors the presence of gluten in the diet, but the frequency is not always parallel to the level of serum or intestinal TG2 antibodies. These findings increase the knowledge about the development of the TG2 plasma cell responses especially in the early phases of coeliac disease.

     

Integrin expression in normal,hyperplastic, dysplastic,and malignant oral epithelium

We have examined the distribution of a range of integrin subunits in normal and lesional oral mucosa. The α₂, α₃, α₆, β₁β, and β₄ subunits were highly expressed in normal epithelium, and there was weaker, more variable expression of α₅ and α_v. Expression of all subunits was highest in the basal layer of normal epithelium, but extensive staining above the basal layer was also observed, particularly in the floor of the mouth and the lateral margin of the tongue. In dysplastic lesions and hyperplastic epithelium adjacent to ulcers, suprabasal staining was even more pronounced. Staining patterns in squamous cell carcinomas showed considerable variation, both within and between individual tumours: in some areas there was staining reminiscent of normal epithelium, but uniform staining throughout tumour islands, and patchy and variable cytoplasmic and pericellular staining were also seen. Thirteen out of 17 carcinomas showed some loss of integrin expression: six out of ten moderately well differentiated tumours and all the poorly differentiated tumours. Focal loss of a₆ and β₄ was most commonly observed, but loss of α₂ and β₃ also occurred. Since integrins regulate not only keratinocyte adhesion, but also the initiation of terminal differentiation, the changes in integrin expression that we have observed may have significance for the behaviour of individual tumours.     

Dietary manipulation in experimental inflammatory bowel disease

Eicosanoids are major mediators of defensive and inflammatory processes of the gut mucosa. The activity of the eicosanoid system is modulated by neural and hormonal pathways, but local factors acting within the gastrointestinal lumen may also be involved. We have studied the influence of dietary fatty acids on eicosanoid synthesis by the gastrointestinal mucosa. Since omega-3 fatty acids compete with the omega-6 as precursors of eicosanoid synthesis, we compared the effects of dietary supplementation with either sunflower or cod liver oil as sources of omega-6 or omega-3 fatty acids, respectively. Rats fed with the codliver-oil-supplemented diet for four weeks showed high omega-3 and low omega-6 plasma fatty acid levels compared to rats fed with the sunflower oil diet. Synthesis of arachidonic-acid-derived eicosanoids (6-keto-PGF_1α, PGE₂, TXB₂, LTB₄, and LTC₄) by gastric and intestinal mucosa was found to be lower in the cod liver group as compared to the sunflower group. However, significant generation of eicosapentaenoic-acid-derived ecosanoids (PGE₃ and LTC₅) was observed only in the cod liver group.We used the (trinitrobenzenesulphonic acid) TNBS model of inflammatory colitis to test the effect of the dietary fat on the development of inflammatory lesions of the bowel. A single intracolonic instillation of the hapten TNBS dissolved in 10% ethanol induces chronic granulomatous lesions of the colonic mucosa that persist for up to 8 weeks. Luminal release of eicosanoid mediators, as measured by intracolonic dialysis, was lower in the cod liver group than in the sunflower group, particularly during the chronic stage of the disease. Macroscopical and microscopical assessment of the lesions, serially performed for up to 8 weeks, showed significant differences between both groups, suggesting that the fish oil diet diminishes the severity of the lesions and their progression to chronicity. In conclusion, changes in the pattern of eicosanoid synthesis by the gastrointestinal mucosa can be induced by altering the dietary intake of their fatty acid precursors. These changes may be relevant for the expression of disease.     

Gluten-sensitive enteropathy coincides with decreased capability of intestinal T cells to secrete IL-17 and IL-22 in a macaque model for celiac disease

Celiac disease (CD) is an autoimmune disorder caused by intolerance to dietary gluten. The interleukin (IL)-17 and IL-22 function as innate regulators of mucosal integrity. Impaired but not well-understood kinetics of the IL-17/22 secretion was described in celiac patients. Here, the IL-17 and IL-22-producing intestinal cells were studied upon their in vitro stimulation with mitogens in class II major histocompatibility complex-defined, gluten-sensitive rhesus macaques. Pediatric biopsies were collected from distal duodenum during the stages of disease remission and relapse. Regardless of dietary gluten content, IL-17 and IL-22-producing cells consisted of CD4 + and CD8 + T lymphocytes as well as of lineage-negative (Lin −) cells. Upon introduction of dietary gluten, capability of intestinal T cells to secrete IL-17/22 started to decline (p < 0.05), which was paralleled with gradual disruption of epithelial integrity. These data indicate that IL-17/22-producing cells play an important role in maintenance of intestinal mucosa in gluten-sensitive primates.     

Intestinal inflammation in children with atopic eczema: faecal eosinophil cationic protein and tumour necrosis factor-α as non-invasive indicators of food allergy

Background Food allergy is contemplated in atopic eczema. Early recognition of food allergies is difficult and the diagnosis is often missed because of the non-specificity of symptoms. New non-invasive tests are clearly needed. Objeetive and methods We measured the concentrations of tumour necrosis factor-α, eosinophil cationic protein and α-1 antitrypsin in faeces as indicators of intestinal inflammation induced by double-blind placebo-controlled oral cow's milk challenge in infants and young children with atopic eczema. Results An increased α-l antitrypsin concentration (>2mg/g) after cow's milk challenge was detected in 43% of the infants positive as compared with 11% of the infants negative to challenge P= 0.02. The concentration of eosinophil cationic protein in faeces increased after cow's milk challenge in patients positive to challenge (P=0.02) but not in those negative to challenge (P=0.79). The concentration of eosinophil cationic protein was enhanced particularly in patients manifesting immediate-type reactions to the cow's milk challenge. The concentration of tumour necrosis factor-α increased after cow's milk challenge in patients positive to challenge (P=0.005) but not in those negative to challenge (P =0.25). The concentration of tumour necrosis factor-α in faeces was enhanced particularly in patients manifesting delayed-type reactions to the cow's milk challenge. Conclusion We conclude that in children with atopic eczema food allergy is associated with intestinal inflammation indicating that more general immunologic disturbances than previously thought take place in these patients. We further suggest that faecal eosinophil cationic protein, tumour necrosis factor-α and α-1 antitrypsin distinctly indicate various reaction types of food allergy. Parallel testing with eosinophil cationic protein and tumour necrosis factor-α may signiticantly enhance the accuracy in diagnosis of food allergy in patients with atopic eczema.     

Dietary manipulation in experimental inflammatory bowel disease

Eicosanoids are major mediators of defensive and inflammatory processes of the gut mucosa. The activity of the eicosanoid system is modulated by neural and hormonal pathways, but local factors acting within the gastrointestinal lumen may also be involved. We have studied the influence of dietary fatty acids on eicosanoid synthesis by the gastrointestinal mucosa. Since omega-3 fatty acids compete with the omega-6 as precursors of eicosanoid synthesis, we compared the effects of dietary supplementation with either sunflower or cod liver oil as sources of omega-6 or omega-3 fatty acids, respectively. Rats fed with the codliver-oil-supplemented diet for four weeks showed high omega-3 and low omega-6 plasma fatty acid levels compared to rats fed with the sunflower oil diet. Synthesis of arachidonic-acid-derived eicosanoids (6-keto-PGF₁, PGE₂, TXB₂, LTB₄, and LTC₄) by gastric and intestinal mucosa was found to be lower in the cod liver group as compared to the sunflower group. However, significant generation of eicosapentaenoic-acid-derived ecosanoids (PGE₃ and LTC₅) was observed only in the cod liver group.We used the (trinitrobenzenesulphonic acid) TNBS model of inflammatory colitis to test the effect of the dietary fat on the development of inflammatory lesions of the bowel. A single intracolonic instillation of the hapten TNBS dissolved in 10% ethanol induces chronic granulomatous lesions of the colonic mucosa that persist for up to 8 weeks. Luminal release of eicosanoid mediators, as measured by intracolonic dialysis, was lower in the cod liver group than in the sunflower group, particularly during the chronic stage of the disease. Macroscopical and microscopical assessment of the lesions, serially performed for up to 8 weeks, showed significant differences between both groups, suggesting that the fish oil diet diminishes the severity of the lesions and their progression to chronicity. In conclusion, changes in the pattern of eicosanoid synthesis by the gastrointestinal mucosa can be induced by altering the dietary intake of their fatty acid precursors. These changes may be relevant for the expression of disease.     

Milk allergy and vitamin D deficiency rickets: a common disorder associated with an uncommon disease.

BACKGROUND: Cow's milk allergy is one of the most common allergies in infancy. It has an excellent prognosis since most cases resolve by 4 years of age. The complications associated with milk allergy include delayed growth and atopic conditions, such as asthma, allergic rhinitis, atopic dermatitis, and other food allergies. OBJECTIVE: To report a case of vitamin D deficiency rickets in a 2-year-old boy with cow's milk allergy. METHODS: We describe a patient with clinical and biochemical evidence of rickets, including decreased serum calcium, phosphate, and 25-hydroxy vitamin D levels and an elevated alkaline phosphatase level. A dietary history revealed the prolonged absence of dietary vitamin D because the child did not tolerate cow's milk. Skin prick testing and measurement of specific IgE to cow's milk were performed to determine whether there was an allergy to cow's milk. RESULTS: Results of skin prick testing and measurement of specific IgE to cow's milk confirmed an IgE-mediated sensitivity to cow's milk. Introduction of appropriate supplementation into the child's diet resulted in complete resolution of his symptoms. CONCLUSIONS: This case emphasizes that the management of cow's milk allergy involves strict avoidance of the allergenic food while also ensuring that essential dietary requirements are met. A dietary history is crucial at all pediatric visits, and inquiry about supplementation of vitamins and minerals is important, especially in children with food allergies.     

Milk-whey diet substantially suppresses seizure-like phenotypes of paraShu,a Drosophila voltage-gated sodium channel mutant

The Drosophila mutant para^Shu harbors a dominant, gain-of-function allele of the voltage-gated sodium channel gene, paralytic (para). The mutant flies display severe seizure-like phenotypes, including neuronal hyperexcitability, spontaneous spasms, ether-induced leg shaking, and heat-induced convulsions. We unexpectedly found that two distinct food recipes used routinely in the Drosophila research community result in a striking difference in severity of the para^Shu phenotypes. Namely, when para^Shu mutants were raised on the diet originally formulated by Edward Lewis in 1960, they showed severe neurological defects as previously reported. In contrast, when they were raised on the diet developed by Frankel and Brousseau in 1968, these phenotypes were substantially suppressed. Comparison of the effects of these two well-established food recipes revealed that the diet-dependent phenotypic suppression is accounted for by milk whey, which is present only in the latter. Inclusion of milk whey in the diet during larval stages was critical for suppression of the adult para^Shu phenotypes, suggesting that this dietary modification affects development of the nervous system. We also found that milk whey has selective effects on other neurological mutants. Among the behavioral phenotypes of different para mutant alleles, those of para^GEFS+ and para^bss were suppressed by milk whey, while those of para^DS and para^ts1 were not significantly affected. Overall, our study demonstrates that different diets routinely used in Drosophila labs could have considerably different effects on neurological phenotypes of Drosophila mutants. This finding provides a solid foundation for further investigation into how dietary modifications affect development and function of the nervous system and, ultimately, how they influence behavior.