近藤 康人

BACKGROUND: To evaluate the long-term safety of subcutaneous immunotherapy with TO-204, a standardized house dust mite (HDM) allergen extracts, we conducted a multicenter, open label clinical trial. METHODS: Japanese patients aged 5-65 years were eligible for the study, if they had HDM-induced allergic rhinitis (AR), allergic bronchial asthma (BA), or both. TO-204 was administered in a dose titration scheme, and the maintenance dose was determined according to the predefined criteria. The treatment period was 52 weeks, and patients who were willing to continue the treatment received TO-204 beyond 52 weeks. This clinical trial is registered at the Japan Pharmaceutical Information Center (Japic CTI-121900). RESULTS: Between July 2012 and May 2015, 44 patients (28 with AR and 16 with allergic BA) were enrolled into the study. All patients were included in the analysis. The duration of treatment ranged from 23 to 142 weeks and the median maintenance dose was 200 Japanese allergy units (JAU). Adverse events occurred in 22 patients (50%). The most common adverse event was local reactions related to the injection sites. Four patients experienced anaphylactic reactions when they were treated with the dose of 500 JAU. Two patients experienced anaphylactic shock with the doses of 1000 JAU at onset. These 6 patients could continue the study with dose reduction. CONCLUSIONS: Safety profile of TO-204 was acceptable in Japanese patients with HDM-induced AR or allergic BA. Higher doses should be administered carefully, because the risk of anaphylaxis increased at doses of 500 or 1000 JAU.

Background: We evaluated the clinical significance of the spontaneous histamine release ratio (SHR/T) and low responders in the automated basophil histamine release test (Allerport® HRT). Methods: This study analyzed the outcomes of 101 oral food challenges (OFC) with egg, milk or wheat (challenge-positive: n=79) in relation to the SHR/T. The traditional HRT low responders (n=27) were separated into two groups: "LOW" responders (n=10), who showed a ≥10% concentration-dependent maximum histamine release in response to the anti-human IgE stimulation, and "NON" responders who did not fulfill the criteria (n=17). Results: Among the 34 patients with ≥20% SHR/T, 32 patients (94%) had a positive OFC with a low threshold dose which provoked severe symptoms. Among the "LOW" responders, four cases showed ≥10% allergen-specific maximum histamine release. On the other hand, concentration-dependent histamine release was not seen in the "NON" responders, suggesting the basophil function was not detected in this subgroup. Conclusion: The present study suggested that SHR/T could be an indicator of basophil activation and hypersensitivity in vivo. We also suggested that significant basophil functions might be detected among the "LOW" responders, but not among the "NON" responders.

Background: Allergen-specific T-helper type 2 (TH2) cells play an important role in the development of allergic inflammation; however, investigations of the properties of allergen-specific T cells have been challenging in humans. Despite clear evidence that forkhead box p3 (Foxp3) is expressed in conventional effector T cells, its function has remained unknown. Objective: To characterize allergen-specific TH2 cells in milk allergy, with particular focus on the expression of Foxp3. Methods: Twenty-one children with milk allergy and 11 children without milk allergy were studied. Peripheral blood mononuclear cells from subjects were stimulated with milk allergen for 6 hours and analyzed using multicolor flow cytometry to identify CD154(+) allergen-specific T-helper cells. Simultaneously, the expression of intracellular cytokines and Foxp3 was analyzed. Results: The milk allergy group had significantly larger numbers of milk allergen-specific interleukin (IL)-4 and IL-5-producing CD4(+) T cells than the control group. Subjects in the milk allergy group had significantly more CD154(+)CD4(+)IL-10-producing cells and CD154(+)Foxp3(+)CD4(+) cells than those in the control group. In addition, the number of milk allergen-specific CD154(+)Foxp3(+)CD4(+) cells strongly correlated with that of CD154(+)IL4(+)CD4(+) cells. Bcl-2 expression in CD154(+)IL-4(+)Foxp3(+) T-helper cells was significantly lower compared with that in total CD4 cells. Conclusion: Increased numbers of IL-4-producing allergen-specific T-helper cells were found in patients with milk allergy. In addition, Foxp3 was coexpressed with IL-4 in allergen-specific TH2 cells from patients. This coexpression was associated with lower Bcl-2 levels and could contribute to the phenotype and function of TH2 cells. (C) 2015 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

Background: In recent years, it has been proposed that the primary mechanism for the development of food allergies is percutaneous sensitization. Since 2010, in Japan, the number of immediate-type wheat allergy due to hydrolyzed wheat protein has dramatically increased among those who have been using soap containing hydrolyzed wheat. This incidence supports the hypothesis that food allergens arise through percutaneous sensitization. Clinical Summary: A 25-year-old man (case 1) and an 18-year-old girl (case 2) with atopic dermatitis visited our Department because of food allergy and hand eczema. After starting their work with fish, severe itchy eczema appeared on their hands. They subsequently started to experience oral allergic symptoms, intraoral itchiness and dyspnea after eating fish. Specific IgE antibodies were detected for many fishes, and skin prick tests showed positive reactions for a variety of fishes in both cases. Furthermore, the fluorescence intensities of specific IgE antibodies against parvalbumin from various types of fish in microarray immunoassay analysis showed positive reactions. We diagnosed them as contact urticaria caused by percutaneous sensitization to parvalbumin through job-related physical contact with fish. Conclusion: The patients' histories and findings indicate the possibility of percutaneous sensitization through occupational exposure to parvalbumin, leading to food allergy.

Background: Some patients with Japanese cedar pollen (JCP)-induced allergic rhinitis develop pollen-food allergy syndrome (PFAS) as a reaction to tomato fruit. Pollen allergen-specific subcutaneous immunotherapy (SCIT) is reportedly beneficial for some associated food allergies; however, the reported changes in food allergen-specific immunoglobulin (Ig)E and IgG(4) levels are inconsistent. Here, we investigated immunologic reactivity to tomato fruit after JCP-based SCIT. Methods: Twenty-three children (aged 6-17 years) with JCP-induced allergic rhinitis and sensitized to tomato (serum tomato fruit-specific IgE level > 0.34 UA/ml) received JCP-based SCIT. Basophil activation by tomato and JCP extracts and serum-specific IgE and IgG(4) levels against these allergens were determined before and after 4 or 5 months of maintenance SCIT. Basophil activation was assessed by monitoring CD203c upregulation on flow cytometry. Results: JCP-based SCIT significantly reduced the basophil activation caused by tomato fruit (p = 0.03) and JCP (p < 0.001) extracts. JCP-specific IgG(4) levels markedly increased after SCIT (p < 0.001), whereas tomato fruit-specific IgG 4 levels did not. After SCIT, no significant changes were observed in specific IgE levels for tomato fruit (p = 0.11) or JCP (p = 0.19). Conclusions: Tomato fruit-specific basophil activation decreases after JCP-based SCIT, suggesting that it is efficacious in relieving and preventing the symptoms of PFAS in patients with JCP-induced allergic rhinitis. (C) 2015 The Author(s) Published by S. Karger AG, Basel

Enokitake (Flammnlina velutipes, winter mushroom) is a common edible mushroom in Japan. We experienced a case of anaphylaxis after enokitake ingestion. There are no reports describing anaphylaxis caused by the ingestion of this mushroom. Enokitake allergen has also not been reported. We thus attempted to identify enokitake allergen using the patient's serum. The patient was a seventeen-year-old woman who had had no episodes of food allergy and experienced anaphylaxis after the ingestion of sukiyaki (beef, pork, tofu, vegetables, enokitake, etc.). She had previously eaten sukiyaki (the same ingredients) without any symptoms. The result of enokitake skin prick to prick test was positive. Oral food challenge was positive, inducing anaphylaxis. We performed western blotting with enokitake extract and the patient's serum. Three enokitake protein bands (18kDa, 39 kDa, 50kDa) reacted specifically with the patient's IgE.

The patient was a 10-year-old girl who presented with a history of anaphylactic episodes on three occasions, that developed in association with exercise after she ate citrus fruit. She underwent tolerance tests, as food-dependent exercise-induced anaphylaxis (FDEIA) induced by citrus fruit was suspected. The result of the test for the combination of intake of oranges and exercise was negative. The patient presented with swollen eyelid and wheezing following combined intake of orange and aspirin, based on which she was diagnosed as having FDEIA. Many patients developing an allergic reaction to fruit are diagnosed as having oral allergy syndrome (OAS), and only few cases of FDEIA are reported. Immunoblot tests revealed antigens of 9kDa, 39kDa and 53kDa in this patient, and an inhibition study with oranges revealed that the 39kDa and 53kDa antigens were probably antigenspecific allergens. Although the studied patient showed a strongly positive result for IgE antibodies specifically directed at cedar pollen, no common antigenicity with cedar pollen could be recognized. The final diagnosis was a type of FDEIA caused by 39kDa and 53kDa proteins, which are different from antigens previously identified in patients with citrus fruits allergy. It should be the first report of such a case.

Dedicator of cytokinesis 8 (DOCK8) deficiency is an autosomal recessive type of combined immunodeficiency with elevated IgE. In this report, we describe a Japanese girl of non-consanguineous family suffering from acute eosinophilic pneumonia (AEP) as a presenting feature of DOCK8 deficiency. Although AEP was self-limiting, consecutively experienced recurrent respiratory infections, severe atopic dermatitis, and vulnerability to viral infections, prompted us to evaluate the possibility of DOCK8 deficiency. Immunological assessments demonstrated decreased IgM, increased IgE, T lymphocytepenia, especially in CD4 T cells, decreased PHA blastogenesis, and decreased CD27(+)CD19(+) memory B cells. Western blotting revealed the absence of DOCK8 protein. Investigation of genomic DNA by multiplex ligation-dependent probe amplification (MLPA) revealed a heterozygous large deletion of 77kb spanning from intron 5 to exon 22. DOCK8 cDNA sequencing revealed a nonsense mutation at position 740 (E740X). As far as we know, this is the first Japanese case of DOCK8 deficiency. Pediatr Pulmonol. 2014; 49:E52-E55. (c) 2013 Wiley Periodicals, Inc.

Background: Sea urchin roe can cause anaphylactic reactions the first time they are consumed; therefore, careful clinical attention should be paid to their effects. However, no previous study has examined the allergens in sea urchin roe using sera from more than one patient. We attempted to identify sea urchin allergens using sera from 5 patients with sea urchin allergies. Methods: We enrolled 5 patients with relevant medical histories, positive results on a skin prick test and/or a food challenge test, and high levels of sea urchin-specific IgE in an enzyme-linked immunosorbent assay. We performed SDS-PAGE, immunoblotting, immunoblot inhibition, and N-terminal amino acid sequence detection. Results: Ten protein bands ranging from 18 to 170 kDa were detected in more than 2 patients' sera. In immunoblotting, the protein band for the 170-kDa major yolk protein was recognized by 4 of the 5 sera. Furthermore, the reaction between IgE and the protein band for egg cortical vesicle protein (18 kDa) was inhibited by the addition of salmon roe extract. Conclusion: Major yolk protein was confirmed to be one of the main allergens in sea urchin roe. In addition, egg cortical vesicle protein (18 kDa) was demonstrated to be an important protein for cross-reactivity with salmon roe. (C) 2014 S. Karger AG, Basel

Background: Salmon is one of the most widely consumed seafoods in Japan and many other countries around the world. Due to the confirmed cases of salmon-induced allergy, the food sanitation law in Japan stipulates salmon as one of the specific food items for which labeling is recommended when used as an ingredient of processed foods. However, trout, the landlocked form of anadromous salmon, is not subject to the allergen-labeling requirements, even though both populations belong to a single species. Since no supporting data have been demonstrated to make a clear distinction between these two populations in terms of allergenicity, we comparatively examined their allergenic properties using sera from patients allergic to fish. Methods: Extracts of Oncorhynchus nerka from different habitats were obtained: kokanee (landlocked) and red salmon (anadromous). Control extracts were derived from four other species. This study focused on the (1) IgE-binding capacity of the fish extracts in patients' sera (n = 50), (2) ELISA inhibition test (n = 6), and (3) inhibition immunoblot test (n = 8) between the kokanee and red salmon. Results: The extracts from kokanee and red salmon showed the highest correlation with each other in terms of the IgE-binding capacity, and showed complete (100%) reciprocal cross-inhibition in the ELISA inhibition test. On immunoblotting, there was no marked difference in the staining pattern between the two extracts, and each IgE-binding band gradually disappeared when the patients' sera were preincubated with the counterpart antigen in a dose-dependent manner. Conclusions: These results suggest that kokanee has similar allergenic properties to red salmon. © 2009 Japanese Society of Allergology.

Background Although changes in the fine balance of allergen-specific T cells are crucial in the pathogenesis of allergic diseases, their roles in the allergic reaction to hen's eggs (HE) have not yet been fully analysed. Objective Using microarray technology, allergen-stimulated T cells from HE-allergic children were analysed to identify genes that are specifically up-regulated in these cells. Methods RNA from CD4(+) CD14(-) cells, fractionated from allergen-stimulated peripheral mononuclear cells, was analysed using a whole -genome microarray and real-time RT-PCR. The protein expression of selected genes was ascertained by flow cytometry. Results In microarray analyses of allergen-stimulated T cells, 43 genes were up-regulated in HE-allergic children but not in non-HE-allergic children. Among these, up-regulation of three genes, cytokine -inducible SH2-containing protein (CISH), nuclear factor of kappa light polypeptide gene enhancer in B-cell inhibitor Z (NFKBIZ) and B-cell CLL/lymphoma 2 (BCL2), was confirmed by real-time quantitative RT-PCR. CISH, but not NFKBIZ or BCL2, showed a significantly higher ratio of antigen-stimulated cell transcription over unstimulated cells in HE-allergic than in non-HE-allergic children (P < 0.01). Flow-cytometric analysis revealed that the percentage of CD25(+)CISH(+) cells in CD4(+) cells from patients with HE allergy was significantly higher than that in controls (P < 0.01). The expression level of CISH was significantly higher in IL-4(+) Th2 cells than in IFN-gamma(+) Th1 cells. Conclusion We noted that CISH expression in allergen-stimulated CD4(+) T cells from HE-allergic patients was significantly increased in both mRNA and protein levels compared with that from non-HE-allergic children.

Background: Children with allergy to raw egg white might tolerate low amounts of heated egg. Ovomucoid-specific IgE antibodies have been suggested to be predictors of whether children could tolerate heat-treated egg. Objective: The aim was to evaluate the clinical usefulness and added diagnostic value of measurements of IgE antibodies to egg white, ovalbumin, and ovomucoid in children with egg allergy. Methods: One hundred eight patients (median age, 34.5 months) with suspected egg allergy underwent double-blind, placebo-controlled food challenges with raw and heated egg. The outcomes of the challenges were related to the serum concentration of specific IgE antibodies and total IgE by using ImmunoCAP. Results: Reactions to heated egg white were observed in 38 patients (considered allergic to raw and heated egg), 29 patients reacted to only raw egg white, and 41 patients were tolerant. Correlation was observed between the serologic parameters studied. Receiver operating characteristic analysis showed that egg white ImmunoCAP was useful in the diagnosis of allergy to raw egg white. The positive decision point, based on 95% clinical specificity, was 7.4 kU(A)/L, and the negative decision point, based on 95% clinical sensitivity, was 0.6 kU(A)/L. For reaction to heated egg white, ovomucoid ImmunoCAP was superior. The positive decision point was 10.8 kU(A)/L, and the negative decision point was 1.2 kU(A)/L. Conclusions: Quantitative measurements of specific IgE antibodies to both egg white and ovomucoid and the evaluation against the suggested positive and negative decision points for specific IgE will be useful in the diagnosis of egg allergy.

Background: Phenytoin can induce diversified adverse reactions including generalized eruptions and the hypersensitivity syndrome. Delayed-type allergic mechanisms have been postulated to underlie these reactions. The tests most widely used to detect T-cell sensitization to drugs are the patch test and the lymphocyte transformation test (LTT), but their sensitivity is not sufficient. Simultaneous assessment of both the frequencies and the cytokine-producing phenotypes of allergen-specific T cells has become possible with the recently introduced carboxyfluorescein succinimidyl ester (CFSE) assay. Methods: Seven patients who presented with phenytoin-induced maculopapular exanthema with and without fever were included in this study. Peripheral blood mononuclear cells (PBMCs) were labeled with CFSE and cultured with phenytoin for seven days. The cells were stained with anti-CD4 and cytokine-specific monoclonal antibodies (MoAbs), and analyzed with FACSCalibur. Results: The phenytoin-specific proliferation of CD4+ cells in patients was significantly higher than in the four controls exposed to phenytoin, and in seven healthy children with no previous phenytoin intake. A significant difference in the percentages of CD4+ IFN-γ+ cells between patients and the seven healthy children was observed. The sensitivity and specificity of proliferation were 100% and 90.9%, and those of IFN-γ secretion were 71.4% and 100%, respectively. Conclusions: Phenytoin-specific proliferation may be detected with greater sensitivity by the CFSE dilution assay than the conventional LTT. The assay revealed that both CD4+ and CD4- T cells proliferated and produced IFN-γ and TNF-α after stimulation with phenytoin. The CFSE dilution assay might be useful for the diagnosis and understanding of drug hypersensitivity. ©2007 Japanese Society of Allergology.

Background The role of antigen-specific T cells in the allergic reaction to cow's milk or in tolerance induction is not yet fully understood. Objective This study was designed to analyse both cow's milk protein (CMP)-specific T cell proliferation and cytokine production simultaneously in children with cow's milk allergy (CMA) in comparison with subjects with various allergic backgrounds. Methods Carboxyfluorescein succinimidyl ester was used to detect cow's milk-specific T cells by flow cytometry. The intra-cytoplasmic cytokine production of these antigen-specific T cells was also analysed. Results Significant differences of both CMP-specific CD4(+) cell proliferation and cytokine production between CDAA and non-allergic children were observed. While the proliferative responses of children who recently outgrew CMA were not significantly different from those of patients, the patterns of cytokine production were similar to those of non-allergic children. Conclusion These results suggest that the presence of CMP-specific T cell clones per se does not produce CMA, but that the T-helper type 2-skewed pattern of those T cells is associated with adverse reactions. Although it is not possible to distinguish between individual patients with and without CMA on the basis of CFSE assays, these results contribute to the understanding of the pathogenesis and tolerance induction of CMA.

The group I allergens are a major cause of cedar pollen hypersensitivity in several geographic areas. Allergens from several taxa have been shown to cross-react. The goal of these studies was to compare the structural features of the shared and unique epitopes of the group 1 allergen from mountain cedar (Jun a 1) and Japanese cedar (Cry j 1). An array of overlapping peptides from the sequence of Jun a I and a panel of monoclonal anti-Cry j I antibodies were used to identify the IgE epitopes recognized by cedar-sensitive patients from Texas and Japan. IgE from Japanese patients reacted with peptides representing one of the two linear epitopes within the highly conserved beta-helical core structure and both epitopes within less ordered loops and turns near the N- and C-termini of Jun a 1. A three-dimensional (3D) model of the Cry j 1, based on the crystal structure of Jun a 1, indicated a similar surface exposure for the four described epitopes of Jun a I and the homologous regions of Cry j 1. The monoclonal antibodies identified another shared epitope, which is most likely conformational and a unique Cry j I epitope that may be the previously recognized glycopeptide IgE epitope. Defining the structural basis for shared and unique epitopes will help to identify critical features of IgE epitopes that can be used to develop mimotopes or identify allergen homologues for vaccine development. (C) 2005 Elsevier Ltd. All rights reserved.

Background: Hypersensitivity reactions to fish are a common food allergy, but IgE-binding activity to fish species have not been fully elucidated. The aim of this study was to identify fish with high binding activity to IgE in sera from Japanese fish-hypersensitive individuals. Methods: 38 children with a history of at least one episode of hypersensitivity after ingestion of fish were enrolled and 34 children with no history of reactions and negative IgE results for at least five kinds of fish antigen were included as controls. Using a radioallergosorbent test, we examined IgE-binding to each fish species using sera from fish-hypersensitive subjects. Fish were then graded according to IgE-binding activity. Results: Many fish species, including red salmon, silver salmon, yellowfin tuna, big eyed tuna, Atlantic tuna, saurel, skipper, yellowtail, Japanese sardine, bonita and mackerel had high IgE-binding activity. All of these fish are abundantly consumed in Japan. The hypersensitivity reactions experienced by many subjects occurred after ingestion of species with high IgE-binding activity. Only halibut (Osteichthyes) and sharks (Chondrichthyes) had low IgE-binding activity. Conclusions: A correlation was observed between IgE levels and expression of symptoms after fish ingestion. High consumption of salmon, tuna, scad (including saurel), skipper, yellowtail, sardine, bonita and mackerel in Japan might be the cause of the high IgE-binding activity of these species. The grades of fish species consumed widely in Japan are likely to be useful for nutritional instruction of fish-allergic patients. ©2006 Japanese Society of Allergology.

Background: Salmon roe (SR) anaphylaxis has often been reported and SR-containing foods are designated as 'recommended for allergic labeling' however, there have been no reports about its allergenicity, including its cross-reactivity. Because its cross-reactivity is controversial, clinicians are often confused concerning education regarding its dietary elimination. The purpose of this study was to examine the cross-reactivity between SR and other kinds of fish roe, salmon, or chicken egg (CE). Methods: We measured the specific-IgE to SR, herring roe (HR), pollock roe (PR), salmon and CE using RAST in 27 patients with a fish allergy and 26 control subjects. Then, using the sera of 2 patients with SR anaphylaxis, an ELISA inhibition study was performed to examine the cross-reactivity between SR and HR, PR, salmon or CE. We then compared the IgE binding patterns to SR between the anaphylaxis patients and fish allergy patients with immunoblotting. Results: There were positive correlations between SR and HR or PR, but none between SR and salmon or CE. In the ELISA study using sera from two patients with SR anaphylaxis, IgE-binding to SR was inhibited more than 50% only when the sera were pre-incubated with HR, inhibited almost 50% by PR in a dose-dependent manner, but not inhibited by CE or anisakis. Salmon inhibited the IgE binding to SR more than 50% in a SR-anaphylaxis patient. The IgE binding patterns to SR from anaphylaxis patients were almost identical and unlike those of patients with fish allergy. Conclusions: There was a cross-reactivity between SR and HR, but no relationship between SR and CE. ©2005 Japanese Society of Allergology.

Background An association between pollinosis and sensitivity to fruits and vegetables has been reported. Although Japanese cedar (Cryptomeria japonica) pollinosis is one of the most widespread diseases in Japan, there have been no reports demonstrating cross-reactivity between Japanese cedar pollen and other plant food. Objective The aim of this study was to demonstrate cross-reactivity between Japanese cedar pollen and tomato fruit (Lycopersicon esculentum) using RAST inhibition and immunoblot inhibition. Methods The RAST and immunoblot inhibition were performed using sera from patients with oral allergy syndrome (OAS) after ingesting fresh tomatoes. We identified some proteins that took part in cross-reactive IgE by the determination of N-terminal animo acid sequences and a homology search through the SWISS-PROT database. Results In the RAST inhibition. the bindings of IgE from the sera from four out of five (4/5) subjects to Japanese cedar pollen discs were inhibited by more than 50% by preincubation of the serum with tomato fruit extracts, Likewise. the IgE bindings to tomato fruit discs were inhibited more than 50% by Japanese cedar pollen extracts in 315 sera. In immunoblot inhibition, IgE binding activities of some protein bands on both membranes were decreased by heterologous inhibitors. However, the combinations of these protein bands involved in cross-reactivity were different between patients. Conclusion We have demonstrated cross-reactivity between Japanese cedar pollen and tomato fruit using RAST inhibition and immunoblot inhibition.

Background: Although the tomato fruit (Lycopersicon esculentum) has been widely investigated for breeding purposes, there have been few studies on tomato allergenicity. We attempted to identify the tomato fruit allergens and to compare the concentrations of IgE-binding proteins among the different growth stages with sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting. Methods. An immunoblot experiment on tomato fruit extracts was performed using sera from 11 patients with oral allergy syndrome (OAS) to tomatoes. Bands reacting with IgE from more than half of the OAS patients' sera were excised and subjected to determination of N-terminal amino acid sequences using the automated Edman degradation method. Moreover, we compared the concentrations of these proteins at each growth stage of the tomato fruit with SDS-PAGE and immunoblotting. Results: Four proteins binding with IgE from more than half of the OAS patients' sera were determined to be polygalacturonase 2A (PG2A), beta-fructofuranosidase, superoxide dismutase (SOD) and pectinesterase (PE). The concentrations of PG2A, beta-fructofuranosidase and PE were highest in the red ripening stage with both SDS-PAGE and immunoblotting. Conclusion: The concentrations of 3 of 4 tomato allergens increased during ripening. Copyright (C) 2002 S. Karger AG, Basel.

Background: We occasionally see egg-allergic children who develop contact urticaria to hen's egg despite the absence of the overt symptoms on ingestion. The mechanisms remain to be elucidated. Methods: Twenty-one subjects with positive reactions to 20-min patch tests for egg-white antigens were divided into subgroups with positive (n = 10) and negative (n = 11) results to oral challenge tests by the same antigens. We measured IgE antibody for egg white and its components, and IgE-binding activities to digestive enzyme-treated ovomucoid by RAST inhibition. Results: There were no significant differences in IgE antibody titers to egg white (positive vs negative: 30.3% vs 15.3%, P = 0.130), ovomucoid (21.5% vs 10.2%, P = 0.078), ovotransferrin (9.9% vs 3.7%, P = 0.105), and lysozyme (3.4% vs 2.9%, P = 0.944), except ovalbumin (16.8% vs 5.6%, P = 0.024), between the positive and negative subjects in the provocation tests. In contrast, the concentration (1.93 mu g/ml) of pepsin-treated ovomucoid needed for 50% RAST inhibition in the challenge-positive subjects was significantly (P = 0.0003) lower than that (114.9 mu g/ml) of negative subjects. Similar but less significant differences were obtained when ovomucoid fragments treated with chymotrypsin (0.91 mu g/ml vs 6.86 mu g/ml, P = 0.014) and trypsin (0.75 mu g/ml vs 4.67 mu g/ml, P = 0.041) were used as inhibitors. Conclusions: We suggest that IgE antibodies from subjects showing contact urticaria despite the absence of reactions to the ingestion of egg white recognize the epitope(s) unstable to digestive enzymes.

Background: We frequently encounter subjects without overt symptoms despite high IgE antibodies to egg white and its components. The measurements of these antibodies are not necessarily efficient for the diagnosis or the prediction of the outcome of egg allergy in children. Methods: Specific IgE antibodies to egg white and its components, including ovomucoid, ovalbumin, ovotransferrin and lysozyme, were measured by direct RAST assays. IgE-binding activity to ovomucoid degraded by pepsin, trypsin and chymotrypsin was examined by FAST inhibition. Thirty subjects were divided into two groups with positive (n = 18; mean age rt SD = 42 +/-25 months) and negative (n = 12; mean age +/- SD = 48 +/-31 months) oral challenge rests with egg white antigens. The individuals with positive results to the first challenge tests were given the second provocation tests at mean intervals of 32 months. IgE-binding activity of the sera collected on the first challenge to these ovomucoid fragments was compared between subjects with positive and negative reactions to the Follow-up challenge tests. Results: There were no significant differences in IgE antibody titers to egg white and its components between the positive and negative groups at the first and the second challenge tests. IgE-binding activity to ovomucoid digests after treatments with pepsin (p = 0.000008) and trypsin (p = 0.037), except chymotrypsin (p = 0.062), were significantly higher in subjects with positive challenge tests than in those with negative results. The difference was most remarkable in the IgE-binding to pepsin digests; the average concentrations (mean - SD and mean + SD) needed for 50% RAST inhibition in the positive group and in the negative group were 2.6 mu g/ml (0.3 and 25) and 94.2 mu g/ml (24.7 and 358.7), respectively. A significant difference was still observed in the inhibition tests using filtrates of pepsin digests with a membrane with MW 10,000 (p=0.014) and 3,000 (p = 0.042) of cutoff. The concentration (mean = 0.8, mean-SD = 0.2, mean + SD = 3.4; mu g/ml) of pepsin-treated ovomucoid required for 50% FAST inhibition in the subjects with positive second challenge results was significantly (p = 0.033) lower than that (mean = 6.8, mean - SD = 0.6, mean + SD = 73.9) of the negative group. Conclusion: IgE-binding activity to pepsin-digested ovomucoid was of diagnostic value to distinguish the challenge-positive subjects from the negative subjects. Subjects with high IgE-binding activity to pepsin-treated ovomucoid are unlikely to outgrow egg white allergy. Copyright (C) 1999 S. Karger AG, Basel.

Japanese cedar (Cryptomeria japonica) pollinosis has been a serious allergic disease in Japan. There are two kinds of Japanese cedar trees; the popular one is diploid, the less popular is triploid. These trees are not very different morphologically. However, the relative allergenicity of their pollens is unknown, although both major allergens, Cry j 1 and Cry j 2, have been purified and cloned from the diploid line. Triploid trees are sterile and the allergenicity of their pollen may differ. Using Japanese-cedar-allergic patient sera, we compared the concentration of these two major allergens in both kinds of pollen. Pollen collected from different years and regions was also studied. The results indicate that triploid tree pollen extract has lower concentrations of both major allergens; therefore, the pollen may be less allergenic.

Background: Immediate hypersensitive reactions induced by buckwheat ingestion are considered to be IgE-mediated. Some subjects, however, develop no immediate adverse reactions after buckwheat ingestion despite high levels of buckwheat-specific antigens IgE, The mechanism is unknown. Objective: To investigate the mechanisms. Methods: RAST for buckwheat and rice and RAST inhibition between these antigens were performed using sera from 23 buckwheat-sensitive subjects and 30 buckwheat-tolerant subjects who had IgE antibodies for both buckwheat and rice. Results: RAST values for buckwheat and rice were significantly correlated with each other (P < .01) in the buckwheat-tolerant group, but not in the buckwheat-sensitive group, This suggests the IgE antibodies from the subjects without any overt symptoms after buckwheat ingestion recognize the cross-reactive epitope between buckwheat and rice, whereas the IgE antibodies from those with immediate reactions to buckwheat ingestion do not. RAST inhibition assays were performed to evaluate this, RAST inhibition of heterogeneous combination of inhibitor and disc antigen such as rice and buckwheat was significantly smaller than that of homologous combination of rice and rice or buckwheat and buckwheat in the group with immediate symptoms after buckwheat ingestion. There was no significant difference in RAST inhibition between homologous and heterogeneous combinations in the group without the symptoms. Conclusions: There was cross-reactivity with IgE antibodies between buckwheat and rice and IgE antibodies from the buckwheat-tolerant subjects with high levels of IgE antibodies from the buckwheat might recognize the epitopes on buckwheat antigens which cross-react with rice antigens, whereas IgE antibodies from the buckwheat-sensitive subjects might bind to buckwheat-specific epitopes.

Cross-allergenicity between five cereal grains including rice, wheat, corn, Japanese millet (Panicum crus-galli L. var. frumentaceum Trin.) and Italian millet (Setaria italica Beauv. var. germanica schrad.) was examined by radioallergosorbent test (RAST) and RAST inhibition assay. There were significant close correlations between every combinations of RAST values for the five cereal grain extracts. RAST inhibition assay of each extract against RAST discs coupled with other cereal grain extracts indicated marked cross-reactivity of IgE binding between these cereal grain extracts. Rice protein 16KD (RP16KD) was shown to be one of major allergens in rice grain extracts by immunoblotting analysis, histamine release assay from human leukocytes and RAST inhibition. Next, the involvement of RP16KD in the cross-allergenicity between these cereals was investigated. RAST values for RP16KD significantly correlated with that for Italian millet as well as rice but not with those for corn and wheat. There was a trend of positive correlation between RAST values for RP16KD and Japanese millet. In the RAST inhibition assay using sera with positive RAST for these five cereal grain extracts and RP16KD, RP16KD inhibited IgE binding to these all cereal discs in a dose-dependent manner. Similarly, all of the five cereal grain extracts showed an effective decrease in IgE binding to the RP16KD disc. These results indicated possible participation of IgE binding structure on RP16KD in cross-allergenicity between these cereal grain extracts in the Poaceae family.