[A CASE OF FOOD-DEPENDENT EXERCISE-INDUCED ANAPHYLAXIS DUE TO POLISHED RICE].

A 14-year-old girl presented to our hospital with food-dependent exercise-induced anaphylaxis (FDEIA), possibly caused by rice. Despite experiencing four previous episodes of suspected FDEIA, she did not seek medical attention at her own discretion. On the fifth occurrence of symptoms, the general practitioner suspected FDEIA and referred the patient to our hospital. The only common factor in all five episodes was the consumption of rice, leading to the examination of the patient under suspicion of FDEIA caused by rice. Skin prick test results were positive for bran and polished rice, and exercise after consumption of polished rice resulted in anaphylaxis. Therefore, we diagnosed FDEIA caused by polished rice. Immunoblotting confirmed the presence of immunoglobulin E reacting with 14-16kDa rice bran protein in the patient's serum. The immunoblot inhibition test suggested that the rice bran protein to which the patient's serum reacted was also present in polished rice and no wash rice. As the patient may experience FDEIA after ingestion of no wash rice or rice flour, she was advised to eliminate these from her diet, treating them similarly to brown rice or polished rice.

[ANTIGENICITY OF SOY PROTEIN IN SOY-BASED INFANT FORMULA BONLACT® i].

BACKGROUND: The purpose of this study was to compare the antigenicity of Bonlact® i (BL) with that of defatted soy protein (SP) and soy protein isolate (SPI), which is the original source of BL, using sera from patients with soybean allergy. METHODS: Proteins were extracted from SP, SPI, and BL using PBS. Proteins in each sample were analyzed for antigenicity using inhibition ELISA with SP-specific IgE (sIgE), SDS-PAGE, and immunoblotting. Sere from patients with soybean allergy confirmed by an oral food challenge (OFC) (n=6, OFC+ Pt), and from patients who were positive for soy-sIgE without symptoms ( n = 7, sIgE+ Pt) were used for these assays. The cross-antigenicity of SP and BL with cow's milk (CM) proteins was also analyzed in the sera from patients with CM allergy using inhibition ELISA. RESULTS: SDS-PAGE showed that the proteins in BL produced a smear-like band in the low-molecular-weight region compared with that in SP and SPI. Inhibition ELISA against SP-sIgE showed that BL had a significantly lower inhibition rate than that of SP in both OFC+ Pt and sIgE+ Pt. Immunoblotting analysis showed that the bands of BL were thinner than those of SP and SPI. Additionally, SP and BL showed no cross-antigenicity with CM proteins. CONCLUSION: The proteins in BL was partially digested, and its antigenicity was lower than that of SP and SPI.

Evaluation of cross-reactivity between casein components using inhibition assay and in silico analysis.

BACKGROUND: We previously reported that the specific IgE levels to αs1-casein (CN) and ß-CN in patients with cow's milk allergy decreased with similar dynamics during oral immunotherapy. Therefore, we hypothesized that αs1- and ß-CN have strong cross-reactivity among CN components, despite the low similarity in the full-length amino acid sequences. METHODS: The αs1-, ß-, and κ-CN were purified from commercial cow's milk. We recruited 39 patients with cow's milk allergy, and the serum IgE levels for each CN component were measured by enzyme-linked immunosorbent assay (ELISA). Cross-reactivity between CN components was investigated by competitive ELISA against αs1-CN. Sequence homology between CN components at the peptide level was calculated using in silico analysis and quantified by the property distance (PD) value. RESULTS: The αs1-CN-specific IgE levels exhibited a strong positive correlation with the ß-CN-specific IgE (r = 0.945, P < .001). Complete competition was observed by ß-CN against αs1-CN, suggesting the presence of common epitopes between them. In silico analysis detected 24 peptide sets with PD values lower than 10 between αs1- and ß-CN, and 14 sets between αs1- and κ-CN. The amino acid sequences of αs1-CN (E61-E70) and ß-CN (I12-E21) that showed the lowest PD value (5.30) were present in the characteristic sequence known as casein phosphopeptide (CPP). CONCLUSION: We detected strong cross-reactivity between CN components. Furthermore, we found highly homologous sequences in the CPP region, which contains a core sequence of "SSSEE" with phosphorylated serine residues.

Changes in passively-sensitized basophil activation to αS1-casein after oral immunotherapy.

INTRODUCTION: Immune response to cow's milk allergen (CMA) has been analyzed mostly using crude milk antigen or a mixture of various caseins. This study aimed to assess the changes in the immunological response against αS1-casein during oral immunotherapy (OIT) and to investigate the mechanism of tolerance. METHODS: We have performed rush OIT to 39 patients with CMA and obtained the serum samples up to 3 years after OIT. Immunoglobulin E (IgE) and IgG4 antibodies specific to highly purified αS1-casein as well as passively-sensitized basophil activation were evaluated using the serial samples. Furthermore, we examined whether basophil activation led by the pre-OIT serum was suppressed by the post-OIT serum, or by the tolerant serum obtained from naturally outgrown patients. RESULTS: Specific IgE to αS1-casein was significantly reduced after OIT. Specific IgG4 (sIgG4) to αS1-casein was also detected in most of the pre-OIT sera, which was not significantly increased after OIT. Activation of passively-sensitized basophils to αS1-casein was significantly reduced after 2 years (14% ± 19%) and 3 years (19% ± 18%) post-OIT compared with pre-OIT (%CD63high basophils; 51% ± 27%). Furthermore, the addition of post-OIT or tolerant serum to pre-OIT serum significantly suppressed the basophil activation. This suppression was abrogated by washing the supernatant after passive sensitization, but not by depleting IgG antibodies from post-OIT or tolerant sera, nor by blocking FcγRIIb using an anti-FcγR antibody. CONCLUSIONS: αS1-casein-sIgG4 plays a minor role in tolerance mechanisms in cases of CMA; humoral factors other than antigen-sIgG4 may be involved.

Exercise-induced allergic reactions on desensitization to wheat after rush oral immunotherapy.

BACKGROUND: The effect of oral immunotherapy (OIT) on wheat allergy is promising in terms of the potential to obtain desensitization; however, the frequency of exercise-induced allergic reactions on desensitization (EIARDs) and the associated risk factors remain to be determined. METHODS: Twenty-five patients underwent rush OIT for wheat allergy, and 21 achieved the full-dose intake of wheat products (5 g of wheat protein). Exercise-provocation tests were repeatedly performed after the ingestion of a full-dose wheat product. The time-course of the levels of the specific IgEs (sIgE) to wheat extract, total gliadin, deamidated gliadin, recombinant gliadin components (α/ß-, γ- and ω-5-), and glutenin (high and low molecular weight) components was analyzed using ImmunoCAP® , ELISA, or IgE immunoblotting. RESULTS: Fourteen patients (66.7%) were diagnosed as EIARD+, which remained 5 years after rush OIT in 11 patients (52.4%). There were no differences in the clinical backgrounds of the EIARD+ and EIARD- patients. However, EIARD+ patients showed significantly higher sIgE levels to all gliadin and glutenin components than EIARD- patients before OIT. The sIgE levels to each component decreased equally after 1 and 2 years of OIT. On IgE immunoblotting, sera from all patients reacted to the multiple gluten bands, and some reacted to the water-soluble bands. The intensity of all IgE-reactive bands also became equally lighter after OIT. CONCLUSIONS: EIARDs were frequently observed and remained for a long period after successful OIT for wheat allergy. None of the specific wheat components were found to contribute to EIARDs.

INSOLUBILITY AND ALTERATION OF ALLERGENIC ACTIVITY OF WHEAT PROTEINS IN PROCESSED FOODS.

BACKGROUND: Food processing causes decomposition, denaturation or polymerization of protein, which may alter an allergic reaction. This study aimed to investigate the insolubility and alteration of wheat allergens in processed foods and the reactivity to patient sera. METHODS: We extracted proteins from wheat flour, udon and bread using different extracts and conducted SDS-polyacrylamide gel electrophoresis. IgE-immunoblotting was also conducted using sera from children with wheat allergy. RESULTS: Soluble protein was extracted from wheat flour, and gluten fractions were also extracted by adding SDS. However, no proteins were able to be extracted from udon or bread witout severing the disulfide bonds under reducing condition. Only trace amounts of protein were detected in the water after boiling udon noodles. The reactivity of IgE antibody to the extracted protein did not differ among the different processed food types. CONCLUSIONS: Wheat allergens became strongly insolubilized after gluten formation and heating. However, the reactivity of IgE antibody to each allergen was not affected by food processing. Further studies are needed for the effects on clinical symptoms.

Development of oral immunotherapy model using B10.A mice and egg white lysozyme.

Oral immunotherapy for food allergy has been the focus of a lot of attention recently. The patients have to eat allergenic food instead of eliminating it in this therapy and there is no established standard method yet. To promote clear understanding and improvement of oral immunotherapy, the present study using B10.A mice investigated the effect of multiple oral administration of a model antigen, egg-white lysozyme, on both the antibody response and the anaphylactic reaction induced by subsequent administration of lysozyme. Various doses of egg-white lysozyme (0-100 mg/mouse) were administered to mice intragastrically for 6 d; then additional lysozyme was administered via the intraperitoneal route in all groups. Lysozyme-specific antibody responses were promptly induced by the first oral administration and enhanced by intraperitoneal administration. An anaphylactic reaction was further induced in these sensitized mice by intragastric administration of lysozyme, and the symptoms of shock were compared in order to evaluate the effects of pretreatment. Interestingly, the decrease in rectal temperature which is one of the common anaphylactic symptoms in mice was suppressed in all of the oral pre-administration groups, and the effects were highest in the group that received 20 mg. Consequently, this study using B10.A mice has shown that sensitization can be induced by intragastric administration of lysozyme instead of oral tolerance; however, anaphylactic shock induced by subsequent intragastric administration of lysozyme is suppressed. This mouse model would be useful for assessing the method of oral immunotherapy.

Digestion and gastrointestinal absorption of the 14-16-kDa rice allergens.

The digestibility and gastrointestinal absorption of 14-16-kDa rice allergens (RAs) were investigated. RAs and bovine serum albumin (BSA) were first evaluated for their digestibility. BSA was digested completely by in vitro incubation with some proteases, but RAs remained almost intact. Administered orally (20 mg per mouse), intact RAs were clearly detected in the small intestine even 60 min after the administration, the amount of total RAs in the small intestine being estimated to be 0.59 mg. RAs were then biotinylated and infused into the duodenal lumen of anesthetized mice, and portal blood was collected. The RA concentrations in the portal plasma were respectively estimated to be 0.4-0.9 and 0.3-2.5 microg/ml for 0.4 and 4 mg doses. These results suggest that RAs are highly resistant to digestive enzymes and that about 1/100 of orally administered RAs remain intact in the small intestine, while at least 1/1,000-1/10,000 is absorbed and delivered into circulated blood.

Degradation of soluble proteins including some allergens in brown rice grains by endogenous proteolytic activity during germination and heat-processing.

The effect of germination and subsequent heat-processing on the degradation of soluble proteins, including some allergenic proteins, in brown rice grains was investigated. The content of soluble proteins, including 14-16-kDa and 26-kDa allergens, in the germinated and processed brown rice grains (GPR) was much lower than that of non-germinated brown rice. These proteins in brown rice grains were also much lower after subsequent heat-processing during the manufacturing process. The protease activity of germinated brown rice (GR) was detected and increased 1.5 times after germination. The optimum pH values for degradation of the 26-kDa and 14-16-kDa allergens in the GR grains were 4 and between 5 and 7, respectively. These results suggest that the decrease in the soluble proteins and allergens was induced in part by proteolytic degradation. The presence of a detergent enhanced the proteolytic degradation of the soluble proteins, especially of the 26-kDa allergen, in the brown rice grains. The degradation of the 26-kDa allergen was weakly inhibited by NEM, suggesting cysteine protease(s) may have been involved in its degradation. These results suggest that the two abundant allergens were degraded in a different manner and probably by different proteases in the grains during germination.