Sustained silencing peanut allergy by xanthopurpurin is associated with suppression of peripheral and bone marrow IgE-producing B cell.

Introduction: Peanut allergy is an immunoglobulin E (IgE) mediated food allergy. Rubia cordifolia L. (R. cordifolia), a Chinese herbal medicine, protects against peanut-induced anaphylaxis by suppressing IgE production in vivo. This study aims to identify IgE-inhibitory compounds from the water extract of R. cordifolia and investigate the underlying mechanisms using in vitro and in vivo models. Methods: Compounds were isolated from R. cordifolia water extract and their bioactivity on IgE production was assessed using a human myeloma U266 cell line. The purified active compound, xanthopurpurin (XPP), was identified by LC-MS and NMR. Peanut-allergic C3H/HeJ mice were orally administered with or without XPP at 200µg or 400µg per mouse per day for 4 weeks. Serum peanut-specific IgE levels, symptom scores, body temperatures, and plasma histamine levels were measured at challenge. Cytokines in splenocyte cultures were determined by ELISA, and IgE + B cells were analyzed by flow cytometry. Acute and sub-chronic toxicity were evaluated. IL-4 promoter DNA methylation, RNA-Seq, and qPCR analysis were performed to determine the regulatory mechanisms of XPP. Results: XPP significantly and dose-dependently suppressed the IgE production in U266 cells. XPP significantly reduced peanut-specific IgE (>80%, p <0.01), and plasma histamine levels and protected the mice against peanut-allergic reactions in both early and late treatment experiments (p < 0.05, n=9). XPP showed a strong protective effect even 5 weeks after discontinuing the treatment. XPP significantly reduced the IL-4 level without affecting IgG or IgA and IFN-γ production. Flow cytometry data showed that XPP reduced peripheral and bone marrow IgE + B cells compared to the untreated group. XPP increased IL-4 promoter methylation. RNA-Seq and RT-PCR experiments revealed that XPP regulated the gene expression of CCND1, DUSP4, SDC1, ETS1, PTPRC, and IL6R, which are related to plasma cell IgE production. All safety testing results were in the normal range. Conclusions: XPP successfully protected peanut-allergic mice against peanut anaphylaxis by suppressing IgE production. XPP suppresses murine IgE-producing B cell numbers and inhibits IgE production and associated genes in human plasma cells. XPP may be a potential therapy for IgE-mediated food allergy.

Double-Blind, Placebo-Controlled Study of E-B-FAHF-2 in Combination With Omalizumab-Facilitated Multiallergen Oral Immunotherapy.

BACKGROUND: Oral immunotherapy (OIT) is limited by adverse events, and most patients require continued treatment to maintain their increased threshold. Adjunctive treatments have been explored to increase the safety and efficacy of OIT. OBJECTIVE: This study aimed to determine the safety and efficacy of enhanced, butanol purified Food Allergy Herbal Formula-2 (E-B-FAHF-2) for inducing remission in subjects undergoing omalizumab-facilitated multiallergen OIT (multi-OIT). METHODS: In this double-blind, placebo-controlled clinical trial, subjects were randomized 1:1 to receive either E-B-FAHF-2 or placebo, starting 2 months before OIT and continuing throughout OIT. All subjects received a 4-month course of omalizumab, starting 2 months before OIT through the 2-month OIT build-up phase. After 24 months of multi-OIT (maintenance dose of 1000 mg of each allergen), desensitization and remission were assessed. The primary objective was to determine if subjects in the E-B-FAHF-2 group (EOIT) were more likely than the placebo group (OIT) to develop remission to all 3 allergens treated with multi-OIT, as defined by the absence of dose-limiting symptoms to a cumulative dose of 4444 mg of protein after discontinuing treatment for 3 months. RESULTS: Thirty-three subjects were randomized. A total of 63.6% were desensitized to 4444 mg of protein for each allergen at 26 months, and 24.2% met the primary outcome of remission at 29 months, with no difference between the treatment groups. There was good adherence (>85%) with study medications, with no difference between the treatment groups. There was no difference in reported overall adverse events between the treatment groups. CONCLUSION: Omalizumab-facilitated multifood OIT was safe and effective, and remission was achieved in about a quarter of subjects. However, outcomes were not improved by the addition of E-B-FAHF-2.

Soy Gly m 8 sIgE Has Limited Value in the Diagnosis of Soy Allergy in Peanut Ara h 2-Sensitized Adults.

INTRODUCTION: Recently, specific IgE (sIgE) sensitization against Gly m 8 (soy 2S albumin) has been described as a good diagnostic marker for soy allergy (SA). The aim of this study was to evaluate the diagnostic value of Gly m 8 by determining the sensitization profiles based on the homologues soy allergens Bet v 1, Ara h 1, Ara h 2, and Ara h 3. METHODS: Thirty soy-allergic adults were included; sIgE to total soy extract, Gly m 8, Gly m 4, Gly m 5, Gly m 6, Bet v 1, Ara h 1, Ara h 2, and Ara h 3 were determined. Sensitization patterns were analyzed and determined. The clinical relevance of sIgE of Gly m 8 sensitization was measured by assessing its capacity to degranulate basophils in Gly m8-sensitized patients by an indirect basophil activation test (iBAT). RESULTS: Based on the sIgE patterns of sensitization, two groups of SA patients were identified: (i) peanut-associated SA group (all patients were sensitized to one or more of the peanut compounds) and (ii) non-peanut/PR-10-associated SA group (22 patients were sensitized to Gly m 4 and Bet v 1 but not to any of the peanut compounds). A high and significant correlation between total soy extract and Gly m 6 (R2 = 0.97), Gly m 5 (R2 = 0.85), and Gly m 8 (R2 = 0.78) was observed. A nonsignificant correlation was observed between the levels of sIgE of Gly m 8 versus Ara h2. The iBAT results showed that Gly m 8 did not induce basophil degranulation in any of the peanut-associated patients, indicating that the Gly m8 sensitizations were not clinically relevant. CONCLUSIONS: Gly m 8 was not a major allergen in the selected soy-allergic population. The iBAT results indicated that Gly m 8 was not able to induce basophil degranulation in sIgE Gly m 8-sensitized soy-allergic patients. Thus, Gly m 8 would have no added value in the diagnosis of SA in the present study population.

Targeting Ara h 2 with human-derived monoclonal antibodies prevents peanut-induced anaphylaxis in mice.

BACKGROUND: Peanut allergy is a type-I hypersensitivity immune reaction mediated by the binding of peanut allergens to IgE-FcεRI complexes on mast cells and basophils and by their subsequent cellular degranulation. Of all major peanut allergens, Ara h 2 is considered the most anaphylactic. With few options but allergen avoidance, effective treatment of allergic patients is needed. Passive immunotherapy (herein called PIT) based on prophylactic administration of peanut-specific monoclonal antibodies (mAbs) may present a promising treatment option for this under-served disease. METHOD: Fully human recombinant anti-peanut IgG mAbs were tested in mice sensitized to peanut allergen extract. Allergic mice received intravenous immunotherapy with anti-peanut Ara h 2-specific IgG1 or IgG4 mAbs cocktails, and were then challenged by a systemic injection of high-dose peanut allergen extract. The protection from allergic anaphylaxis was measured by monitoring the core body temperature. RESULTS: PIT with peanut-specific mAbs was associated with a significant and dose-dependent reduction of anaphylactic reactions in peanut-sensitized mice challenged with peanut allergen extract. Complete protection was observed at doses approximately 0.3-0.6 mg mAbs. Mixtures of mAbs were more effective than single mAbs, and effective treatment could be obtained with mAbs of both IgG1 and IgG4 subclasses. The therapeutic effect of anti-Ara h 2 mAbs was based on allergen neutralization and independent of the Fcγ receptor and mast-cell inhibition. CONCLUSION: This is the first report that shows that human-derived anti-peanut mAbs can prevent allergic anaphylaxis in mice. The study demonstrates that neutralizing allergenic epitopes on Ara h 2 by mAbs may represent a promising treatment option in peanut-allergy.

Development of sensitization to peanut and storage proteins and relation to markers of airway and systemic inflammation: A 24-year follow-up.

BACKGROUND: Long-time data of peanut allergy over time is sparse. We aimed to study the longitudinal development of sensitization to peanut extract and storage protein allergen molecules and associations with asthma status, airway and systemic inflammation markers. METHODS: The Swedish birth cohort BAMSE followed 4089 participants with questionnaires, clinical investigations and blood sampling between 0 and 24 years. Information on (i) background factors at 2 months, (ii) peanut allergy symptoms and IgE data (ImmunoCAP) at 4, 8, 16, and 24 years, and (iii) IgE to storage proteins, lung function data including exhaled nitric oxide (FENO) as well as systemic inflammatory markers at 24 years of age were collected. RESULTS: The prevalence of peanut extract sensitization, defined as IgE ≥ 0.35 kUA /L, was 5.4%, 8.0%, 7.5%, and 6.2% at 4, 8, 16, and 24 years of age, respectively. Between 8 and 24 years of age, (33/1565) participants developed IgE-ab to peanut extract (median 1,4, range 0.7-2.6 kUA /L), and among those 85% were also sensitized to birch. Only six individuals developed sensitization to Ara h 2 (≥0.1 kUA /L) between 8 and 24 years of age, of whom three had an IgE-ab level between 0.1-0.12 kUA /L. Storage protein sensitization was associated with elevated FENO, blood eosinophils and type 2 inflammation-related systemic proteins. CONCLUSION: Sensitization to peanut extract after 4 years of age is mainly induced by birch cross-sensitization and IgE to Ara h 2 rarely emerges after eight years of age. Storage protein sensitization is associated with respiratory and systemic inflammation.

Comprehensive Care and Education Can Improve Nutritional and Growth Outcomes in Children with Persistent Cow's Milk, Egg, or Peanut Allergies: A Five-Year Follow-Up Study of Nutritional Status.

BACKGROUND: Data suggest that food allergies greatly impact a child's health and growth due to inadequate nutrient intake. Our study aimed to establish the long-term outcome of children with food allergies compared to a control group. METHODS: This study was a retrospective cohort study with longitudinal follow-up with a mean period of 4.85 years from the diagnosis to the last study visit. The patients' nutritional intake was assessed using a three-day food diary and analysed by a dietitian. Patients (61 boys and 33 girls, mean age 6.9 years) had a single food allergy including 21 patients with cow's milk, 34 with egg, and 39 with peanut allergies. The control group included 36 children (19 boys and 17 girls, mean age 8.03 years). Blood analysis was performed on all participants. RESULTS: Data from our study showed that patients with cow's milk, egg or peanut allergies had normal growth and achieved catch-up growth from the diagnosis until the last study visit. In the cow's milk allergy group, the allergy was shown to affect calcium intake (p < 0.05), while egg and peanut allergies did not impact the dietary intake of nutrients. None of the investigated food allergies affected blood results (p < 0.05). CONCLUSIONS: In the present study, we showed that single food allergies do not compromise growth in children if they are provided with appropriate support and that the affected children reach catch-up growth from the diagnosis.

Fatty-Acid-Based Membrane Lipidome Profile of Peanut Allergy Patients: An Exploratory Study of a Lifelong Health Condition.

Peanut allergy is a lifelong, increasingly prevalent, and potentially life-threatening disease burdening families and communities. Dietary, particularly polyunsaturated fatty acids (PUFAs), intakes can exert positive effects on immune and inflammatory responses, and the red blood cell (RBC) membrane lipidome contains stabilized metabolic and nutritional information connected with such responses. The fatty-acid-based membrane lipidome profile has been exploratorily evaluated in a small cohort of patients (eight males and one female, age range 4.1−21.7 years old, body mass index BMI < 25) with angioedema and/or anaphylaxis after peanut ingestion. This analysis was performed according to an ISO 17025 certified robotic protocol, isolating mature RBCs, extracting membrane lipids, and transforming them to fatty acid methyl esters for gas chromatography recognition and quantification. Comparison with a group of age- and BMI-matched healthy individuals and with benchmark interval values of a healthy population evidenced significant differences, such as higher levels of ω-6 (arachidonic acid), lower values of ω-3 eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), together with an increased ω-6/ω-3 ratio in allergic patients. A significant inverse correlation was also found between specific immunoglobulin E (IgE) levels and ω-6 di-homo-gamma-linolenic acid (DGLA) and total PUFAs. Results of this preliminary study encourage screenings in larger cohorts, also in view of precision nutrition and nutraceuticals strategies, and stimulate interest to expand basic and applied research for unveiling molecular mechanisms that are still missing and individuating treatments in chronic allergic disorders.

Anti-IgE effect of small-molecule-compound arctigenin on food allergy in association with a distinct transcriptome profile.

BACKGROUND: Excessive production of IgE plays a major role in the pathology of food allergy. In an attempt to identify anti-IgE natural products, Arctium Lappa was one of the most effective herbs among approximately 300 screened medicinal herbs. However, little is known about its anti-IgE compounds. OBJECTIVE: To identify compounds from Arctium Lappa for targeted therapy on IgE production and explore their underlying mechanisms. METHODS: Liquid-liquid extraction and column chromatographic methods were used to purify the compounds. IgE inhibitory effects were determined on IgE-producing human myeloma U266 cells, peanut-allergic murine model and PBMCs from food-allergic patients. Genes involved in IgE inhibition in PBMCs were studied by RNA sequencing. RESULTS: The main compounds isolated were identified as arctiin and arctigenin. Both compounds significantly inhibited IgE production in U266 cells, with arctigenin the most potent (IC50=5.09µg/mL). Arctigenin (at a dose of 13 mg/kg) markedly reduced peanut-specific IgE levels, blocked hypothermia and histamine release in a peanut-allergic mouse model. Arctigenin also significantly reduced IgE production and Th2 cytokines (IL-5, IL-13) by PBMCs. We found 479 differentially expressed genes in PBMCs with arctigenin treatment (p < .001 and fold-change ≥1.5), involving 24 gene ontology terms (p < .001, FDR <0.05); cell division was the most significant. Eleven genes including UBE2C and BCL6 were validated by qPCR. CONCLUSION: Arctigenin markedly inhibited IgE production in U266 cells, peanut-allergic murine model and PBMCs from allergic patients by down-regulating cell division, cell cycle-related genes and up-regulating anti-inflammatory factors.

Early Introduction of Food Allergens and Risk of Developing Food Allergy.

There is increasing evidence that early introduction of allergenic foods may decrease the risk of developing IgE-mediated food allergy. Patterns of food introduction before the 2015 publication of the Learning Early about Peanut Allergy (LEAP) trial are not well-studied, but are important as a baseline for evaluating subsequent changes in infant feeding practices and potentially food allergy. We performed a retrospective longitudinal study using data from a multicenter cohort of infants hospitalized with bronchiolitis between 2011-2014. The primary outcomes were IgE-mediated egg or peanut allergy by age 3 years. Of 770 participants included in the analysis, 635 (82%) introduced egg, and 221 (27%) introduced peanut by age 12 months per parent report. Four participants had likely egg allergy, and eight participants had likely peanut allergy by age 3 years. Regular infant egg consumption was associated with less egg allergy. The association was suggestive for infant peanut consumption with zero peanut allergy cases. Overall, our results suggest that early introduction of peanut was uncommon before 2015. Although limited by the small number of allergy cases, our results suggest that early introduction of egg and peanut are associated with a decreased risk of developing food allergy, and support recent changes in practice guidelines.

Study protocol of a phase 2, dual-centre, randomised, controlled trial evaluating the effectiveness of probiotic and egg oral immunotherapy at inducing desensitisation or sustained unresponsiveness (remission) in participants with egg allergy compared with placebo (Probiotic Egg Allergen Oral Immunotherapy for Treatment of Egg Allergy: PEAT study).

INTRODUCTION: Egg allergy is the most common food allergy in children but recent studies have shown persistence or delayed resolution into adolescence. As there is currently no effective long-term treatment, definitive treatments that improve quality of life and prevent fatalities for food allergies are required. We have previously shown that a novel treatment comprising a combination of the probiotic Lactobacillus rhamnosus CGMCC 1.3724 with peanut oral immunotherapy (OIT) is highly effective at inducing sustained unresponsiveness, with benefit persisting to 4 years after treatment cessation in the majority of initial treatment responders. In this study, we plan to extend the probiotic food OIT platform to another allergen, namely egg. We describe the protocol for a phase 2, dual-centre, randomised, controlled trial evaluating the effectiveness of probiotic and egg OIT at inducing desensitisation or sustained unresponsiveness (remission) in participants with egg allergy compared with placebo. METHODS AND ANALYSIS: 80 participants aged 5-30 years of age with current egg allergy confirmed by double-blind placebo-controlled food challenge at study screening will be recruited from Australia and Singapore. There are two intervention arms-probiotic and egg OIT (active) or placebo. Interventions are administered once daily for 18 months. The primary outcome is the proportion of participants who attain 8-week sustained unresponsiveness in the active group versus placebo group. ETHICS AND DISSEMINATION: This study has been approved by the Human Research Ethics Committees at the Royal Children's Hospital (HREC 2019.082) and the National Healthcare Group Domain Specific Review Board (2019/00029). Results will be published in peer-reviewed journals and disseminated via presentations at international conferences. TRIAL REGISTRATION NUMBER: ACTRN12619000480189.

Identification and Purification of Novel Low-Molecular-Weight Lupine Allergens as Components for Personalized Diagnostics.

Lupine flour is a valuable food due to its favorable nutritional properties. In spite of its allergenic potential, its use is increasing. Three lupine species, Lupinus angustifolius, L. luteus, and L. albus are relevant for human nutrition. The aim of this study is to clarify whether the species differ with regard to their allergen composition and whether anaphylaxis marker allergens could be identified in lupine. Patients with the following characteristics were included: lupine allergy, suspected lupine allergy, lupine sensitization only, and peanut allergy. Lupine sensitization was detected via CAP-FEIA (ImmunoCAP) and skin prick test. Protein, DNA and expressed sequence tag (EST) databases were queried for lupine proteins homologous to already known legume allergens. Different extraction methods applied on seeds from all species were examined by SDS-PAGE and screened by immunoblotting for IgE-binding proteins. The extracts underwent different and successive chromatography methods. Low-molecular-weight components were purified and investigated for IgE-reactivity. Proteomics revealed a molecular diversity of the three species, which was confirmed when investigated for IgE-reactivity. Three new allergens, L. albus profilin, L. angustifolius and L. luteus lipid transfer protein (LTP), were identified. LTP as a potential marker allergen for severity is a valuable additional candidate for molecular allergy diagnostic tests.

Ara h 2-specific IgE is superior to whole peanut extract-based serology or skin prick test for diagnosis of peanut allergy in infancy.

BACKGROUND: Screening of high-risk infants for peanut allergy (PA) before introduction is now recommended in the United States, but the optimal approach is not clear. OBJECTIVE: We sought to compare the diagnostic test characteristics of peanut skin prick test (SPT), peanut-specific IgE (sIgE), and sIgE to peanut components in a screening population of infants before known peanut exposure. METHODS: Infants aged 4 to 11 months with (1) no history of peanut ingestion, testing, or reaction and (2) (a) moderate-severe eczema, (b) history of food allergy, and/or (c) first-degree relative with a history of PA received peanut SPT, peanut-sIgE and component-IgE testing, and, depending on SPT wheal size, oral food challenge or observed feeding. Receiver-operator characteristic areas under the curve (AUCs) were compared, and diagnostic sensitivity and specificity were calculated. RESULTS: A total of 321 subjects completed the enrollment visit (median age, 7.2 months; 58% males), and 37 (11%) were found to have PA. Overall, Ara h 2-sIgE at a cutoff point of 0.1 kUa/L discriminated between allergic and nonallergic best (AUC, 0.96; sensitivity, 94%; specificity, 98%), compared with peanut-sIgE at 0.1 kUa/L (AUC, 0.89; sensitivity, 100%; specificity, 78%) or 0.35 kUa/L (AUC, 0.91; sensitivity, 97%; specificity, 86%), or SPT at wheal size 3 mm (AUC, 0.90; sensitivity, 92%; specificity, 88%) or 8 mm (AUC, 0.87; sensitivity, 73%; specificity, 99%). Ara h 1-sIgE and Ara h 3-sIgE did not add to prediction of PA when included in a model with Ara h 2-sIgE, and Ara h 8-sIgE discriminated poorly (AUC, 0.51). CONCLUSIONS: Measurement of only Ara h 2-sIgE should be considered if screening of high-risk infants is performed before peanut introduction.

Induction of Peanut Allergy Through Inhalation of Peanut in Mice.

Peanut (PN) allergy is a common life-threatening disease; however, our knowledge on the immunological mechanisms remains limited. Here, we describe the first mouse model of inhalation-driven peanut allergy. We administered PN flour intranasally to naïve wild-type mice twice a week for 4 weeks, followed by intraperitoneal challenge with PN extract. Exposure of mice to PN flour sensitized them without addition of adjuvants, and mice developed PN-specific IgE, IgG1, and IgG2a. After challenge, mice displayed lower body temperature and other clinical signs of anaphylaxis. This inhalation model is an ideal system to allow for future examination of immunological mechanisms critical for the development of PN allergy.

Preventing food allergy in infancy and childhood: Systematic review of randomised controlled trials.

BACKGROUND: This systematic review of ways to prevent immediate-onset/IgE-mediated food allergy will inform guidelines by the European Academy of Allergy and Immunology (EAACI). METHODS: The GRADE approach was used. Eleven databases were searched from 1946 to October 2019 for randomized controlled trials (and large prospective cohort studies in the case of breastfeeding). The studies included heterogeneous interventions, populations, and outcomes and so were summarized narratively. RESULTS: Forty-six studies examined interventions to reduce the risk of food allergy in infancy (up to 1 year) or early childhood. The following interventions for pregnant or breastfeeding women and/or infants may have little to no effect on preventing food allergy, but the evidence is very uncertain: dietary avoidance of food allergens, vitamin supplements, fish oil, probiotics, prebiotics, synbiotics, and emollients. Breastfeeding, hydrolyzed formulas, and avoiding cow's milk formula may not reduce the risk of cow's milk protein allergy; however, temporary supplementation with cow's milk formula in the first week of life may increase the risk of cow's milk allergy. Introducing well-cooked egg, but not pasteurized raw egg, from 4 to 6 months probably reduces the risk of hen's egg allergy. Introducing regular peanut consumption into the diet of an infant at increased risk beginning from 4 to 11 months probably results in a large reduction in peanut allergy in countries with a high prevalence. These conclusions about introducing peanut are based on moderate certainty evidence, from single trials in high-income countries. CONCLUSIONS: Sixty percent of the included studies were published in the last 10 years, but much still remains to be understood about preventing food allergy. In particular, there is a need to validate the potential benefits of early introduction of food allergens in a wider range of populations.

IgE cross-reactivity measurement of cashew nut, hazelnut and peanut using a novel IMMULITE inhibition method.

Background Tree nut-allergic individuals are often sensitised towards multiple nuts and seeds. The underlying cause behind a multi-sensitisation for cashew nut, hazelnut, peanut and birch pollen is not always clear. We investigated whether immunoglobulin E antibody (IgE) cross-reactivity between cashew nut, hazelnut and peanut proteins exists in children who are multi-allergic to these foods using a novel IMMULITE®-based inhibition methodology, and investigated which allergens might be responsible. In addition, we explored if an allergy to birch pollen might play a role in this co-sensitisation for cashew nut, hazelnut and peanut. Methods Serum of five children with a confirmed cashew nut allergy and suffering from allergic symptoms after eating peanut and hazelnut were subjected to inhibition immunoassays using the IMMULITE® 2000 XPi. Serum-specific IgE (sIgE) to seed storage allergens and pathogenesis-related protein 10 (PR10) allergens were determined and used for molecular multicomponent allergen correlation analyses with observed clinical symptoms and obtained inhibition data. Results IgE cross-reactivity was observed in all patients. Hazelnut extract was a strong inhibitor of cashew nut sIgE (46.8%), while cashew nut extract was less able to inhibit hazelnut extract (22.8%). Peanut extract showed the least inhibition potency. Moreover, there are strong indications that a birch pollen sensitisation to Bet v 1 might play a role in the observed symptoms provoked upon ingestion of cashew nut and hazelnut. Conclusions By applying an adjusted working protocol, the IMMULITE® technology can be used to perform inhibition assays to determine the risk of sIgE cross-reactivity between very different food components.

Effectiveness of different proteases in reducing allergen content and IgE-binding of raw peanuts.

The effectiveness of some non-specific proteases in reducing raw peanut allergenicity was investigated. Peanut kernels were treated by Alcalase, papain, Neutrase and bromelain, respectively. The residues of major peanut allergens Ara h 1, Ara h 2 and Ara h 6 were determined by sandwich ELISA and SDS-PAGE, and the allergenicities of treated peanuts were compared to that of untreated peanuts by western blot. All tested proteases were effective in reducing Ara h 1, but their effectiveness in hydrolyzing Ara h 2 and Ara h 6 varied greatly. The maximal reductions of extractable Ara h 1, Ara h 2 and Ara h 6 were 100%, 100% and 99.8%, respectively, achieved by Alcalase hydrolysis. Alcalase was more effective in overall allergenicity reduction; bromelain and Neutrase were the least effective in reducing Ara h 2 and Ara h 6, respectively. The hydrolysis of original allergens also produced some smaller peptides with strong IgE-binding.

Fructo-Oligosaccharides Modify Human DC Maturation and Peanut-Induced Autologous T-Cell Response of Allergic Patients In Vitro.

Background: Dendritic cells (DCs) play an important role in antigen presentation, and are an interesting target for immune-modulation in allergies. Short- and long-chain fructo-oligosaccharides (scFOS/lcFOS, FF) have immunomodulatory capacities, and may influence the outcome of DC antigen presentation. Objective: This study investigated the effect of FF during DC maturation and allergen presentation using cells of peanut-allergic patients in an autologous DC-T cell assay. Methods: CD14+ and CD4+ T cells were isolated from peanut-allergic patients. CD14+ monocytes were differentiated into immature DCs (imDCs), and matured (matDCs) in the presence or absence of crude peanut-extract (CPE) and/or FF, and co-cultured in an autologous DC-T cell assay. T cell polarization, proliferation and cytokine production were measured. Results: Expression of maturation surface molecule markers on matDCs was not affected by CPE and/or FF. By contrast, the IL-10 secretion by matDCs increased compared to imDCs, upon exposure to CPE and FF compared to CPE alone. Also the IP-10 secretion increased in CPE/FF-matDCs compared to imDC. CPE-matDCs enhanced IL-13 release in the DC-T-cell assay and Treg polarization in presence or absence of FF. CPE/FF-DCs tended to increase the Treg/Th1 and Treg/Th2 ratios compared to matDCs. The proliferation of both Treg and Th2 cells tended to increase when T cells were co-cultured with CPE-matDCs compared to matDCs, which became significant when CPE-matDCs were also exposed to FF and a same tendency was shown for Th1 proliferation. Conclusion: Only in the presence of FF, CPE-matDCs produced increased regulatory and Th1-related mediators. CPE-matDCs modified T cell polarization and proliferation, and additional exposure to FF tended to enhance Treg/Th2 and Treg/Th1 ratios instructed by CPE/FF-matDCs. However this effect was not strong enough to suppress CPE-matDCs induced IL-13 release by Th-cells. This indicates the ability of FF to modify DC maturation in the presence of an allergen supporting a more Treg/Th1 prone direction of the successive allergen specific Th2 cell response.