Extracellular adenosine triphosphate skews the T helper cell balance and enhances neutrophil activation in mice with food allergies.

Exposure to food allergens elicits fast changes in the intestinal microenvironment, which guides the development of allergic reactions. Investigating the key information about these changes may help in better understanding food allergies. In this research, we explored the relationship between a food allergy and extracellular adenosine triphosphate (ATP), a danger molecule that has been proved to regulate the onset of allergic asthma and dermatitis but has not been studied in food allergies, by developing a unique animal model through allergen-containing diet feeding. After consuming an allergen-containing diet for 7 days, the allergic mice exhibited severe enteritis with elevated luminal ATP levels. The dysregulated luminal ATP worsened food-induced enteritis by enhancing Th17 cell responses and increasing mucosal neutrophil accumulation. In vitro experiments demonstrated that ATP intervention facilitated Th17 cell differentiation and neutrophil activation. In addition, the diet-induced allergy showed noticeable gut dysbiosis, characterized by decreased microbial diversity and increased diet-specific microbiota signatures. As the first, we show that food-induced enteritis is associated with an elevated concentration of luminal ATP. The dysregulated extracellular ATP exacerbated the enteritis of mice to a food challenge by manipulating intestinal Th17 cells and neutrophils.

Lipid matrix-specific pretreatment method for enhancing the extractability and allergenicity maintenance of bovine milk allergens in ELISA detection.

The presence of various components in the food matrix makes allergen detection difficult and inaccurate, and pretreatment is an innovative breakthrough point. Food matrices were categorised based on their composition. Subsequently, a pretreatment method was established using a combination of ultrasound-assisted n-hexane degreasing and weakly alkaline extraction systems to enhance the detection accuracy of bovine milk allergens. Results showed that more allergens were obtained with less structural destruction, as demonstrated using immunological quantification and spectral analysis. Concurrently, allergenicity preservation was confirmed through liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis, a KU812 cell degranulation model, and western blotting. The method exhibited good accuracy (bias, 8.47%), repeatability (RSDr, 1.52%), and stability (RSDR, 5.65%). In foods with high lipid content, such as chocolate, the allergen content was 2.29-fold higher than that of commercial kits. Laser confocal scanning microscopy (LCSM) and scanning electron microscopy (SEM) analyses revealed a significant decrease in fat content after post-pretreatment using our method. In addition, colloidal stability surpassed that achieved using commercial kits, as indicated through the PSA and zeta potential results. The results demonstrated the superiority of the extractability and allergenicity maintenance of lipid matrix-specific pretreatment methods for improving the accuracy of ELISA based allergen detection in real food.

Effects of medicine food homologous materials on food allergy-associated factors: intestinal oxidative stress, intestinal inflammation and Th2 immune response.

BACKGROUND: Food allergies could be regulated via Th1/Th2 balance, intestinal oxidative stress and inflammation, which were considered as food allergy-associated factors. Medicine-food homologous materials (MFHM) were considered as a significant factor with respect to preventing human diseases. To evaluate the associations between MFHM and food allergy-associated factors, two types of MFHM with the remarkable function of anti-oxidation and anti-inflammation, Gardeniae fructus (Gar) and Sophorae glos (Sop), were chosen. RESULTS: By constructing an H2O2-induced oxidative stress model of Caco-2 cells and an intestinal inflammatory cell model of Caco-2 cells with tumor necrosis factor-α and interleukin (IL)-13, the contents of anti-oxidative enzymes (SOD and GSH), inflammatory factor (IL-8) and tight junction proteins (zonula occludens-1, occludin and claudin-1) in Caco-2 cells were determined. Moreover, the anti-allergic effects of digestive Sop and Gar were evaluated by measuring the levels of Th1/Th2/Treg cytokines in the spleen cells of sensitized mice. The results showed that the SOD and GSH were obviously increased and the gene and protein expression of IL-8 and claudin-1 were improved with the incubation of digested Sop. Th2 cytokine was reduced and Th1/Th2 balance was promoted on coincubation with ovalbumin (OVA) and digested Sop in the splenocytes. However, the digested Gar had no effect. CONCLUSION: The digested Sop not only had suppressive effects on intestinal oxidative stress and inflammation, but also had regulative effects on Th1/Th2 balance. This finding demonstrated that not all of the MFHM with anti-oxidant and anti-inflammatory effects have anti-allergic activities. The present study may be contributing toward establishing a screening model to identify the anti-allergic MFHM. © 2024 Society of Chemical Industry.

Changes in structural and functional properties of whey protein cross-linked by polyphenol oxidase.

The natural whey protein is unstable, to achieve more efficient utilization, the functional properties of whey protein were modified by changing its structure, and enzymatic cross-linking is one of the common methods in dairy products to change the functional characterization. This study was conducted with objective to evaluate the structural and functional of whey protein which was cross-linked by polyphenol oxidase from Agaricus bisporus. Whey protein was cross-linked by polyphenol oxidase, and the polymers and dimers were revealed by SDS-PAGE and LC-MS/MS, the structural alterations of the polymers were analyzed by UV-vis, fluorescence spectroscopy and SEM, and the effects of functional properties of whey protein after cross-linked were also explored. Results showed that dimer and high polymer of ß-lactoglobulin were formed, the secondary structure of whey protein was exhibited a significant variation, and the microstructure changed obviously. Moreover, the foaming and antioxidant activity of whey protein was enhanced although the emulsifying was reduced after cross-linked. These findings emphasize the feasible application of enzymatic cross-linking in improving the functional properties of whey protein, and provide a new direction for changing the traditional processing technology of whey protein and developing high-quality products.

Allergenicity evaluation of peptides from milk and yogurt after gastrointestinal digestion based on epitopes.

As we know, milk and yogurt have good nutritional value and it is reported that some peptides can induce tolerance to alleviate or eliminate cow's milk allergy (CMA). However, there is a lack of detailed information on the peptides after digestion which could induce tolerance. In this study, the distribution pattern of digested proteins was detected during gastrointestinal digestion in infants and adults by Tricine-SDS-PAGE and RP-HPLC, and the digestive products were transported by the model of Caco-2 cells and the sequence of peptides was identified by LC-MS/MS. Residual allergenicity was evaluated by indirect ELISA during gastrointestinal digestion and the released peptides were aligned with T cell/IgE epitopes and biological functions by prediction software and previous information. These results indicated that the major allergens in yogurt were more easily digested with stronger transport capacity and had lower IgE-binding capacity. We obtained 113 peptides from the major allergens in the digested products and 38 of these peptides existed in all the digested products, among which 13 peptides had specific biological functions, such as ACE-inhibitory, antimicrobial and DPP-IV inhibitory properties. Although there was no obvious difference in the number of epitopes between fresh milk and yogurt, the difference in the properties and content of specific peptides might be the key factor for the difference in allergenicity. Most importantly, 11 peptides that contained T cell epitopes but not IgE epitopes might induce immune tolerance in CMA, which should be confirmed further.

Identification of immunoglobulin E epitopes on major allergens from dairy products after digestion and transportation in vitro.

Dairy processing can alter the digestion stability and bioavailability of cow milk proteins in the gastrointestinal tract. However, analysis of stable linear epitopes on cow milk allergens that could enter into intestinal mucosal is limited. Thus, this study aimed to investigate the digestion and transportation properties and residual allergen epitopes entering into gastrointestinal mucosa of 3 commercial dairy products, including pasteurized milk (PM), ultra-heat-treated milk (UHTM), and dried skim milk (DSM). In this work, the digestive stability of the 3 kinds of dairy products has been performed in a standard multistep static digestion model in vitro and characterized by Tricine-SDS-polyacrylamide gel electrophoresis and reversed-phase HPLC. With respect to gastrointestinal digestion in vitro, the main allergens including ß-lactoglobulin (ß-LG), α-lactalbumin (α-LA), and caseins were degraded gradually, and the resistance peptides remained in the PM with a molecular weight of range from 3.4 to 5.0 kDa. Simultaneously, the potential allergenicity of the cow milk proteins was diminished gradually and is basically consistent after 60 min of gastrointestinal digestion. After gastrointestinal digestion, the remaining peptides were transported via an Ussing chamber and identified by liquid chromatography-MS/MS. By alignment, 10 epitopes peptides were identified from 16 stable peptides, including 5 peptides (AA 92-100, 125-135, 125-138, and 149-162) in ß-LG, 2 peptides in α-LA (AA 80-93 and 63-79), 2 peptides in αS1-casein (AA 84-90 and 125-132), and 1 peptide (AA 25-32) in αS2-casein were identified by dot-blotting mainly exist in UHTM and PM. This study demonstrates dairy processing can affect the digestion and transport characteristics of milk proteins and in turn alter epitope peptides release.

Antioxidant and Anti-Inflammatory Potential of Peptides Derived from In Vitro Gastrointestinal Digestion of Germinated and Heat-Treated Foxtail Millet (Setaria italica) Proteins.

This study aimed at identifying antioxidant and anti-inflammatory peptides derived from the in vitro gastrointestinal digestion of germinated and heated (microwave and boiling) foxtail millet. The protein digest fraction containing low-molecular-weight peptides (<3 kDa) and the most hydrophobic subfraction (F4) abundant in random coil structure were responsible for the bioactivity. Then, seven novel peptides were identified using liquid chromatography with tandem mass spectrometry (LC-MS/MS) from the most potent F4 subfraction derived from boiled germinated millet. All seven synthesized peptides significantly (p < 0.05) reduced reactive oxygen species production and increased glutathione content and superoxide dismutase activity in Caco-2 cells, whereas two peptides (EDDQMDPMAK and QNWDFCEAWEPCF) were superior in inhibiting nitric oxide, tumor necrosis factor-α (reduced to 42.29 and 44.07%, respectively), and interleukin-6 (reduced to 56.59 and 43.45%, respectively) production in a RAW 264.7 cell model. This study is the first to report about the potential role of germinated and heated foxtail millet as a source of dual antioxidant and anti-inflammatory peptides.

The dietary c9,t11-conjugated linoleic acid enriched from butter reduces breast cancer progression in vivo.

The c9,t11-conjugated linoleic acid (CLA), which is the minor polyunsaturated fatty acid (PUFA) naturally present in butter, has gained attention due to its important preventive effect against breast cancer in vitro. In this paper, the enrichment of c9,t11-CLA from butter was optimized and the preventive effect of dietary c9,t11-CLA against breast cancer in vivo was investigated. Results showed that the concentration of c9,t11-CLA increased more than 10 times via a one-step urea complexation. Furthermore, the dietary c9,t11-CLA showed obvious preventive effect against breast cancer in decreasing the tumor weight and volume, and reducing the tumor incidence up to 50%. In addition, the expression of progesterone receptor and Ki-67 decreased significantly with the treatment of c9,t11-CLA. In conclusion, the dietary c9,t11-CLA enriched from butter showed a preventive effect against breast cancer in vivo via the inhibition of the hormonal receptor and cell proliferation. PRACTICAL APPLICATIONS: This paper provided new insight into the preparation of specific c9,t11-CLA isomer. It can be enriched from butter in large-scale with low-cost by urea complexation. Meanwhile, the enriched dietary c9,t11-CLA can be further processed into cancer prevention functional foods.

The Profile of Human Milk Metabolome, Cytokines, and Antibodies in Inflammatory Bowel Diseases Versus Healthy Mothers, and Potential Impact on the Newborn.

BACKGROUND AND AIMS: For women with inflammatory bowel disease [IBD], it is not very well known how IBD or IBD treatment affects their breast milk components. We aimed to investigate whether breast milk composition differs in healthy control [HC] versus IBD mothers in terms of antibodies, cytokines, and metabolite,s to identify potential impact of IBD breast milk on neonatal immune system. METHODS: Breast milk specimens from HC [n = 17] and IBD [n = 31 for Crohn's disease [CD]; and n = 41 for ulcerative colitis [UC]; were collected at 3 and 6 months postpartum [PP3] and [PP6], respectively. Faecal samples were also collected. Cytokines and immunoglobulins [IgA/IgG/IgE] were analysed by multiplex Meso Scale Discovery [MSD] and commercial kits. Moreover, breast milk metabolites were analysed by 1H nuclear magnetic resonance [NMR]. RESULTS: We found that breast milk from IBD mothers showed significantly lower levels of IgA, sugar metabolite [lactose], and 2-aminobutyrate. In contrast, we observed that breast milk from mothers with IBD had increased levels of pro-inflammatory cytokines and higher energy metabolites [lactate and succinate] than milk from healthy mothers. In addition, we noticed that the type of treatment [5-aminosalicylic acid versus biologics] influenced the milk cytokines and metabolites profile. CONCLUSIONS: The reduction in immunoprotective components of IBD breast milk such as sIgA and lactose theoretically may modulate the potential protective effects of breastfeeding. On the other hand, presence of higher levels of pro-inflammatory cytokines, lactate, and succinate may predispose the offspring to an inflammatory condition or impact on the gut microbiome. Better understanding of the role of succinate in infants and its potential effects on microbiome or mucosal immunity merits further investigations.

Lower Abundance and Impaired Function of CD71+ Erythroid Cells in Inflammatory Bowel Disease Patients During Pregnancy.

BACKGROUND AND AIMS: CD71+ erythroid cells are enriched during pregnancy with immuno suppressive properties. We investigated the frequency and functionality of CD71+ erythroid cells in peripheral blood, cord blood, and placenta of inflammatory bowel disease [IBD] patients versus healthy controls [HCs]. We aimed to determine their role in IBD pathogenesis during pregnancy. METHODS: Peripheral blood was collected at preconception, the first, second and third trimesters, and postpartum. Cord blood and placental tissues were collected at the time of birth. Cells from different specimens were subjected to immune-phenotyping and functional assays. CD71+ erythroid cells were purified for quantitative polymerase chain reaction [qPCR] analysis. Using an allogeneic mouse model of pregnancy, the effects of CD71+ erythroid cells depletion on intestinal homeostasis and dysbiosis was studied. RESULTS: IBD patients had lower CD71+ erythroid cells during pregnancy compared with HCs. Placenta and cord blood CD71+ erythroid cells from IBD patients exhibited impaired functionality and expressed lower inhibitory molecules including VISTA, TGF-ß, and reactive oxygen species [ROS]. Lower CD71+ erythroid cells were correlated with reduced regulatory T cells and increased immune-activation in IBD patients. Depletion of CD71+ erythroid cells in an allogeneic pregnancy model resulted in upregulation of TLRs, IL-6, and CXCL-1, and enhanced production of TNF-α, in intestinal tissues. In contrast, TGF-ß gene expression was reduced. Excessive inflammatory response in the gut [e.g. TNF-α] affects intestinal integrity and CD71+ erythroid cells impact on the gut's bacterial composition. CONCLUSIONS: Reduced frequency and/or impaired functionality of CD71+ erythroid cells during pregnancy may predispose IBD patients to a more pro-inflammatory milieu in their gastrointestinal tract, characterised by lower Tregs, higher IL-6, and TNF-α, and dysbiosis.

Potential allergenicity response to structural modification of irradiated bovine α-lactalbumin.

Bovine α-lactalbumin (α-La) is a major food allergen found in milk and is characterized by high conformational stability because of its four disulfide bridges and being calcium bound. This study aimed to describe the influence of gamma irradiation on the structure and potential allergenicity of α-La. The prepared α-La was irradiated at doses of 1-10 kGy. The changes in structure were characterized through SDS-PAGE, circular dichroism spectroscopy, ultraviolet absorption spectroscopy, and fluorescence spectroscopy. The potential allergenicity of the irradiated α-La was evaluated in vitro through IgG/IgE inhibition ELISA and the human basophil KU812 degranulation assay. The results showed that the secondary and tertiary structures of α-La significantly changed and caused extensive protein denaturation and aggregation. IgG and IgE binding properties remarkably decreased, and the degranulation capacity of basophils weakened. The results suggested that structural damage of α-La induced by irradiation significantly reduces the potential allergenicity of α-La.