Differential Effects of Oligosaccharides, Antioxidants, Amino Acids and PUFAs on Heat/Hypoxia-Induced Epithelial Injury in a Caco-2/HT-29 Co-Culture Model.

(1) Exposure of intestinal epithelial cells to heat and hypoxia causes a (heat) stress response, resulting in the breakdown of epithelial integrity. There are indications that several categories of nutritional components have beneficial effects on maintaining the intestinal epithelial integrity under stress conditions. This study evaluated the effect of nine nutritional components, including non-digestible oligosaccharides (galacto-oligosaccharides (GOS), fructo-oligosaccharides (FOS), chitosan oligosaccharides (COS)), antioxidants (α-lipoic acid (ALA), resveratrol (RES)), amino acids (l-glutamine (Glu), l-arginine (Arg)) and polyunsaturated fatty acids (PUFAs) (docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA)), on heat/hypoxia-induced epithelial injury. (2) Two human colonic cell lines, Caco-2 and HT-29, were co-cultured and pre-treated with the nutritional components for 48 h. After pre-treatment, the cells were exposed to heat/hypoxia (42 °C, 5% O2) for 2 h. Epithelial integrity was evaluated by measuring trans-epithelial electrical resistance (TEER), paracellular Lucifer Yellow (LY) permeability, and tight junction (TJ) protein expression. Heat stress and oxidative stress levels were evaluated by determining heat-shock protein-70 (HSP-70) expression and the concentration of the lipid peroxidation product malondialdehyde (MDA). (3) GOS, FOS, COS, ALA, RES, Arg, and EPA presented protective effects on epithelial damage in heat/hypoxia-exposed Caco-2/HT-29 cells by preventing the decrease in TEER, the increase in LY permeability, and/or decrease in TJ proteins zonula occludens-1 (ZO-1) and claudin-3 expression. COS, RES, and EPA demonstrated anti-oxidative stress effects by suppressing the heat/hypoxia-induced MDA production, while Arg further elevated the heat/hypoxia-induced increase in HSP-70 expression. (4) This study indicates that various nutritional components have the potential to counteract heat/hypoxia-induced intestinal injury and might be interesting candidates for future in vivo studies and clinical trials in gastrointestinal disorders related to heat stress and hypoxia.

Remote sensing and signaling in kidney proximal tubules stimulates gut microbiome-derived organic anion secretion.

Membrane transporters and receptors are responsible for balancing nutrient and metabolite levels to aid body homeostasis. Here, we report that proximal tubule cells in kidneys sense elevated endogenous, gut microbiome-derived, metabolite levels through EGF receptors and downstream signaling to induce their secretion by up-regulating the organic anion transporter-1 (OAT1). Remote metabolite sensing and signaling was observed in kidneys from healthy volunteers and rats in vivo, leading to induced OAT1 expression and increased removal of indoxyl sulfate, a prototypical microbiome-derived metabolite and uremic toxin. Using 2D and 3D human proximal tubule cell models, we show that indoxyl sulfate induces OAT1 via AhR and EGFR signaling, controlled by miR-223. Concomitantly produced reactive oxygen species (ROS) control OAT1 activity and are balanced by the glutathione pathway, as confirmed by cellular metabolomic profiling. Collectively, we demonstrate remote metabolite sensing and signaling as an effective OAT1 regulation mechanism to maintain plasma metabolite levels by controlling their secretion.

Curdlan, zymosan and a yeast-derived ß-glucan reshape tumor-associated macrophages into producers of inflammatory chemo-attractants.

Anti-cancer T-cell responses are often halted due to the immune-suppressive micro-environment, in part related to tumor-associated macrophages. In the current study, we assessed indigestible ß-glucans (oatßG, curdlan, grifolan, schizophyllan, lentinan, yeast whole glucan particles (yWGP), zymosan and two additional yeast-derived ß-glucans a and b) for their physicochemical properties as well as their effects on the plasticity of human monocyte-derived macrophages that were polarized with IL-4 to immune-suppressive macrophages. Beta-glucans were LPS/LTA free, and tested for solubility, molecular masses, protein and monosaccharide contents. Curdlan, yeast-b and zymosan re-polarized M(IL-4) macrophages towards an M1-like phenotype, in particular showing enhanced gene expression of CCR7, ICAM1 and CD80, and secretion of TNF-α and IL-6. Notably, differential gene expression, pathway analysis as well as protein expressions demonstrated that M(IL-4) macrophages treated with curdlan, yeast-b or zymosan demonstrated enhanced production of chemo-attractants, such as CCL3, CCL4, and CXCL8, which contribute to recruitment of monocytes and neutrophils. The secretion of chemo-attractants was confirmed when using patient-derived melanoma-infiltrating immune cells. Taken together, the bacterial-derived curdlan as well as the yeast-derived ß-glucans yeast-b and zymosan have the unique ability to preferentially skew macrophages towards a chemo-attractant-producing phenotype that may aid in anti-cancer immune responses.

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.

Phenylthiourea Binding to Human Tyrosinase-Related Protein 1.

Tyrosinase-related protein 1 (TYRP1) is one of the three human melanogenic enzymes involved in the biosynthesis of melanin, a pigment responsible for the color of the skin, hair, and eyes. It shares high sequence identity with tyrosinase, but has two zinc ions in its active site rather than two copper ions as in tyrosinase. Typical tyrosinase inhibitors do not directly coordinate to the zinc ions of TYRP1. Here, we show, from an X-ray crystal structure determination, that phenylthiourea, a highly potent tyrosinase inhibitor, does neither coordinate the active site zinc ions, but binds differently from other structurally characterized TYRP1-inhibitor complexes. Its aromatic ring is directed outwards from the active site, apparently as a result from the absence of polar oxygen substituents that can take the position of water molecules bound in the active site. The compound binds via hydrophobic interactions, thereby blocking substrate access to the active site.

Binding of CML-Modified as Well as Heat-Glycated ß-lactoglobulin to Receptors for AGEs Is Determined by Charge and Hydrophobicity.

Intake of dietary advanced glycation end products (AGEs) is associated with inflammation-related health problems. Nε-carboxymethyl lysine (CML) is one of the best characterised AGEs in processed food. AGEs have been described as ligands for receptors present on antigen presenting cells. However, changes in protein secondary and tertiary structure also induce binding to AGE receptors. We aimed to discriminate the role of different protein modifications in binding to AGE receptors. Therefore, ß-lactoglobulin was chemically modified with glyoxylic acid to produce CML and compared to ß-lactoglobulin glycated with lactose. Secondary structure was monitored with circular dichroism, while hydrophobicity and formation of ß-sheet structures was measured with ANS-assay and ThT-assay, respectively. Aggregation was monitored using native-PAGE. Binding to sRAGE, CD36, and galectin-3 was measured using inhibition ELISA. Even though no changes in secondary structure were observed in all tested samples, binding to AGE receptors increased with CML concentration of CML-modified ß-lactoglobulin. The negative charge of CML was a crucial determinant for the binding of protein bound CML, while binding of glycated BLG was determined by increasing hydrophobicity. This shows that sRAGE, galectin-3, and CD36 bind to protein bound CML and points out the role of negatively charged AGEs in binding to AGE receptors.

IgE Cross-Reactivity of Cashew Nut Allergens.

BACKGROUND: Allergic sensitisation towards cashew nut often happens without a clear history of eating cashew nut. IgE cross-reactivity between cashew and pistachio nut is well described; however, the ability of cashew nut-specific IgE to cross-react to common tree nut species and other Anacardiaceae, like mango, pink peppercorn, or sumac is largely unknown. OBJECTIVES: Cashew nut allergic individuals may cross-react to foods that are phylogenetically related to cashew. We aimed to determine IgE cross-sensitisation and cross-reactivity profiles in cashew nut-sensitised subjects, towards botanically related proteins of other Anacardiaceae family members and related tree nut species. METHOD: Sera from children with a suspected cashew nut allergy (n = 56) were assessed for IgE sensitisation to common tree nuts, mango, pink peppercorn, and sumac using dot blot technique. Allergen cross-reactivity patterns between Anacardiaceae species were subsequently examined by SDS-PAGE and immunoblot inhibition, and IgE-reactive allergens were identified by LC-MS/MS. RESULTS: From the 56 subjects analysed, 36 were positive on dot blot for cashew nut (63%). Of these, 50% were mono-sensitised to cashew nuts, 19% were co-sensitised to Anacardiaceae species, and 31% were co-sensitised to tree nuts. Subjects co-sensitised to Anacardiaceae species displayed a different allergen recognition pattern than subjects sensitised to common tree nuts. In pink peppercorn, putative albumin- and legumin-type seed storage proteins were found to cross-react with serum of cashew nut-sensitised subjects in vitro. In addition, a putative luminal binding protein was identified, which, among others, may be involved in cross-reactivity between several Anacardiaceae species. CONCLUSIONS: Results demonstrate the in vitro presence of IgE cross-sensitisation in children towards multiple Anacardiaceae species. In this study, putative novel allergens were identified in cashew, pistachio, and pink peppercorn, which may pose factors that underlie the observed cross-sensitivity to these species. The clinical relevance of this widespread cross-sensitisation is unknown.

Structure and Function of Human Tyrosinase and Tyrosinase-Related Proteins.

Melanin is the main pigment responsible for the color of human skin, hair and eye. Its biosynthesis requires three melanogenic enzymes, tyrosinase (TYR), and the tyrosinase-related proteins TYRP1 and TYRP2. The difficulty of isolating pure and homogeneous proteins from endogenous sources has hampered their study, and resulted in many contradictory findings regarding their physiological functions. In this review, we summarize recent advances on the structure and function of TYR and TYRPs by virtue of the crystal structure of human TYRP1, which is the first available structure of a mammalian melanogenic enzyme. This structure, combined with tyrosinase structures from other lower eukaryotes and mutagenesis studies of key active site residues, sheds light on the mechanism of TYR and TYRPs. Furthermore, a TYRP1-based homology model of TYR provides a high-quality platform to map and analyze albinism-related mutations, as well as the design of specific antimelanogenic compounds. Finally, we provide perspectives for future structure/function studies of TYR and TYRPs.

Small percentage of anaphylactic reactions treated with epinephrine during food challenges in Dutch children.

BACKGROUND: Severe allergic reactions, including anaphylaxis, occur during oral food challenges (OFCs) and the first-line treatment of anaphylaxis is epinephrine. OBJECTIVE: To evaluate the percentage of anaphylactic reactions treated with epinephrine during OFCs and to identify associated factors for the administration of epinephrine. METHODS: Children who underwent an OFC with peanut, hazelnut, cow's milk, hen's egg, or cashew nut from 2005 through 2015 in the Netherlands were evaluated. Children with reactions meeting the criteria for anaphylaxis according to the European Academy of Allergy and Clinical Immunology guidelines for food allergy and anaphylaxis were included. Children with an anaphylactic reaction treated with vs without epinephrine were compared. Possible factors associated with the administration of epinephrine, such as age, sex, symptoms consistent with asthma, history of an allergic reaction to the tested allergen, and symptom types during the anaphylactic reaction, were evaluated using logistic regression analysis. RESULTS: Eighty-three children in clinical and research settings (43% boys; median age, 7 years; range, 1-17) who met the criteria for anaphylaxis were included in this study. Thirty-two of 83 children (39%) with anaphylaxis were treated with epinephrine. Respiratory symptoms during the OFC were treated significantly more often with epinephrine than gastrointestinal symptoms (P = .01). CONCLUSION: Only 39% of children with anaphylaxis, according to the guideline criteria, were treated with epinephrine during the OFC and most of these children had respiratory symptoms. There is need for an easy-to-use international guideline for the treatment of allergic symptoms during OFCs.

The Effect of Tomatine on Gene Expression and Cell Monolayer Integrity in Caco-2.

More understanding of the risk-benefit effect of the glycoalkaloid tomatine is required to be able to estimate the role it might play in our diet. In this work, we focused on effects towards intestinal epithelial cells based on a Caco-2 model in order to analyze the influence on the cell monolayer integrity and on the expression levels of genes involved in cholesterol/sterol biosynthesis (LDLR), lipid metabolism (NR2F2), glucose and amino acid uptake (SGLT1, PAT1), cell cycle (PCNA, CDKN1A), apoptosis (CASP-3, BMF, KLF6), tight junctions (CLDN4, OCLN2) and cytokine-mediated signaling (IL-8, IL1ß, TSLP, TNF-α). Furthermore, since the bioactivity of the compound might vary in the presence of a food matrix and following digestion, the influence of both pure tomatine and in vitro digested tomatine with and without tomato fruit matrix was studied. The obtained results suggested that concentrations <20 µg/mL of tomatine, either undigested or in vitro digested, do not compromise the viability of Caco-2 cells and stimulate cytokine expression. This effect of tomatine, in vitro digested tomatine or in vitro digested tomatine with tomato matrix differs slightly, probably due to variations of bioactivity or bioavailability of the tomatine. The results lead to the hypothesis that tomatine acts as hormetic compound that can induce beneficial or risk toxic effects whether used in low or high dose.

Adaptation of exercise-induced stress in well-trained healthy young men.

NEW FINDINGS: What is the central question of this study? Exercise is known to induce stress-related physiological responses, such as changes in intestinal barrier function. Our aim was to determine the test-retest repeatability of these responses in well-trained individuals. What is the main finding and its importance? Responses to strenuous exercise, as indicated by stress-related markers such as intestinal integrity markers and myokines, showed high test-retest variation. Even in well-trained young men an adapted response is seen after a single repetition after 1 week. This finding has implications for the design of studies aimed at evaluating physiological responses to exercise. Strenuous exercise induces different stress-related physiological changes, potentially including changes in intestinal barrier function. In the Protégé Study (ISRCTN14236739; www.isrctn.com), we determined the test-retest repeatability in responses to exercise in well-trained individuals. Eleven well-trained men (27 ± 4 years old) completed an exercise protocol that consisted of intensive cycling intervals, followed by an overnight fast and an additional 90 min cycling phase at 50% of maximal workload the next morning. The day before (rest), and immediately after the exercise protocol (exercise) a lactulose and rhamnose solution was ingested. Markers of energy metabolism, lactulose-to-rhamnose ratio, several cytokines and potential stress-related markers were measured at rest and during exercise. In addition, untargeted urine metabolite profiles were obtained. The complete procedure (Test) was repeated 1 week later (Retest) to assess repeatability. Metabolic effect parameters with regard to energy metabolism and urine metabolomics were similar for both the Test and Retest period, underlining comparable exercise load. Following exercise, intestinal permeability (1 h plasma lactulose-to-rhamnose ratio) and the serum interleukin-6, interleukin-10, fibroblast growth factor-21 and muscle creatine kinase concentrations were significantly increased compared with rest only during the first test and not when the test was repeated. Responses to strenuous exercise in well-trained young men, as indicated by intestinal markers and myokines, show adaptation in Test-Retest outcome. This might be attributable to a carry-over effect of the defense mechanisms triggered during the Test. This finding has implications for the design of studies aimed at evaluating physiological responses to exercise.

Structure of Human Tyrosinase Related Protein†1 Reveals a Binuclear Zinc Active Site Important for Melanogenesis.

Tyrosinase-related protein 1 (TYRP1) is one of three tyrosinase-like glycoenzymes in human melanocytes that are key to the production of melanin, the compound responsible for the pigmentation of skin, eye, and hair. Difficulties with producing these enzymes in pure form have hampered the understanding of their activity and the effect of mutations that cause albinism and pigmentation disorders. Herein we show that the typical tyrosinase-like subdomain of TYRP1 contains two zinc ions in the active site instead of copper ions as found in tyrosinases, which explains why TYRP1 does not exhibit tyrosinase redox activity. In addition, the structures reveal for the first time that the Cys-rich subdomain, which is unique to vertebrate melanogenic proteins, has an epidermal growth factor-like fold and is tightly associated with the tyrosinase subdomain. Our structures suggest that most albinism-related mutations of TYRP1 affect its stability or activity.

Favourable effects of consuming a Palaeolithic-type diet on characteristics of the metabolic syndrome: a randomized controlled pilot-study.

BACKGROUND: The main goal of this randomized controlled single-blinded pilot study was to study whether, independent of weight loss, a Palaeolithic-type diet alters characteristics of the metabolic syndrome. Next we searched for outcome variables that might become favourably influenced by a Paleolithic-type diet and may provide new insights in the pathophysiological mechanisms underlying the metabolic syndrome. In addition, more information on feasibility and designing an innovative dietary research program on the basis of a Palaeolithic-type diet was obtained. METHODS: Thirty-four subjects, with at least two characteristics of the metabolic syndrome, were randomized to a two weeks Palaeolithic-type diet (n = 18) or an isoenergetic healthy reference diet, based on the guidelines of the Dutch Health Council (n = 14). Thirty-two subjects completed the study. Measures were taken to keep bodyweight stable. As primary outcomes oral glucose tolerance and characteristics of the metabolic syndrome (abdominal circumference, blood pressure, glucose, lipids) were measured. Secondary outcomes were intestinal permeability, inflammation and salivary cortisol. Data were collected at baseline and after the intervention. RESULTS: Subjects were 53.5 (SD9.7) year old men (n = 9) and women (n = 25) with mean BMI of 31.8 (SD5.7) kg/m2. The Palaeolithic-type diet resulted in lower systolic blood pressure (-9.1 mmHg; P = 0.015), diastolic blood pressure (-5.2 mmHg; P = 0.038), total cholesterol (-0.52 mmol/l; P = 0.037), triglycerides (-0.89 mmol/l; P = 0.001) and higher HDL-cholesterol (+0.15 mmol/l; P = 0.013), compared to reference. The number of characteristics of the metabolic syndrome decreased with 1.07 (P = 0.010) upon the Palaeolithic-type diet, compared to reference. Despite efforts to keep bodyweight stable, it decreased in the Palaeolithic group compared to reference (-1.32 kg; P = 0.012). However, favourable effects remained after post-hoc adjustments for this unintended weight loss. No changes were observed for intestinal permeability, inflammation and salivary cortisol. CONCLUSIONS: We conclude that consuming a Palaeolithic-type diet for two weeks improved several cardiovascular risk factors compared to a healthy reference diet in subjects with the metabolic syndrome. TRIAL REGISTRATION: Nederlands Trial Register NTR3002.

Differential effects of heating modes on the immunogenic potential of soy-derived peptides released after in vitro infant digestion.

During production of soy-based infant formula, soy protein undergoes heating processes. This study investigated the differential impact of heating modes on the immunogenic potential of peptides in soy protein digests. Wet or dry heating was applied, followed by in vitro gastrointestinal infant digestion. The released peptides were analyzed by LC-MS/MS. Bioinformatics tools were utilized to predict and identify potential linear B-cell and T-cell epitopes, as well as to explore cross-reactivity with other legumes. Subsequently, the peptide intensities of the same potential epitope across different experimental conditions were compared. As a result, we confirmed the previously observed enhancing effect of wet heating on infant digestion and inhibitory effect of dry heating. A total of 8,546 peptides were detected in the digests, and 6,684 peptides were with a score over 80. Among them, 29 potential T-cell epitopes and 27 potential B-cell epitopes were predicted. Cross-reactivity between soy and other legumes, including peanut, pea, chickpea, lentil, kidney bean, and lupine, was also detected. Overall, heating and digestion time could modulate the potential to trigger peptide-induced immune responses.

Glycation of goat milk with different casein-to-whey protein ratios and its effects on simulated infant digestion.

This research compared the effects of dry heating on the digestion of goat milk proteins with different casein-to-whey ratios (40% casein, C40 and 80% casein, C80). The glycation markers of heated samples were determined by LC-MS. Heating at 60 °C for 8 h induced early glycation while heating at 60 °C for 72 h induced advanced glycation. Unheated C80 samples showed a higher digestibility than unheated C40 samples, which may be due to their higher protein solubility. After dry heating for 72 h, no significant difference in digestibility was observed between C80 and C40 samples. Heating for 72 h decreased the digestibility of C40 samples compared to unheated samples, probably due to glycation, while protein aggregation was the main reason for the reduced digestibility of heated C80 samples. Overall, this study showed that dry heating for 72 h induced a lower digestibility of C80 and C40 samples, although with different underlying mechanisms.

sRAGE-binding and antimicrobial bioactivities of soy and pea protein after heating and in vitro infant digestion.

During infant formula production, proteins are always heated, potentially affecting their digestibility and the bioactivities of resulting peptides. Although plant proteins are a promising dairy alternative for infant formula, they remain understudied, necessitating further investigations. Therefore, this research aimed to fill this gap by assessing the impact of different heating modes on soy protein (SP) and pea protein (PP), focusing on glycation levels, peptide formation during in vitro infant digestion, and immune protection potential (sRAGE-binding and antimicrobial activities) of the resulting peptides. Consequently, dry heating led to increased glycation and glycated peptide production, particularly with higher glycation in PP than SP. Moreover, PP exhibited an overall stronger sRAGE-binding capacity than SP, regardless of heating and digestion conditions. Regarding antimicrobial activity, both SP and PP-derived peptides displayed reduced effectiveness against Enterobacter cloacae after dry heating. Additionally, Staphylococcus epidermidis was differently inhibited, where PP-derived peptides showed inherent inhibition. The primary determinant of sRAGE-binding and antimicrobial potential in digestion-derived peptides was the protein source. Subsequent bioinformatics analysis predicted 519 and 133 potential antimicrobial peptides in SP and PP, respectively. This study emphasises the importance of protein source for infant formula to ensure infant health.

Heating affects protein digestion of skimmed goat milk under simulated infant conditions.

The objective of this research was to analyse the effects of heating on digestion of skimmed goat milk proteins. Most previous goat milk digestion studies evaluated the digestion only based on the supernatant. In this study, digestion of skimmed goat milk was studied in both supernatant and gastric clot. The results indicated that, compared to mild temperature heated samples (≤75 °C), samples heated at ≥80 °C showed more extensive gastric clot formation with a higher protein digestion rate, but also resulted in a larger amount of undigested whey proteins due to its severe aggregation. For the peptidome, ß-casein was the major source of bioactive peptides. The samples heated at 65 °C showed higher bioactive peptide abundances, whereas at temperatures higher than 75 °C, it was reduced due to cleavage into smaller peptides. Overall, this study showed that different heating temperatures induced different whey protein denaturation degrees, which affected their digestion in skimmed goat milk..

Evaluating and comparing tolerance, nutritional quality and bio-functional activity of bovine-plasma, corn and whey proteins, outcomes of a randomized double blind controlled trial.

Important considerations in the choice of future sustainable protein sources for human application are tolerance, nutritional quality, and potential health benefits. We evaluated, in a double-blind cross-over intervention trial, tolerance, nutritional quality, and potential health effects of two sustainable protein sources. Thirty-six apparently healthy older adults (age 62.3 ± 7.2yrs, BMI 25 ± 3 kg/m2) received 40 g/day bovine-plasma protein (BP), corn protein (CP) or, as a benchmark, whey protein (WP) for one week with a washout period of one week in-between. In 12 participants, we also determined postprandial amino acid (PAA) uptake kinetics upon consumption of 20 g BP, CP, or WP. Changes in self-reported gastrointestinal complaints and intestinal permeability assessed using a multi-sugar acetylsalicylic acid test did not differ between the interventions. Clear differences in PAA responses were observed after consumption of the different proteins, but clear essential amino acid responses were observed for all proteins. BP consumption resulted in a small but significant increase in blood pressure outcomes, and CP consumption resulted in a small but significant decrease in insulin levels when compared to the other interventions. In conclusion, alternative protein concentrates and isolates studied here can be consumed in relative high quantities without experiencing unwanted GI complaints or gut barrier dysfunction and they can be a good source of essential amino acids. The rise in blood pressure observed during the BP intervention, potentially linked to the elevated salt content of the BP, constitutes a potential health issue. Future studies with longer intervention periods might however be recommended.

Tolerance Induction in Cow's Milk Allergic Children by Heated Cow's Milk Protein: The iAGE Follow-Up Study.

Accelerating the induction of tolerance to cow's milk (CM) reduces the burden of cow's milk allergy (CMA). In this randomised controlled intervention study, we aimed to investigate the tolerance induction of a novel heated cow milk protein, the iAGE product, in 18 children with CMA (diagnosed by a paedriatric allergist). Children who tolerated the iAGE product were included. The treatment group (TG: n = 11; mean age 12.8 months, SD = 4.7) consumed the iAGE product daily with their own diet, and the control group (CG: n = 7; mean age 17.6 months, SD = 3.2) used an eHF without any milk consumption. In each group, 2 children had multiple food allergies. The follow-up procedures consisted of a double-blind placebo-controlled food challenge (DBPCFC) with CM t = 0, t = 1 (8 months), t = 2 (16 months), and t = 3 (24 months). At t = 1, eight (73%) of 11 children in the TG had a negative DBPCFC, versus four out of seven (57%) in the CG (BayesFactor = 0.61). At t = 3, nine of the 11 (82%) children in the TG and five of seven (71%) in the CG were tolerant (BayesFactor = 0.51). SIgE for CM reduced from a mean of 3.41 kU/L (SD = 5.63) in the TG to 1.24 kU/L (SD = 2.08) at the end of intervention, respectively a mean of 2.58 (SD = 3.32) in the CG to 0.63 kU/L (SD = 1.06). Product-related AEs were not reported. CM was successfully introduced in all children with negative DBPCFC. We found a standardised, well-defined heated CM protein powder that is safe for daily OIT treatment in a selected group of children with CMA. However, the benefits of inducing tolerance were not observed.