Infant Immune System Research Articles

The Human Milk-derived Peptide Drives Rapid Regulation of Macrophage Inflammation Responses in the Neonatal Intestine

Background and AimsThe interactions between human milk and the regulation of innate immune homeostasis in newborns, and their impact on intestinal health, are not fully understood. This study aimed to explore the role of peptides in human milk extracellular vesicles (EVs) in this process. MethodsA comprehensive screening of peptides within human milk EVs was performed, leading to the identification of a beta-casein-derived peptide (CASB135-150). The effects of CASB135-150 on intestinal injury were evaluated in a rat necrotizing enterocolitis (NEC) model. Immunofluorescence analysis was used to determine its distribution, and its impact on NF-κB signaling and inflammation was studied in bone marrow-derived macrophages (BMDMs) and intestinal macrophages. Protein-protein interaction (PPI) analysis, single-cell RNA-seq (scRNA-seq), and co-immunoprecipitation (co-IP) experiments were conducted to explore the mechanism underlying CASB135-150 function. ResultsCASB135-150 significantly mitigated intestinal injury in the rat NEC model. Immunofluorescence analysis revealed that CASB135-150 could target intestinal macrophages and rapidly inhibited NF-κB signaling and reduced inflammation. ScRNA-seq analyses indicated a strong association between FHL2 and NEC development, and co-IP confirmed the interaction between CASB135-150 and FHL2. CASB135-150 disrupted the FHL2/TRAF6 complex, reducing TRAF6 protein levels. Mutation of key amino acids in CASB135-150 disrupted its interaction with FHL2 and abolished its ability to inhibit NF-κB signaling, which also prevented its protective effect in vivo. RNA-seq of intestinal tissue further highlighted the impact of CASB135-150 on the NF-κB signaling pathway. ConclusionsOur study identifies CASB135-150, a novel peptide in human milk EVs, that rapidly regulates macrophage inflammatory responses and protects against NEC-induced intestinal injury. These findings provide new insights into the role of human milk in modulating the infant immune system and intestinal health.

Plasma adenosine deaminase-1 and -2 activities are lower at birth in Papua New Guinea than in The Gambia but converge over the first weeks of life.

Dynamic cellular and molecular adaptations in early life significantly impact health and disease. Upon birth, newborns are immediately challenged by their environment, placing urgent demands on the infant immune system. Adenosine deaminases (ADAs) are enzymatic immune modulators present in two isoforms - ADA-1 and ADA-2. Infants exhibit low ADA activity, resulting in high plasma adenosine concentrations and a consequent anti-inflammatory/anti-Th1 bias. While longitudinal studies of plasma ADA have been conducted in infants in The Gambia (GAM), little is known regarding ADA trajectories in other parts of the world. Herein, we characterized plasma ADA activity in an infant cohort in Papua New Guinea (PNG; n=83) and compared to ontogeny of ADA activity in a larger cohort in GAM (n=646). Heparinized peripheral blood samples were collected at day of life (DOL) 0, DOL7, DOL30, and DOL128. Plasma ADA-1, ADA-2, and total ADA activities were measured by chromogenic assay. Compared to GAM infants, PNG infants had significantly lower ADA-1 (0.9-fold), ADA-2 (0.42-fold), and total ADA (0.84-fold) activities at birth which converged by DOL30. Overall, discovery of a distinct baseline and a consistent pattern of increasing plasma ADA activity in early life in two genetically and geographically distinct populations validates and extends previous findings on the robustness of early life immune ontogeny.

Birth and household exposures are associated with changes to skin bacterial communities during infancy

Abstract Microbial exposures during infancy shape the development of the microbiome, the collection of microbes living in and on the body, which in turn directs immune system training. Newborns acquire a substantial quantity of microbes during birth and throughout infancy via exposure to microbes in the physical and social environment. Alterations to early life microbial environments may give rise to mismatches, where environmental, cultural, and behavioral changes that outpace the body’s adaptive responses can lead to adverse health outcomes, particularly those related to microbiome development and immune system regulation. This study explored the development of the skin microbiome among infants born in Chicago, USA. We collected skin swab microbiome samples from 22 mother-infant dyads during the first 48 hours of life and again at six weeks postpartum. Mothers provided information about social environments and hygiene behaviors that may impact infants’ microbial exposures. Analysis of 16S rRNA bacterial gene sequencing data revealed correlations between infant skin bacterial abundance shortly after birth and factors such as antibiotic exposure and receiving a bath in the hospital. The composition of the infant microbiome at six weeks of age was associated with interactions with caregivers and infant feeding practices. We also found shifts in maternal skin microbiomes that may reflect increased hygiene practices in the hospital. Our data suggest that factors related to the birth and household environment can impact the development of infant skin microbiomes and point to practices that may produce mismatches for the infant microbiome and immune system.

Recent Progress in Physiological Significance and Biosynthesis of Lacto-N-triose II: Insights into a Crucial Biomolecule.

Lacto-N-triose II (LNTri II), an important precursor for human milk oligosaccharide (HMOs) synthesis, has garnered significant attention due to its structural features and physiological properties. Composed of galactose (Gal), N-acetylglucosamine (GlcNAc), and glucose (Glc), with the chemical structure GlcNAcβ1,3Galβ1,4Glc, the distinctive structure of LNTri II confers various physiological functions such as promoting the growth of beneficial bacteria, regulating the infant immune system, and preventing certain gastrointestinal diseases. Extensive research efforts have been dedicated to elucidating efficient enzymatic synthesis pathways for LNTri II production, with particular emphasis on the transglycosylation activity of β-N-acetylhexosaminidases and the action of β-1,3-N-acetylglucosaminyltransferases. Additionally, metabolic engineering and cell factory approaches have been explored, harnessing the potential of engineered microbial hosts for the large-scale biosynthesis of LNTri II. This review summarizes the structure, derivatives, physiological effects, and biosynthesis of LNTri II.

Transcriptome and metabolome analyses reveal the effects of formula and breast milk on the growth and development of human small intestinal organoids

Breast milk is widely acknowledged as the ideal nutritional resource for infants and can well meet the nutritional requirements for baby’s growth and development. Infant formula is a substitute for breast milk, designed to closely mimic its composition and function for breast milk. Most of the previous studies used tumor colorectal cancer cell lines to study the nutritional potency of formula and its components, so realistic data closer to the baby could not be obtained. Small intestinal organoids, derived from differentiated human embryonic stem cells, can be used to simulate nutrient absorption and metabolism in vitro. In this experiment, we used small intestinal organoids to compare the nutrient absorption and metabolism of three infant formulae for 0–6 months with breast milk samples. Transcriptome and metabolome sequencing methods were used to analyze the differentially expressed genes (DEGs) and differentially expressed metabolites (DEMs). The pathways related to DEGs, DEMs were enriched using GO, KEGG, GSEA and other methods to investigate their biological characteristics. We have found that both formula and breast milk promote the development of the infant’s immune system, nutrient absorption and intestinal development. In PMH1 we found that the addition of oligofructose to milk powder promoted lipid metabolism and absorption. In PMH2 we found that whey protein powder favours the development of the immune system in infants. In PMH3 we found that oligogalactans may act on the brain-gut axis by regulating the intestinal flora, thereby promoting axon formation and neural development. By linking these biological properties of the milk powder with its composition, we confirmed the effects of added ingredients on the growth and development of infants. Also, we demonstrated the validity of small intestine organoids as a model for absorption and digestion in vitro. Through the above analyses, the advantages and disadvantages of the roles of formula and breast milk in the growth and metabolism of infants were also compared.

Effects of pregnancy and lactation prebiotics supplementation on infant allergic disease: A randomized controlled trial

BackgroundIngestion of prebiotics during pregnancy and lactation may have immunomodulatory benefits for the developing fetal and infant immune system and provide a potential dietary strategy to reduce the risk of allergic diseases. ObjectiveThe aim of this trial was to determine whether maternal supplementation with dietary prebiotics reduces the risk of allergic outcomes in infants with hereditary risk. MethodsWe undertook a double-blind, randomized controlled trial in which pregnant women were allocated to consume prebiotics (14.2g daily of galacto-oligosaccharides and fructo-oligosaccharides in ratio 9:1) or placebo (8.7g daily maltodextrin) powder from <21 weeks gestation until 6-months postnatal during lactation. Eligible women had infants with a first-degree relative with a history of medically diagnosed allergic disease. The primary outcome was infant medically diagnosed eczema by 1-year of age, and secondary outcomes included allergen sensitization, food allergy, and recurrent wheeze by 1-year of age. Results652 women were randomized between June 2016 and November 2021 (n=329 prebiotics, n=323 placebo). There was no significant difference between groups in the percentage of infants with medically diagnosed eczema by 1-year of age (prebiotics 31.5% (103/327 infants) compared to placebo 32.6% (105/322 infants); adjusted relative risk 0.98 (95% CI 0.77, 1.23; p=0.84). Secondary outcomes and safety measures also did not significantly differ between groups. ConclusionWe found little evidence that maternal prebiotics supplementation during pregnancy and lactation reduces the risk of infant medically diagnosed eczema by 1-year of age in infants who are at hereditary risk of allergic disease.

A newborn's perspective on immune responses to food.

In this review, we will highlight infants' immune responses to food, emphasizing the unique aspects of early-life immunity and the critical role of breast milk as a food dedicated to infants. Infants are susceptible to inflammatory responses rather than immune tolerance at the mucosal and skin barriers, necessitating strategies to promote oral tolerance that consider this susceptibility. Breast milk provides nutrients for growth and cell metabolism, including immune cells. The content of breast milk, influenced by maternal genetics and environmental exposures, prepares the infant's immune system for the outside world, including solid foods. To do this, breast milk promotes immune system development through antigen-specific and non-antigen-specific immune education by exposing the newborn to food and respiratory allergens and acting on three key targets for food allergy prevention: the gut microbiota, epithelial cells, and immune cells. Building knowledge of how the maternal exposome and human milk composition influence offspring's healthy immune development will lead to recommendations that meet the specific needs of the developing immune system and increase the chances of promoting an appropriate immune response to food in the long term.

The Role of Breastfeeding in Acute Respiratory Infections in Infancy.

Acute respiratory infections (ARIs) affect the respiratory tract, are often caused by viruses such as respiratory syncytial virus and rhinovirus, and present symptoms such as coughing, fever, respiratory distress, and breathing difficulty. The global adherence to exclusive breastfeeding (BF) for the first 6 months of life has reached 44%, supported by the World Health Organization and United Nations International Children's Emergency Fund efforts. BF provides vital nutrients and contributes to infant immune system development, protecting against infections. The role of BF in preventing and reducing complications of ARIs in infants is gaining attention, prompting a review of current data and future research needs. This review aims to summarize the evidence on the role of BF in reducing the risk and severity of ARIs in infants, elucidate the adaptations in breast milk composition during infections, and identify relevant research needs. Human milk (HM) is rich in immunoglobulins, antimicrobial peptides, and immunomodulatory factors that protect against various pathogens, including respiratory viruses. Several studies have demonstrated that BF is associated with a significant reduction in hospitalization, oxygen requirements, and mortality in infants with ARIs. The effectiveness of BF varies according to the specific respiratory virus, and a longer duration of exclusive BF appears to enhance its protective effect. It is documented that the composition of HM adjusts dynamically in response to infections, fortifying the infant's immune defenses. Specific immunological components of HM, including leukocytes and immunoglobulins, increase in response to infection in the infant, contributing to the enhancement of the immune defense in infants. Immune-boosting microRNAs enhance immune transfer to the infants and promote early gut maturation, and the HM microbiome along with other factors modifies the infant's gut microbiome and immune system. BF defends infants from respiratory infections, and the investigation of the microRNAs in HM offers new insights into its antiviral properties. The promotion of BF, especially in vulnerable communities, is of paramount importance in alleviating the global burden of ARIs in infancy.

Live Rotavirus Vaccination Appears Low-risk in Infants Born to Mothers With Inflammatory Bowel Disease on Biologics

Biologic therapies in the context of inflammatory bowel disease and pregnancy lead to improved maternal and fetal outcomes. Placental transfer results in detectable drug concentrations in infants. Rotavirus infection results in diarrheal related hospitalizations; however, the live oral vaccine is not currently recommended in biologic exposed infants. The aim of this study was to assess the effect of maternal biologic therapies on the infant immune system and safety of live rotavirus vaccination in biologic-exposed infants. Biologic-exposed infants underwent standardized clinical assessments, drug concentration, and immune function testing (complete blood count, differential, immunoglobulin levels, extended B-cell and T-cell subset enumeration, Recent Thymic Emigrants, regulatory T-cell numbers, mitogen stimulation assays, and review of T-cell Receptor Excision Circles in the newborn screen). Rotavirus vaccine-specific adverse effects following immunizations up to 42 days post the last dose of the vaccine series were recorded. There were 57 infants born to 52 mothers with inflammatory bowel disease exposed to infliximab (n= 21), adalimumab (n= 19), vedolizumab (n= 10), and ustekinumab (n= 7) in the third trimester for a median of 39 weeks (interquartile range, 38-39 weeks) at delivery. Immunologic assessments validated for age were normal in all infants despite median infliximab concentrations of 6.1 ug/mL (range, 0.4-28.8 ug/mL), adalimumab concentrations of 1.7ug/mL (range, 0.7-7.9 ug/mL), ustekinumab concentrations of 0.6 ug/mL (range, 0-1.1), and undetectable for vedolizumab at 10.7 weeks (interquartile range, 9.4-12.4) of age. The live oral rotavirus vaccine series was provided to 50 infants with the first dose given at a median of 13weeks of age. No adverse effects following immunization were reported. Immune function testing was normal, and administration of live rotavirus vaccination appeared low-risk in biologic-exposed infants irrespective of circulating drug levels.

Role of breastfeeding in disease prevention.

Human milk provides the infant with many bioactive factors, including immunomodulating components, antimicrobials and prebiotics, which modulate the infant microbiome and immune system maturation. As a result, breastfeeding can impact infant health from infancy, through adolescence, and into adulthood. From protecting the infant from infections, to reducing the risk of obesity, type 1 diabetes and childhood leukaemia, many positive health outcomes are observed in infants receiving breastmilk. For the mother, breastfeeding protects against postpartum bleeding and depression, increases weight loss, and long-term lowers the risk of type 2 diabetes, breast and ovarian cancer, and cardiovascular diseases. Beyond infants and mothers, the wider society is also impacted because of avoidable costs relating to morbidity and mortality derived from a lack of human milk exposure. In this review, Medline was used to search for relevant articles to discuss the health benefits of breastfeeding and its societal impact before exploring future recommendations to enhance our understanding of the mechanisms behind breastfeeding's positive effects and promote breastfeeding on a global scale.

Exploring the Impact of Extra Virgin Olive Oil on Maternal Immune System and Breast Milk Composition in Rats.

Maternal breast milk plays a key role in providing newborns with passive immunity and stimulating the maturation of an infant's immune system, protecting them from many diseases. It is known that diet can influence the immune system of lactating mothers and the composition of their breast milk. The aim of this study was to establish if a supplementation during the gestation and lactation of Lewis rats with extra virgin olive oil (EVOO), due to the high proportion of antioxidant components in its composition, has an impact on the mother's immune system and on the breast milk's immune composition. For this, 10 mL/kg of either EVOO, refined oil (control oil) or water (REF group) were orally administered once a day to rats during gestation and lactation periods. Immunoglobulin (Ig) concentrations and gene expressions of immune molecules were quantified in several compartments of the mothers. The EVOO group showed higher IgA levels in both the breast milk and the mammary glands than the REF group. In addition, the gene expression of IgA in mammary glands was also boosted by EVOO consumption. Overall, EVOO supplementation during gestation and lactation is safe and does not negatively affect the mother's immune system while improving breast milk immune composition by increasing the presence of IgA, which could be critical for an offspring's immune health.

Maternal immunoglobulin G affects brain development of mouse offspring

Maternal immunoglobulin (Ig)G is present in breast milk and has been shown to contribute to the development of the immune system in infants. In contrast, maternal IgG has no known effect on early childhood brain development. We found maternal IgG immunoreactivity in microglia, which are resident macrophages of the central nervous system of the pup brain, peaking at postnatal one week. Strong IgG immunoreactivity was observed in microglia in the corpus callosum and cerebellar white matter. IgG stimulation of primary cultured microglia activated the type I interferon feedback loop by Syk. Analysis of neonatal Fc receptor knockout (FcRn KO) mice that could not take up IgG from their mothers revealed abnormalities in the proliferation and/or survival of microglia, oligodendrocytes, and some types of interneurons. Moreover, FcRn KO mice also exhibited abnormalities in social behavior and lower locomotor activity in their home cages. Thus, changes in the mother-derived IgG levels affect brain development in offsprings.

Pediatric immunotherapy and HIV control.

Highlighting opportunities/potential for immunotherapy by understanding dynamics of HIV control during pediatric HIV infection with and without antiretroviral therapy (ART), as modeled in Simian immunodeficiency virus (SIV) and Simian-human immunodeficiency virus (SHIV)-infected rhesus macaques and observed in clinical trials. This review outlines mode of transmission, pathogenesis of pediatric HIV, unique aspects of the infant immune system, infant macaque models and immunotherapies. During the earliest stages of perinatal HIV infection, the infant immune system is characterized by a unique environment defined by immune tolerance and lack of HIV-specific T cell responses which contribute to disease progression. Moreover, primary lymphoid organs such as the thymus appear to play a distinct role in HIV pathogenesis in children living with HIV (CLWH). Key components of the immune system determine the degree of viral control, targets for strategies to induce viral control, and the response to immunotherapy. The pursuit of highly potent broadly neutralizing antibodies (bNAbs) and T cell vaccines has revolutionized the approach to HIV cure. Administration of HIV-1-specific bNAbs, targeting the highly variable envelope improves humoral immunity, and T cell vaccines induce or improve T cell responses such as the cytotoxic effects of HIV-1-specific CD8+ T cells, both of which are promising options towards virologic control and ART-free remission as evidenced by completed and ongoing clinical trials. Understanding early events during HIV infection and disease progression in CLWH serves as a foundation for predicting or targeting later outcomes by harnessing the immune system's natural responses. The developing pediatric immune system offers multiple opportunities for specific long-term immunotherapies capable of improving quality of life during adolescence and adulthood.

Maternal circadian rhythm disruption affects neonatal inflammation via metabolic reprograming of myeloid cells.

Disruption of circadian rhythm during pregnancy produces adverse health outcomes in offspring; however, the role of maternal circadian rhythms in the immune system of infants and their susceptibility to inflammation remains poorly understood. Here we show that disruption of circadian rhythms in pregnant mice profoundly aggravates the severity of neonatal inflammatory disorders in both male and female offspring, such as necrotizing enterocolitis and sepsis. The diminished maternal production of docosahexaenoic acid (DHA) and the impaired immunosuppressive function of neonatal myeloid-derived suppressor cells (MDSCs) contribute to this phenomenon. Mechanistically, DHA enhances the immunosuppressive function of MDSCs via PPARγ-mediated mitochondrial oxidative phosphorylation. Transfer of MDSCs or perinatal supplementation of DHA relieves neonatal inflammation induced by maternal rhythm disruption. These observations collectively demonstrate a previously unrecognized role of maternal circadian rhythms in the control of neonatal inflammation via metabolic reprograming of myeloid cells.

FcαRI (CD89) is upregulated on subsets of mucosal and circulating NK cells and regulates IgA-class specific signaling and functions

Immunoglobulin A (IgA) is the predominant mucosal antibody class with both anti- and pro-inflammatory roles [1–3]. However, the specific role of the IgA receptor CD89, expressed by a subset of natural killer (NK) cells, is poorly explored. We found that CD89 protein expression on circulating NK cells is infrequent in humans and rhesus macaques, but transcriptomic analysis showed ubiquitous CD89 expression, suggesting an inducible phenotype. Interestingly, CD89+ NK cells were more frequent in cord blood and mucosae, indicating a putative IgA-mediated NK cell function in the mucosae and infant immune system. CD89+ NK cells signaled through upregulated CD3ζ, SYK, ZAP70, and SLAMF1, but also showed high expression of inhibitory receptors such as KLRG1 and reduced activating NKp46 and NKp30. CD89-based activation or ADCC with monomeric IgA1 reduced NK cell functions, while ADCC with combinations of IgG and IgA2 was enhanced compared to IgG alone. These data suggest that functional CD89+ NK cells survey mucosal sites, but CD89 likely serves as regulatory receptor which can be further modulated depending on IgA and IgG subclass. Although the full functional niche of CD89+ NK cells remains unexplored, these intriguing data suggest the CD89 axis could represent a novel immunotherapeutic target in the mucosae or early life.

Human milk oligosaccharides differentially support gut barrier integrity and enhance Th1 and Th17 cell effector responses in vitro.

Human milk oligosaccharides (HMOs) can modulate the intestinal barrier and regulate immune cells to favor the maturation of the infant intestinal tract and immune system, but the precise functions of individual HMOs are unclear. To determine the structure-dependent effects of individual HMOs (representing different structural classes) on the intestinal epithelium as well as innate and adaptive immune cells, we assessed fucosylated (2'FL and 3FL), sialylated (3'SL and 6'SL) and neutral non-fucosylated (LNT and LNT2) HMOs for their ability to support intestinal barrier integrity, to stimulate the secretion of chemokines from intestinal epithelial cells, and to modulate cytokine release from LPS-activated dendritic cells (DCs), M1 macrophages (MØs), and co-cultures with naïve CD4+ T cells. The fucosylated and neutral non-fucosylated HMOs increased barrier integrity and protected the barrier following an inflammatory insult but exerted minimal immunomodulatory activity. The sialylated HMOs enhanced the secretion of CXCL10, CCL20 and CXCL8 from intestinal epithelial cells, promoted the secretion of several cytokines (including IL-10, IL-12p70 and IL-23) from LPS-activated DCs and M1 MØs, and increased the secretion of IFN-γ and IL-17A from CD4+ T cells primed by LPS-activated DCs and MØs while reducing the secretion of IL-13. Thus, 3'SL and 6'SL supported Th1 and Th17 responses while reducing Th2 responses. Collectively, our data show that HMOs exert structure-dependent effects on the intestinal epithelium and possess immunomodulatory properties that confer benefits to infants and possibly also later in life.

Mammary epithelium permeability during established lactation: associations with cytokine levels in human milk.

The cytokine profile of human milk may be a key indicator of mammary gland health and has been linked to infant nutrition, growth, and immune system development. The current study examines the extent to which mammary epithelium permeability (MEP) is associated with cytokine profiles during established lactation within a sample of US mothers. Participants were drawn from a previous study of human milk cytokines. The present analysis includes 162 participants (98 Black, 64 White) with infants ranging from 1 to 18 months of age. Levels of cytokines were determined previously. Here we measure milk sodium (Na) and potassium (K) levels with ion-selective probes. Two approaches were used to define elevated MEP: Na levels ≥10 mmol/L and Na/K ratios greater than 0.6. Associations between maternal-infant characteristics, elevated MEP, and twelve analytes (IL-6, IL-8, TNFα, IL-1β, FASL, VEGFD, FLT1, bFGF, PLGF, EGF, leptin, adiponectin) were examined using bivariate associations, principal components analysis, and multivariable logistic regression models. Elevated MEP was observed in 12 and 15% of milk samples as defined by Na and Na/K cutoffs, respectively. The odds of experiencing elevated MEP (defined by Na ≥ 10 mmol/L) were higher among Black participants and declined with older infant age. All cytokines, except leptin, were positively correlated with either Na or the Na/K ratio. A pro-inflammatory factor (IL-6, IL-8, TNFα, IL-1β, EGF) and a tissue remodeling factor (FASL, VEGFD, FLT1, bFGF, PLGF, adiponectin) each contributed uniquely to raising the odds of elevated MEP as defined by either Na or the Na/K ratio. This exploratory analysis of MEP and cytokine levels during established lactation indicates that elevated MEP may be more common in US populations than previously appreciated and that individuals identifying as Black may have increased odds of experiencing elevated MEP based on current definitions. Research aimed at understanding the role of MEP in mammary gland health or infant growth and development should be prioritized.

Immunoregulation of bovine lactoferrin together with osteopontin promotes immune system development and maturation.

The immune system of infants is partly weak and immature, and supplementation of infant formula can be of vital importance to boost the development of the immune system. Lactoferrin (LF) and osteopontin (OPN) are essential proteins in human milk with immunoregulation function. An increasing number of studies indicate that proteins have interactions with each other in milk, and our previous study found that a ratio of LF : OPN at 1 : 5 (w/w, denoted as LOP) had a synergistic effect on intestinal barrier protection. It remains unknown whether LOP can also exert a stronger effect on immunoregulation. Hence, we used an in vitro model of LPS-induced macrophage inflammation and in vivo models of LPS-induced intestinal inflammation and early life development. We showed that LOP increased the secretion of the granulocyte-macrophage colony-stimulating factor (132%), stem cell factor (167%) and interleukin-3 (176%) in bone marrow cells, as well as thymosin (155%) and interleukin-10 (161%) in the thymus, more than LF or OPN alone during development, and inhibited changes in immune cells and cytokines during the LPS challenge. In addition, analysis of the components of digested proteins in vitro revealed that differentially expressed peptides may provide immunoregulation. Lastly, LOP increased the abundance of Rikenellaceae, Muribaculum, Faecalibaculum, and Elisenbergiella in the cecum content. These results imply that LOP is a potential immunomodifier for infants and offers a new theoretical basis for infant formula innovation.

Effect of protein modification in synbiotic infant formula on probiotic metabolic activity and bacterial composition in an infant gut-model.

Aim: Microbial colonization of the neonatal gut is pivotal in priming the infant's immune system. Human milk (HM) is the best nutrition for infants and supports the development of the microbiota due to prebiotic compounds and probiotic microorganisms. When exclusive breastfeeding is not possible, infant formula (IF) with probiotics is a strategy to support the infant's microbiome development. However, knowledge about the effects of the infant gut microbiota and different compounds in IF on individual probiotic strains is limited, as strain-level detection in a complex ecosystem is challenging. The aim of the present study was to show the effects of IF with different protein forms on the metabolic activity of two probiotic strains isolated from HM in a complex ecosystem. Methods: By using an ex-vivo infant gut model containing infant donor-microbiota, the effects of IF with either intact or extensively hydrolyzed protein on the metabolic activity of the donor microbiota, as well as two probiotic strains [Limosilactobacillus fermentum (L. fermentum) CECT 5716 (Lf) and Bifidobacterium breve (B. breve) DSM 32583 (Bb)], were analyzed. A new bioinformatic pipeline combined with a specific infant microbiome database was used to explore shotgun metagenome datasets (1200 Megabases) for taxonomic identification and strain-level tracking. Results: Both protein forms (i.e., intact or extensively hydrolyzed protein) in IF supported infant gut microbial metabolic activity equally, as evidenced by similar levels of short-chain fatty acids (SCFAs). Interestingly, gut microbial metabolic activity was found to be differently activated in a strain-dependent manner. Taxonomic profiling of the microbiome at the strain level enabled monitoring of the prevalence and abundance of both probiotic strains, even in a complex ecosystem. Conclusion: Food matrix and host microbiota interactions should be considered when evaluating strain-specific probiotic effects in the future.

Transfer of celiac disease-associated immunogenic gluten peptides in breast milk: variability in kinetics of secretion.

Exposure to antigens is crucial for child immune system development, aiding disease prevention and promoting infant health. Some common food antigen proteins are found in human breast milk. However, it is unclear whether gluten antigens linked to celiac disease (CD) are transmitted through breast milk, potentially impacting the development of the infant's immune system. This study aimed to analyze the passage of gluten immunogenic peptides (GIP) into human breast milk. We evaluated the dynamics of GIP secretion after lactating mothers adopted a controlled gluten-rich diet. We prospectively enrolled 96 non-CD and 23 CD lactating mothers, assessing total proteins and casein in breast milk, and GIP levels in breast milk and urine. Subsequently, a longitudinal study was conducted in a subgroup of 12 non-CD lactating mothers who adopted a controlled gluten-rich diet. GIP levels in breast milk and urine samples were assayed by multiple sample collections over 96 hours. Analysis of a single sample revealed that 24% of non-CD lactating mothers on a regular unrestricted diet tested positive for GIP in breast milk, and 90% tested positive in urine, with significantly lower concentrations in breast milk. Nevertheless, on a controlled gluten-rich diet and the collection of multiple samples, GIP were detected in 75% and 100% of non-CD participants in breast milk and urine, respectively. The transfer dynamics in breast milk samples were long-enduring and GIP secretion persisted from 0 to 72h. In contrast, GIP secretion in urine samples was limited to the first 24h, with inter-individual variations. In the cohort of CD mothers, 82.6% and 87% tested negative for GIP in breast milk and urine, respectively. This study definitively established the presence of GIP in breast milk, with substantial inter-individual variations in secretion dynamics. Our findings provide insights into distinct GIP kinetics observed in sequentially collected breast milk and urine samples, suggesting differential gluten metabolism patterns depending on the organ or system involved. Future research is essential to understand whether GIP functions as sensitizing or tolerogenic agents in the immune system of breastfed infants.