Necrotizing enterocolitis (NEC) is characterized by intestinal injury and impaired mucin synthesis. We recently showed that breast milk exosomes from rodents promote intestinal cell viability, ...epithelial proliferation, and stem cell activity, but whether they also affect mucus production is unknown. Therefore, the aim of this study was to investigate the effects of bovine milk-derived exosomes on goblet cell expression in experimental NEC and delineate potential underlying mechanisms of action. Exosomes were isolated from bovine milk by ultracentrifugation and confirmed by Nanoparticle Tracking Analysis and through the detection of exosome membrane markers. To study the effect on mucin production, human colonic LS174T cells were cultured and exposed to exosomes. Compared to control, exosomes promoted goblet cell expression, as demonstrated by increased mucin production and relative expression levels of goblet cell expression markers trefoil factor 3 (TFF3) and mucin 2 (MUC2). In addition, exosome treatment enhanced the expression of glucose-regulated protein 94 (GRP94), the most abundant intraluminal endoplasmic reticulum (ER) chaperone protein that aids in protein synthesis. Furthermore, experimental NEC was induced in mouse pups by hyperosmolar formula feeding, lipopolysaccharide administration and hypoxia exposure on postnatal days 5-9. Milk exosomes were given with each gavage feed. NEC was associated with ileal morphological injury and reduction in MUC2+ goblet cells and GRP94+ cells per villus. Exosome administration to NEC pups prevented these changes. This research highlights the potential novel application of milk-derived exosomes in preventing the development of NEC in high-risk infants when breast milk is not available.
Indigenous microbiota are an essential component in the modern concept of human health, but the composition and functional characteristics of a healthy microbiome remain to be precisely defined. ...Patterns of microbial colonization associated with disease states have been documented, but the health-associated microbial patterns and their functional characteristics are less clear. A healthy microbiome, considered in the context of body habitat or body site, could be described in terms of ecologic stability (i.e., ability to resist community structure change under stress or to rapidly return to baseline following a stress-related change), by an idealized (presumably health-associated) composition or by a desirable functional profile (including metabolic and trophic provisions to the host). Elucidation of the properties of healthy microbiota would provide a target for dietary interventions and/or microbial modifications aimed at sustaining health in generally healthy populations and improving the health of individuals exhibiting disrupted microbiota and associated diseases.
The use of probiotics is increasing in popularity for both the prevention and treatment of a variety of diseases. While a growing number of well-conducted, prospective, randomized, controlled, ...clinical trials are emerging and investigations of underlying mechanisms of action are being undertaken, questions remain with respect to the specific immune and physiological effects of probiotics in health and disease. This Review considers recent advances in clinical trials of probiotics for intestinal disorders in both adult and pediatric populations. An overview of recent in vitro and in vivo research related to potential mechanisms of action of various probiotic formulations is also considered.
Probiotics are derived from traditional fermented foods, from beneficial commensals or from the environment. They act through diverse mechanisms affecting the composition or function of the commensal ...microbiota and by altering host epithelial and immunological responses. Certain probiotic interventions have shown promise in selected clinical conditions where aberrant microbiota have been reported, such as atopic dermatitis, necrotising enterocolitis, pouchitis and possibly irritable bowel syndrome. However, no studies have been conducted that can causally link clinical improvements to probiotic-induced microbiota changes. Whether a disease-prone microbiota pattern can be remodelled to a more robust, resilient and disease-free state by probiotic administration remains a key unanswered question. Progress in this area will be facilitated by: optimising strain, dose and product formulations, including protective commensal species; matching these formulations with selectively responsive subpopulations; and identifying ways to manipulate diet to modify bacterial profiles and metabolism.
The gut microbiota is integral to human health, including maintaining the delicate balance between tolerance and protection against potentially harmful pathogens. A growing body of evidence ...implicates the intestinal microbiome in immune-mediated inflammatory disorders; these data span the spectrum from genetic and environmental disease risk factors, to animal studies (particularly germ-free and gnotobiotic models) and human studies, including evidence of dysbiosis in diseased individuals compared to healthy populations. In this review, we summarize both animal and human data supporting a link between the gut microbiota and inflammatory bowel diseases (IBD) and systemic inflammatory arthritis, as models for chronic inflammatory disorders, while offering a pediatric focus (pediatric IBD and juvenile idiopathic arthritis). We discuss relevant mechanisms related to the crosstalk between the gut microbiota and the innate and adaptive immune system. We close with a brief discussion of emerging microbe-altering interventions, including fecal microbial transplantation and its immunologic effects.
•The gut microbiota influences innate and adaptive immune responses.•Animal models show that experimental colitis and inflammatory arthritis are attenuated in the germ-free state.•Intestinal dysbiosis is observed in IBD and juvenile idiopathic arthritis (JIA).•Both are associated with decreased short-chain fatty acid-producing commensals and increased pathobionts.•Fecal microbial transplantation (FMT) is under investigation for such disorders.
Restoring intestinal flora may improve outcomes in children with acute gastroenteritis. In this multicenter trial, the administration of lactobacillus for 5 days in children with acute ...gastroenteritis was not associated with clinical benefit.
Scope
Necrotizing enterocolitis (NEC) is a leading cause of morbidity and death in preterm infants, occurring more often in formula‐fed than breastfed infants. Studies in both rats and humans show ...that human milk oligosaccharides (HMOs) lower the incidence of NEC, but the mechanism underlying such protection is currently unclear.
Methods and Results
By extracting HMOs from pooled human breastmilk, the impact of HMOs on the intestinal mucin levels in a murine model of NEC are investigated. To confirm the results, the findings are validated by exposing human intestinal epithelial cells and intestinal organoids to HMOs and evaluated for mucin expression. HMO‐gavage to pups increases Muc2 levels and decreases intestinal permeability to macromolecular dextran. HMO‐treated cells have increased Muc2 expression, decreased bacterial attachment and dextran permeability during challenge by enteric pathogens. To identify the mediators involved in HMO induction of mucins, it is demonstrated that HMOs directly induce the expression of chaperone proteins including protein disulfide isomerase (PDI). Suppression of PDI activity removes the protective effects of HMOs on barrier function in vitro as well as NEC protection in vivo.
Conclusions
Taken together, the results provide insights to the possible mechanisms by which HMOs protect the neonatal intestine through upregulation of mucins.
Murine pups are gavage‐fed with pooled human milk oligosaccharides and following induction of necrotizing enterocolitis (NEC) are assessed for intestinal injury and mucin expression. The findings are validated in vitro using human intestinal organoids and two tissue culture epithelial cell lines. It has been shown that human milk oligosaccharide feeding reduces intestinal injury, increases goblet cells, and promotes mucin expression.
Obesity and diabetes are characterized by increased inflammation reflecting disordered control of innate immunity. We reveal a local intestinal intraepithelial lymphocyte (IEL)-GLP-1 receptor ...(GLP-1R) signaling network that controls mucosal immune responses. Glp1r expression was enriched in intestinal IEL preparations and copurified with markers of Tαβ and Tγδ IELs, the two main subsets of intestinal IELs. Exendin-4 increased cAMP accumulation in purified IELs and reduced the production of cytokines from activated IELs but not from splenocytes ex vivo. These actions were mimicked by forskolin, absent in IELs from Glp1r(-/-) mice, and attenuated by the GLP-1R agonist exendin (9-39) consistent with a GLP-1R-dependent mechanism of action. Furthermore, Glp1r(-/-) mice exhibited dysregulated intestinal gene expression, an abnormal representation of microbial species in feces, and enhanced sensitivity to intestinal injury following administration of dextran sodium sulfate. Bone marrow transplantation using wild-type C57BL/6 donors normalized expression of multiple genes regulating immune function and epithelial integrity in Glp1r(-/-) recipient mice, whereas acute exendin-4 administration robustly induced the expression of genes encoding cytokines and chemokines in normal and injured intestine. Taken together, these findings define a local enteroendocrine-IEL axis linking energy availability, host microbial responses, and mucosal integrity to the control of innate immunity.
Neutrophil extracellular traps (NETs) are an essential component of the antimicrobial repertoire and represent an effective means by which neutrophils capture, contain, and kill microorganisms. ...However, the uncontrolled or excessive liberation of NETs also damages surrounding cells and can contribute to disease pathophysiology. Alterations in the gut microbiota, as well as the presence of local and systemic markers of inflammation, are strongly associated with the manifestation of a spectrum of intestinal disorders, including chronic inflammatory bowel disease. Although probiotics exert beneficial effects on gut homeostasis, their direct effect on neutrophils, which are abundant in the setting of intestinal inflammation, remains unclear. In this study, we investigated the effects of nonpathogenic, enteropathogenic, and probiotic bacteria on the dynamics of NET formation. Using murine bone marrow-derived neutrophils and the neutrophil-differentiated human myeloid cell line d.HL-60, we demonstrate for the first time, to our knowledge, that probiotic Lactobacillus rhamnosus strain GG inhibits both PMA- and Staphylococcus aureus-induced formation of NETs. Moreover, probiotic L. rhamnosus strain GG had potent antioxidative activity: dampening reactive oxygen species production and phagocytic capacity of the neutrophils while protecting against cell cytotoxicity. Within the milieu of the gut, this represents a novel mechanism by which probiotics can locally dampen innate immune responses and confer desensitization toward luminal Ags.
Background. Vitamin D, an important modulator of the immune system, has been shown to protect mucosal barrier homeostasis. This study investigates the effects of vitamin D deficiency on ...infection-induced changes in intestinal epithelial barrier function in vitro and on Citrobacter rodentium–induced colitis in mice. Methods. Polarized epithelial Caco2-bbe cells were grown in medium with or without vitamin D and challenged with enterohemorrhagic Escherichia coli O157:H7. Barrier function and tight junction protein expression were assessed. Weaned C57BL/6 mice were fed either a vitamin D-sufficient or vitamin D-deficient diet and then infected with C. rodentium. Disease severity was assessed by histological analysis, intestinal permeability assay, measurement of inflammatory cytokine levels, and microbiome analysis. Results. 1,25(OH)₂D₃ altered E. coli O157:H7-induced reductions in transepithelial electrical resistance (P<.01), decreased permeability (P<.05), and preserved barrier integrity. Vitamin D-deficient mice challenged with C. rodentium demonstrated increased colonic hyperplasia and epithelial barrier dysfunction (P<.0001 and P<.05, respectively). Vitamin D deficiency resulted in an altered composition of the fecal microbiome both in the absence and presence of C. rodentium infection. Conclusions. This study demonstrates that vitamin D is an important mediator of intestinal epithelial defenses against infectious agents. Vitamin D deficiency predisposes to more-severe intestinal injury in an infectious model of colitis.
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