Tamotsu Kato

Commensal microbe-derived butyrate induces the differentiation of colonic regulatory T cells

Gut commensal microbes shape the mucosal immune system by regulating the differentiation and expansion of several types of T cell. Clostridia, a dominant class of commensal microbe, can induce colonic regulatory T (Treg) cells, which have a central role in the suppression of inflammatory and allergic responses. However, the molecular mechanisms by which commensal microbes induce colonic Treg cells have been unclear. Here we show that a large bowel microbial fermentation product, butyrate, induces the differentiation of colonic Treg cells in mice. A comparative NMR-based metabolome analysis suggests that the luminal concentrations of short-chain fatty acids positively correlates with the number of Treg cells in the colon. Among short-chain fatty acids, butyrate induced the differentiation of Treg cells in vitro and in vivo, and ameliorated the development of colitis induced by adoptive transfer of CD4+ CD45RBhi T cells in Rag1−/− mice. Treatment of naive T cells under the Treg-cell-polarizing conditions with butyrate enhanced histone H3 acetylation in the promoter and conserved non-coding sequence regions of the Foxp3 locus, suggesting a possible mechanism for how microbial-derived butyrate regulates the differentiation of Treg cells. Our findings provide new insight into the mechanisms by which host–microbe interactions establish immunological homeostasis in the gut.

Oral Administration of P. gingivalis Induces Dysbiosis of Gut Microbiota and Impaired Barrier Function Leading to Dissemination of Enterobacteria to the Liver

Although periodontitis has been implicated as a risk factor for various systemic diseases, the precise mechanisms by which periodontitis induces systemic disease remain to be elucidated. We have previously revealed that repeated oral administration of Porphyromonas gingivalis elicits endotoxemia via changes in the gut microbiota of the ileum, and thereby induces systemic inflammation and insulin resistance. However, it is not clear to what extent a single administration of P. gingivalis could affect gut microbiota composition, gut barrier function, and subsequent influx of gut microbiota into the liver. Therefore, in the present study, C57BL/6 mice were orally administered P. gingivalis (strain W83) once and compared to sham-inoculated mice. The phylogenetic structure and diversity of microbial communities in the gut and liver were analyzed by pyrosequencing the 16S ribosomal RNA genes. Serum endotoxin activity was determined by a Limulus amebocyte lysate test. Gene expression in the intestine and expression of 16S rRNA genes in the blood and liver were examined by quantitative polymerase chain reaction. Administration of P. gingivalis significantly altered gut microbiota, with an increased proportion of phylum Bacteroidetes, a decreased proportion of phylum Firmicutes, and increased serum endotoxin levels. In the intestinal tissues, gene expression of tjp-1 and occludin, which are involved in intestinal permeability, were downregulated. Higher amounts of bacterial DNA were detected in the liver of infected mice. Importantly, changes in gut microbiota preceded systemic inflammatory changes. These results further support the idea that disturbance of the gut microbiota composition by orally derived periodontopathic bacteria may be a causal mechanism linking periodontitis and systemic disease.

Probiotic Bifidobacterium longum alters gut luminal metabolism through modification of the gut microbial community

Probiotics are well known as health-promoting agents that modulate intestinal microbiota. However, the molecular mechanisms underlying this effect remain unclear. Using gnotobiotic mice harboring 15 strains of predominant human gut-derived microbiota (HGM), we investigated the effects of Bifidobacterium longum BB536 (BB536-HGM) supplementation on the gut luminal metabolism. Nuclear magnetic resonance (NMR)-based metabolomics showed significantly increased fecal levels of pimelate, a precursor of biotin, and butyrate in the BB536-HGM group. In addition, the bioassay revealed significantly elevated fecal levels of biotin in the BB536-HGM group. Metatranscriptomic analysis of fecal microbiota followed by an in vitro bioassay indicated that the elevated biotin level was due to an alteration in metabolism related to biotin synthesis by Bacteroides caccae in this mouse model. Furthermore, the proportion of Eubacterium rectale, a butyrate producer, was significantly higher in the BB536-HGM group than in the group without B. longum BB536 supplementation. Our findings help to elucidate the molecular basis underlying the effect of B. longum BB536 on the gut luminal metabolism through its interactions with the microbial community.

Multiple Omics Uncovers Host-Gut Microbial Mutualism During Prebiotic Fructooligosaccharide Supplementation

Fructooligosaccharide (FOS), a prebiotic well known for its health-promoting properties, can improve the human gut ecosystem most likely through changes in its microbial composition. However, the detailed mechanism(s) of action of FOS in the modulation of the gut ecosystem remain(s) obscure. Traditional methods of profiling microbes and metabolites could barely show any significant features due to the existence of large interindividual differences, but our novel microbe–metabolite correlation approach, combined with faecal immunoglobulin A (IgA) measurements, has revealed that the induction of mucosal IgA by FOS supplementation correlated with the presence of specific bacteria. Furthermore, the metabolic dynamics of butyrate, l-phenylalanine, l-lysine and tyramine were positively correlated with that of these bacteria and IgA production, whereas p-cresol was negatively correlated. Taken together, our focused intraindividual analysis with omics approaches is a powerful strategy for uncovering the gut molecular network and could provide a new vista for understanding the human gut ecosystem.

The Epithelia-Specific Membrane Trafficking Factor AP-1B Controls Gut Immune Homeostasis in Mice

BACKGROUND & AIMS Epithelial cells that cover the intestinal mucosal surface maintain immune homeostasis and tolerance in the gastrointestinal tract. However, little is known about the molecular mechanisms that regulate epithelial immune functions. Epithelial cells are distinct in that they are highly polarized; this polarity is, at least in part, established by the epithelium-specific polarized sorting factor adaptor protein (AP)-1B. We investigated the role of AP-1B-mediated protein sorting in the maintenance of gastrointestinal immune homeostasis. METHODS The role of AP-1B in intestinal immunity was examined in AP-1B-deficient mice (Ap1m2(-/-)) by monitoring their phenotypes, intestinal morphology, and epithelial barrier functions. AP-1B-mediated protein sorting was examined in polarized epithelial cells from AP-1B knockdown and Ap1m2(-/-) mice. RESULTS Ap1m2(-/-) mice developed spontaneous chronic colitis, characterized by accumulation of interleukin-17A-producing, T-helper 17 cells. Deficiency of AP-1B caused epithelial immune dysfunction, such as reduced expression of antimicrobial proteins and impaired secretion of immunoglobulin A. These defects promoted intestinal dysbiosis and increased bacterial translocation within the mucosa. Importantly, AP-1B deficiency led to mistargeting of a subset of basolateral cytokine receptors to the apical plasma membrane in a polarized epithelial cell line and in colonic epithelial cells from mice. AP1M2 expression was reduced significantly in colonic epithelium samples from patients with Crohns disease. CONCLUSIONS AP-1B is required for proper localization of a subset of cytokine receptors in polarized epithelial cells, which allows them to respond to cytokine signals from underlying lamina propria cells. The AP-1B-mediated protein sorting machinery is required for maintenance of immune homeostasis and prevention of excessive inflammation.

High-affinity monoclonal IgA regulates gut microbiota and prevents colitis in mice.

Immunoglobulin A (IgA) is the main antibody isotype secreted into the intestinal lumen. IgA plays a critical role in the defence against pathogens and in the maintenance of intestinal homeostasis. However, how secreted IgA regulates gut microbiota is not completely understood. In this study, we isolated monoclonal IgA antibodies from the small intestine of healthy mouse. As a candidate for an efficient gut microbiota modulator, we selected a W27 IgA, which binds to multiple bacteria, but not beneficial ones such as Lactobacillus casei. W27 could suppress the cell growth of Escherichia coli but not L. casei in vitro, indicating an ability to improve the intestinal environment. Indeed W27 oral treatment could modulate gut microbiota composition and have a therapeutic effect on both lymphoproliferative disease and colitis models in mice. Thus, W27 IgA oral treatment is a potential remedy for inflammatory bowel disease, acting through restoration of host–microbial symbiosis.

In vitro evaluation method for screening of candidate prebiotic foods

The aim of this work was to develop a simple and rapid in vitro evaluation method for screening and discovery of uncharacterised and untapped prebiotic foods. Using a NMR-based metabolomic approach coupled with multivariate statistical analysis, the metabolic profiles generated by intestinal microbiota after in vitro incubation with feces were examined. The viscous substances of Japanese bunching onion (JBOVS) were identified as one of the candidate prebiotic foods by this in vitro screening method. The JBOVS were primarily composed of sugar components, especially fructose-based carbohydrates. Our results suggested that ingestion of JBOVS contributed to lactate and acetate production by the intestinal microbiota, and were accompanied by an increase in the Lactobacillus murinus and Bacteroidetes sp. populations in the intestine and fluctuation of the host-microbial co-metabolic process. Therefore, our approach should be useful as a rapid and simple screening tool for potential prebiotic foods.

IL-22BP dictates characteristics of Peyer’s patch follicle-associated epithelium for antigen uptake

Interleukin-22 (IL-22) acts protectively and harmfully on intestinal tissue depending on the situation; therefore, IL-22 signaling needs to be tightly regulated. IL-22 binding protein (IL-22BP) binds IL-22 to inhibit IL-22 signaling. It is expressed in intestinal and lymphoid tissues, although its precise distribution and roles have remained unclear. In this study, we show that IL-22BP is highly expressed by CD11b+CD8&agr;− dendritic cells in the subepithelial dome region of Peyer’s patches (PPs). We found that IL-22BP blocks IL-22 signaling in the follicle-associated epithelium (FAE) covering PPs, indicating that IL-22BP plays a role in regulating the characteristics of the FAE. As expected, FAE of IL-22BP–deficient (Il22ra2−/−) mice exhibited altered properties such as the enhanced expression of mucus and antimicrobial proteins as well as prominent fucosylation, which are normally suppressed in FAE. Additionally, Il22ra2−/− mice exhibited the decreased uptake of bacterial antigens into PPs without affecting M cell function. Our present study thus demonstrates that IL-22BP promotes bacterial uptake into PPs by influencing FAE gene expression and function.

Aggravation of collagen-induced arthritis by orally administered Porphyromonas gingivalis through modulation of the gut microbiota and gut immune system

Porhyromonas gingivalis, a causative bacterium of periodontitis, is implicated in the etiology of rheumatoid arthritis (RA), mainly because of expressing peptidyl arginine deiminase (PAD) that generates RA-related autoantigens. However, compared with other periodontopathic bacteria, the precise role of P. gingivalis in RA is largely unknown. We found that orally administered P. gingivalis changed the gut microbiome with concomitant elevation of serum endotoxin and inflammatory markers, and impairment of the gut barrier function. Based on findings showing a relationship between gut microbiota and RA, we investigated whether the change of gut microbiota induced by P. gingivalis and Prevotella intermedia, another periodontopathic bacterium without PAD, is associated with collagen-induced arthritis (CIA). DBA/1J mice were orally administered with or without bacteria followed by induction of CIA. P. gingivalis, but not P. intermedia, administration significantly aggravated arthritis with increased interleukin-17 levels in sera and culture supernatants, increased Th17 cell proportions among mesenteric lymphocytes, and a significant change in the gut microbiome. However, P. gingivalis administration did not elevate the level of anti-citrullinated protein antibody. These results suggest a unique role of P. gingivalis in the link between periodontitis and RA by affecting the gut immune system and the gut microbiota composition.

Intestinal IgA as a modulator of the gut microbiota

ABSTRACT Accumulating evidence suggests that dysbiosis plays a role in the pathogenesis of intestinal diseases including inflammatory bowel disease (IBD) as well as extra-intestinal disorders. As a modulator of the intestinal microbiota, we isolated a mouse monoclonal IgA antibody (clone W27) with high affinities for multiple commensal bacteria, but not for beneficial bacteria such as Lactobacillus casei (L. casei). Via specific recognition of an epitope in serine hydroxymethyltransferase (SHMT), a bacterial metabolic enzyme, W27 IgA selectively inhibited the in vitro growth of bound bacteria, including Escherichia coli (E. coli), while having no effect on unbound beneficial bacteria such as L. casei. By modulating the gut microbiota in vivo, oral administration of W27 IgA effectively prevented development of colitis in several mouse models. Here we discuss how intestinal IgA modulates the gut microbiota through recognition of SHMT.