The molecular characterization of antibody binding to a superantigen-like protein from a commensal microbe.

Microorganisms have coevolved diverse mechanisms to impair host defenses. A major one, superantigens, can result in devastating effects on the immune system. While all known superantigens induce vast immune cell proliferation and come from opportunistic pathogens, recently, proteins with similar broad specificity to antibody variable (V) domain families were identified in a commensal microbiota. These proteins, identified in the human commensal Ruminococcus gnavus, are called immunoglobulin-binding protein (Ibp) A and B and have been shown to activate B cells in vitro expressing either human VH3 or murine VH5/6/7. Here, we provide molecular and functional studies revealing the basis of this Ibp/immunoglobulin (Ig) interaction. The crystal structure and biochemical assays of a truncated IbpA construct in complex with mouse VH5 antigen-binding fragment (Fab) shows a binding of Ig heavy chain framework residues to the Ibp Domain D and the C-terminal heavy chain binding domain (HCBD). We used targeted mutagenesis of contact residues and affinity measurements and performed studies of the Fab-IbpA complex to determine the stoichiometry between Ibp and VH domains, suggesting Ibp may serve to cluster full-length IgA antibodies in vivo. Furthermore, in vitro stimulation experiments indicate that binding of the Ibp HCBD alone is sufficient to activate responsive murine B cell receptors. The presence of these proteins in a commensal microbe suggest that binding a broad repertoire of immunoglobulins, particularly in the gut/microbiome environment, may provide an important function in the maintenance of host/microbiome homeostasis contrasting with the pathogenic role of structurally homologous superantigens expressed by pathogens.

Single probiotic supplement suppresses colitis-associated colorectal tumorigenesis by modulating inflammatory development and microbial homeostasis.

BACKGROUND AND AIM: Chronic inflammation is a major contributor to the initiation and progression of cancers. Lactobacillus helveticus NS8, which was originally separated from fermented koumiss, exhibited anti-inflammatory functions in our prior studies. In this study, NS8 was investigated for its potential to prevent colitis-associated colorectal cancer (CAC). METHODS: The protective effects of NS8 against CAC was explored by employing the azoxymethane plus dextran sodium sulfate-induced carcinogenesis mouse model. The prevalences of T cells expressing specific inflammatory cytokines were measured by flow cytometry at the early stage of CAC. Inflammatory modulation by NS8 was also tested in the Caco2-Raw264.7 cell co-culture system. The alternations in the intestinal microbiota following the health-inflammation-cancer sequence were analyzed by 16S rDNA sequencing. RESULTS: Oral intake of NS8 lactobacilli clearly reduced tumor number and the degree of hyperplasia. The increased proliferation of enterocytes at the early stage of CAC was significantly suppressed by NS8, while the level of apoptosis was elevated. The anticancer effects of NS8 were associated with its anti-colitis outcomes before tumor formation. NS8 significantly suppressed the activation of NF-κB and upregulated the anti-inflammatory cytokine IL-10. Further analysis revealed the marked downregulation of IL-17-producing T cells by NS8. Furthermore, NS8 modulated intestinal dysbiosis by promoting beneficial commensal microbes while suppressing cancer-associated microbes. Notably, Bacteroides acidifaciens was the most sensitive commensal bacteria to NS8 intervention. CONCLUSION: These results provide insight into the protective effects of L. helveticus NS8 against colorectal cancer.

Canna starch improves immune functions and the intestinal environment in mice.

The present study was conducted to elucidate the dietary effects of canna starch on the immune functions and intestinal luminal environment in mice. The amylose and resistant starch characteristics were determined for six types of starch, including edible canna. Canna starch was found to be higher in amylose and resistant starch compared with the other starches. BALB/c mice were fed 3.16% (low-canna group) and 6.32% (high-canna group) canna starch for 2 weeks, and then intestinal parameters were measured. Fecal IgA and mucin levels were markedly elevated by canna starch intake. IgA levels in serum and spleen lymphocytes were elevated by canna starch intake in the high-canna group, but not in the low-canna group. When the mice were fed canna starch, the cecum weight increased, and the pH in the cecum decreased. The high-canna group had significantly increased levels of Clostridium subcluster XIVa lactic acid, acetic acid, and n-butyric acid in the cecum compared with the control group. These results suggested that canna starch supplementation changed the intestinal microbiota and enhanced the intestinal immune and barrier functions and cecal organic acids in mice.

The role of commensal microbes in the lifespan of Drosophila melanogaster.

Commensal microbes have mutualistic relationships with their host and mainly live in the host intestine. There are many studies on the relationships between commensal microbes and host physiology. However, there are inconsistent results on the effects of commensal microbes on host lifespan. To clarify this controversy, we generated axenic flies by using two controlled methods - bleaching and antibiotic treatment - and investigated the relationship between the commensal microbes and host lifespan in Drosophila melanogaster. The removal of microbes by using bleaching and antibiotic treatments without detrimental effects increased fly lifespan. Furthermore, a strain of flies colonized with a high load of microbiota showed a greater effect on lifespan extension when the microbes were eliminated, suggesting that commensal bacteria abundance may be a critical determinant of host lifespan. Consistent with those observations, microbial flora of aged fly gut significantly decreased axenic fly lifespan via an increase in bacterial load rather than through a change of bacterial composition. Our elaborately controlled experiments showed that the elimination of commensal microbes without detrimental side effects increased fly lifespan, and that bacterial load was a significant determinant of lifespan. Furthermore, our results indicate the presence of a deterministic connection between commensal microbes and host lifespan.

Microbial and dietary factors modulating intestinal regulatory T cell homeostasis.

To maintain a quiescent gut microenvironment, proper regulation of immune responses initiated by pro-inflammatory immune subsets is required. Several types of regulatory T cells are reported to exert pivotal roles in achieving this. Among various types of regulatory T cells, the crucial role of Foxp3(+) Treg cells has been well documented. Furthermore, accumulating evidence demonstrates that both microbial and dietary factors influence the induction and suppressor functions of intestinal Foxp3(+) Treg cells. Foxp3(+) Treg cells are a highly activated T cell subset which responds rapidly to environmental and nutritional stimuli. Thus, sufficient nutrient supply is required to fuel the high energetic status of Foxp3(+) Treg cells for the regulation of intestinal immunity.