Specific intestinal microbiota has been shown to induce Foxp3+ regulatory T cell development. However, it remains unclear how development of another regulatory T cell subset, Tr1 cells, is regulated in the intestine. Here, we analyzed the role of two probiotic strains of intestinal bacteria, Lactobacillus casei and Bifidobacterium breve in T cell development in the intestine. B. breve, but not L. casei, induced development of IL-10-producing Tr1 cells that express cMaf, IL-21, and Ahr in the large intestine. Intestinal CD103+ dendritic cells (DCs) mediated B. breve-induced development of IL-10-producing T cells. CD103+ DCs from Il10 −/−, Tlr2 −/−, and Myd88 −/− mice showed defective B. breve-induced Tr1 cell development. B. breve-treated CD103+ DCs failed to induce IL-10 production from co-cultured Il27ra −/− T cells. B. breve treatment of Tlr2 −/− mice did not increase IL-10-producing T cells in the colonic lamina propria. Thus, B. breve activates intestinal CD103+ DCs to produce IL-10 and IL-27 via the TLR2/MyD88 pathway thereby inducing IL-10-producing Tr1 cells in the large intestine. Oral B. breve administration ameliorated colitis in immunocompromised mice given naïve CD4+ T cells from wild-type mice, but not Il10 −/− mice. These findings demonstrate that B. breve prevents intestinal inflammation through the induction of intestinal IL-10-producing Tr1 cells.

Colonic epithelial cells are covered by thick inner and outer mucus layers. The inner mucus layer is free of commensal microbiota, which contributes to the maintenance of gut homeostasis. In the small intestine, molecules critical for prevention of bacterial invasion into epithelia such as Paneth-cell-derived anti-microbial peptides and regenerating islet-derived 3 (RegIII) family proteins have been identified. Although there are mucus layers providing physical barriers against the large number of microbiota present in the large intestine, the mechanisms that separate bacteria and colonic epithelia are not fully elucidated. Here we show that Ly6/PLAUR domain containing 8 (Lypd8) protein prevents flagellated microbiota invading the colonic epithelia in mice. Lypd8, selectively expressed in epithelial cells at the uppermost layer of the large intestinal gland, was secreted into the lumen and bound flagellated bacteria including Proteus mirabilis. In the absence of Lypd8, bacteria were present in the inner mucus layer and many flagellated bacteria invaded epithelia. Lypd8(-/-) mice were highly sensitive to intestinal inflammation induced by dextran sulfate sodium (DSS). Antibiotic elimination of Gram-negative flagellated bacteria restored the bacterial-free state of the inner mucus layer and ameliorated DSS-induced intestinal inflammation in Lypd8(-/-) mice. Lypd8 bound to flagella and suppressed motility of flagellated bacteria. Thus, Lypd8 mediates segregation of intestinal bacteria and epithelial cells in the colon to preserve intestinal homeostasis.

Several subsets of innate immune cells, all with unique properties, reside within the intestinal lamina propria. However, compared with intestinal dendritic cells (DCs), intestinal macrophages are less well characterized. In this study, we examined the properties of macrophages in the colonic lamina propria (LMφ). Colonic DCs (LDC) showed LPS-induced production of IL-12p40. In contrast, LMφ showed constitutive IL-10 production and unresponsiveness to LPS in terms of inflammatory cytokine production. Comparison of the gene expression profiles between LMφ and LDC revealed that LMφ preferentially expressed IL-10-related genes. LMφ obtained from mice lacking IL-10 or Stat3 showed hyperproduction of tumour necrosis factor (TNF)-α and IL-6 in response to LPS. IL-10 production in the large intestine was mainly induced by LMφ and regulatory T cells and was dependent on the presence of commensal microbiota. Accordingly, LMφ from germ-free mice showed less production of IL-10 and increased levels of LPS-induced TNF-α and IL-6 production. Taken together, these results demonstrate that the activity of LMφ to produce pro-inflammatory cytokines is negatively regulated through commensal microbiota-dependent IL-10 production in the large intestine.

Complement plays a key role in host defense, but its dysregulation can cause autologous tissue injury. Complement activation is normally controlled by regulatory proteins, including factor H (FH) in plasma and membrane cofactor protein (MCP) on the cell surface. Mutations in FH and MCP are linked to atypical hemolytic uremic syndrome, a type of thrombotic microangiopathy (TMA) that causes renal failure. We describe here that disruption of FH function on the cell surface can also lead to disseminated complement-dependent macrovascular thrombosis. By gene targeting, we introduced a point mutation (W1206R) into murine FH that impaired its interaction with host cells but did not affect its plasma complement-regulating activity. Homozygous mutant mice carrying this mutation developed renal TMA as well as systemic thrombophilia involving large blood vessels in multiple organs, including liver, lung, spleen, and kidney. Approximately 30% of mutant mice displayed symptoms of stroke and ischemic retinopathy, and 48% died prematurely. Genetic deficiency of complement C3 and factor D prevented both the systemic thrombophilia and renal TMA phenotypes. These results demonstrate a causal relationship between complement dysregulation and systemic angiopathy and suggest that complement activation may contribute to various human thrombotic disorders involving both the micro- and macrovasculature.

Our results provide evidence for the renoprotective action of febuxostat against the formation of interstitial fibrosis. A decrease in macrophage infiltration and interstitial fibrosis, along with a decrease of the oxidative stress marker, strongly suggests the existence of a causal relationship between them. Febuxostat may have therapeutic value in slowing or preventing interstitial fibrosis in patients with CKD.

Extracellular ATP is released from live cells in controlled conditions, as well as dying cells in inflammatory conditions, and, thereby, regulates T cell responses, including Th17 cell induction. The level of extracellular ATP is closely regulated by ATP hydrolyzing enzymes, such as ecto-nucleoside triphosphate diphosphohydrolases (ENTPDases). ENTPDase1/CD39, which is expressed in immune cells, was shown to regulate immune responses by downregulating the ATP level. In this study, we analyzed the immunomodulatory function of ENTPDase7, which is preferentially expressed in epithelial cells in the small intestine. The targeted deletion of Entpd7 encoding ENTPDase7 in mice resulted in increased ATP levels in the small intestinal lumen. The number of Th17 cells was selectively increased in the small intestinal lamina propria in Entpd7−/− mice. Th17 cells were decreased by oral administration of antibiotics or the ATP antagonist in Entpd7−/− mice, indicating that commensal microbiota-dependent ATP release mediates the enhanced Th17 cell development in the small intestinal lamina propria of Entpd7−/− mice. In accordance with the increased number of small intestinal Th17 cells, Entpd7−/− mice were resistant to oral infection with Citrobacter rodentium. Entpd7−/− mice suffered from severe experimental autoimmune encephalomyelitis, which was associated with increased numbers of CD4+ T cells producing both IL-17 and IFN-γ. Taken together, these findings demonstrate that ENTPDase7 controls the luminal ATP level and, thereby, regulates Th17 cell development in the small intestine.

Adequate activation of CD4 + T lymphocytes is essential for host defense against invading pathogens; however, exaggerated activity of effector CD4 + T cells induces tissue damage, leading to inflammatory disorders such as inflammatory bowel diseases. Several unique subsets of intestinal innate immune cells have been identified. However, the direct involvement of innate immune cell subsets in the suppression of T-cell-dependent intestinal inflammation is poorly understood. Here, we report that intestinal CX 3 C chemokine receptor 1 high (CX 3 CR1 high ) CD11b + CD11c + cells are responsible for prevention of intestinal inflammation through inhibition of T-cell responses. These cells inhibit CD4 + T-cell proliferation in a cell contact-dependent manner and prevent T-cell-dependent colitis. The suppressive activity is abrogated in the absence of the IL-10/Stat3 pathway. These cells inhibit T-cell proliferation by two steps. Initially, CX 3 CR1 high CD11b + CD11c + cells preferentially interact with T cells through highly expressed intercellular adhesion molecule-1/vascular cell adhesion molecule-1; then, they fail to activate T cells because of defective expression of CD80/CD86. The IL-10/Stat3 pathway mediates the reduction of CD80/CD86 expression. Transfer of wild-type CX 3 CR1 high CD11b + CD11c + cells prevents development of colitis in myeloid-specific Stat3-deficient mice. Thus, these cells are regulatory myeloid cells that are responsible for maintaining intestinal homeostasis. mucosal immunology | innate immunity

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