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Background & Aims: Very early onset inflammatory bowel diseases (VEOIBD), including infant disorders, are a diverse group of diseases found in children younger than 6 years of age. They have been associated with several gene variants. Our aim was to identify the genes that cause VEOIBD. Methods: We performed whole exome sequencing of DNA from 1 infant with severe enterocolitis and her parents. Candidate gene mutations were validated in 40 pediatric patients and functional studies were carried out using intestinal samples and human intestinal cell lines. Results: We identified compound heterozygote mutations in the Tetratricopeptide repeat domain 7 (TTC7A) gene in an infant from non-consanguineous parents with severe exfoliative apoptotic enterocolitis; we also detected TTC7A mutations in 2 unrelated families, each with 2 affected siblings. TTC7A interacts with EFR3 homolog B to regulate phosphatidylinositol 4-kinase at the plasma membrane. Functional studies demonstrated that TTC7A is expressed in human enterocytes. The mutations we identified in TTC7A result in either mislocalization or reduced expression of TTC7A. Phosphatidylinositol 4-kinase was found to co-immunoprecipitate with TTC7A; the identified TTC7A mutations reduced this binding. Knockdown of TTC7A in human intestinal-like cell lines reduced their adhesion, increased apoptosis, and decreased production of phosphatidylinositol 4-phosphate. Conclusions: In a genetic analysis, we identified loss of function mutations in TTC7A in 5 infants with VEOIBD. Functional studies demonstrated that the mutations cause defects in enterocytes and T cells that lead to severe apoptotic enterocolitis. Defects in the phosphatidylinositol 4-kinase−TTC7A−EFR3 homolog B pathway are involved in the pathogenesis of VEOIBD. [Copyright &y& Elsevier]

Graft-versus-host disease (GVHD) is induced by donor T cells cotransplanted with the stem cell graft. The main risk factors for the development of GVHD are the degree of HLA disparity between donor and recipient and the precursor frequency and intensity of T cell responses against major or minor histocompatibility antigens. However, allogeneic T cell responses do not occur in a static and sterile environment, but are modulated by the inflammatory milieu in lymphoid and GVHD target tissues that influences the functional characteristics of antigen presenting cells APCs. Here, we summarize our recent findings concerning the pro- and anti-inflammatory function of (APCs) in the gastrointestinal tract, the role of APCs in the induction of GVH and graft-versus-leukemia (GVL) responses, and their influence on suppressive effects mediated by regulatory T cells. [Copyright &y& Elsevier]

Gut microbes have ascended to prominence as key modulators of host immunity, raising the possibility that they could influence the outcome of cancer immunotherapy. Two recent studies address this question by identifying specific gut-resident bacteria as drivers of checkpoint blockade immunotherapy in pre-clinical tumor models. [ABSTRACT FROM AUTHOR]

Background & Aims: Many models of autoimmunity are associated with lymphopenia. Most involve a T-helper cell (Th)1-type disease, including the diabetic BioBreeding (BB) rat. To investigate the roles of identified susceptibility loci in disease pathogenesis, we bred PVG-RT1u , lymphopenia (lyp)/lyp rats, congenic for the iddm1 (RT1u ) and iddm2 (lyp, Gimap5−/−) diabetes susceptibility loci on the PVG background. Surprisingly, these rats developed a spontaneous, progressive, inflammatory bowel disease. To understand the disease pathogenesis, we undertook investigations at the genetic, histologic, and cellular levels. Methods: Genetically lymphopenic rats and congenic wild-type partners were compared for gross pathologic, histologic, and immunologic parameters, the latter including cytokines and autoantibodies. Results: Genetic analysis demonstrated that homozygosity at the lyp locus was required for disease. All rats developed disease, and the median age at humane killing was ∼36 weeks. This panintestinal disease showed a conspicuous eosinophilic infiltrate in the submucosa and muscle layers, but the villi were unaffected. Diseased rats showed splenomegaly and massive enlargement of the mesenteric lymph nodes. This pathology resembles human eosinophilic gastroenteritis, and several further features indicate a Th2 basis. The rats developed high serum IgE and made IgG autoantibodies that detected a nonleukocytic cell present in the intestinal wall of all rats (including germ free). Conclusions: The T-lymphopenic state associated with GIMAP5 deficiency renders rats generally susceptible to T-cell-mediated autoimmunity, but the immunoregulatory bias (Th1/Th2) of any disease depends on other genetic (or environmental) factors. In the present model, we suggest that defective peripheral tolerance to an intestine-specific autoantigen leads to uncontrolled inflammation of the intestinal wall. [Copyright &y& Elsevier]

Regulatory T (TR) cells are a subset of T cells that function to control immune responses. Different populations of TR cells have been described, including thymically derived CD4+CD25+ TR cells and Tr1 cells induced in the periphery through exposure to antigen. A transcription factor, Foxp3, has been identified that is essential for CD4+CD25+ TR cell development and function. There is now evidence that transforming growth factor-β might play a role in this pathway. CD4+CD25+ TR cells proliferate extensively in vivo in an antigen-specific manner, and can respond to both self and foreign peptides. By suppressing excessive immune responses, TR cells play a key role in the maintenance of self-tolerance, thus preventing autoimmune disease, as well as inhibiting harmful inflammatory diseases such as asthma and inflammatory bowel disease. [Copyright &y& Elsevier]

Pathobionts are members of the gut microbiota with the capacity to cause disease when there is malfunctioning intestinal homeostasis. These organisms are thought to be major contributors to the pathogenesis of inflammatory bowel disease (IBD), a group of chronic inflammatory disorders driven by dysregulated responses towards the microbiota. Over two decades have passed since the discovery of Helicobacter hepaticus , a mouse pathobiont which causes colitis in the context of immune deficiency. During this time, we have developed a detailed understanding of the cellular players and cytokine networks which drive H. hepaticus immunopathology. However, we are just beginning to understand the microbial factors that enable H. hepaticus to interact with the host and influence colonic health and disease. Here we review key H. hepaticus -host interactions, their relevance to other exemplar pathobionts and how when maladapted they drive colitis. Further understanding of these pathways may offer new therapeutic approaches for IBD. [ABSTRACT FROM AUTHOR]

MAIT cells are an unconventional T cell population that can be activated through both TCR-dependent and TCR-independent mechanisms. Here, we examined the impact of combinations of TCR-dependent and TCR-independent signals in human CD8+ MAIT cells. TCR-independent activation of these MAIT cells from blood and gut was maximized by extending the panel of cytokines to include TNF-superfamily member TL1A. RNA-seq experiments revealed that TCR-dependent and TCR-independent signals drive MAIT cells to exert overlapping and specific effector functions, affecting both host defense and tissue homeostasis. Although TCR triggering alone is insufficient to drive sustained activation, TCR-triggered MAIT cells showed specific enrichment of tissue-repair functions at the gene and protein levels and in in vitro assays. Altogether, these data indicate the blend of TCR-dependent and TCR-independent signaling to CD8+ MAIT cells may play a role in controlling the balance between healthy and pathological processes of tissue inflammation and repair. • Activation of human MAIT cells is TCR-dependent or TCR-independent and enhanced by TL1A • TCR-dependent and TCR-independent triggering induces distinct transcriptional responses • TCR-dependent triggering of MAIT cells induces a tissue-repair program • Data integration with in vivo studies in mice indicates a shared transcriptome Leng et al. explore the consequences of activation of human MAIT cells via their TCR and/or cytokines, including the gut-associated TNF-superfamily member TL1A. TCR triggering reveals a transcriptional program linked to tissue-repair functions seen in vivo , consistent with a homeostatic role for these cells in epithelia. [ABSTRACT FROM AUTHOR]

Background & Aims Interactions between commensal microbes and the immune system are tightly regulated and maintain intestinal homeostasis, but little is known about these interactions in humans. We investigated responses of human CD4 + T cells to the intestinal microbiota. We measured the abundance of T cells in circulation and intestinal tissues that respond to intestinal microbes and determined their clonal diversity. We also assessed their functional phenotypes and effects on intestinal resident cell populations, and studied alterations in microbe-reactive T cells in patients with chronic intestinal inflammation. Methods We collected samples of peripheral blood mononuclear cells and intestinal tissues from healthy individuals (controls, n = 13−30) and patients with inflammatory bowel diseases (n = 119; 59 with ulcerative colitis and 60 with Crohn’s disease). We used 2 independent assays (CD154 detection and carboxy-fluorescein succinimidyl ester dilution assays) and 9 intestinal bacterial species ( Escherichia coli, Lactobacillus acidophilus, Bifidobacterium animalis subsp lactis, Faecalibacterium prausnitzii, Bacteroides vulgatus, Roseburia intestinalis, Ruminococcus obeum, Salmonella typhimurium, and Clostridium difficile ) to quantify, expand, and characterize microbe-reactive CD4 + T cells. We sequenced T-cell receptor Vβ genes in expanded microbe-reactive T-cell lines to determine their clonal diversity. We examined the effects of microbe-reactive CD4 + T cells on intestinal stromal and epithelial cell lines. Cytokines, chemokines, and gene expression patterns were measured by flow cytometry and quantitative polymerase chain reaction. Results Circulating and gut-resident CD4 + T cells from controls responded to bacteria at frequencies of 40−4000 per million for each bacterial species tested. Microbiota-reactive CD4 + T cells were mainly of a memory phenotype, present in peripheral blood mononuclear cells and intestinal tissue, and had a diverse T-cell receptor Vβ repertoire. These cells were functionally heterogeneous, produced barrier-protective cytokines, and stimulated intestinal stromal and epithelial cells via interleukin 17A, interferon gamma, and tumor necrosis factor. In patients with inflammatory bowel diseases, microbiota-reactive CD4 + T cells were reduced in the blood compared with intestine; T-cell responses that we detected had an increased frequency of interleukin 17A production compared with responses of T cells from blood or intestinal tissues of controls. Conclusions In an analysis of peripheral blood mononuclear cells and intestinal tissues from patients with inflammatory bowel diseases vs controls, we found that reactivity to intestinal bacteria is a normal property of the human CD4 + T-cell repertoire, and does not necessarily indicate disrupted interactions between immune cells and the commensal microbiota. T-cell responses to commensals might support intestinal homeostasis, by producing barrier-protective cytokines and providing a large pool of T cells that react to pathogens. [ABSTRACT FROM AUTHOR]

Background & Aims: Toll-like receptors (TLR) are innate immune receptors involved in recognition of the intestinal microflora; they are expressed by numerous cell types in the intestine, including epithelial cells, myeloid cells, and lymphocytes. Little is known about the relative contributions of TLR signaling in distinct cellular compartments to intestinal homeostasis. We aimed to define the roles of TLR signals in distinct cell types in the induction and regulation of chronic intestinal inflammation. Methods: We assessed the roles of the shared TLR signaling adaptor protein, MyD88, in several complementary mouse models of inflammatory bowel disease, mediated by either innate or adaptive immune activation. MyD88-deficient mice and bone marrow chimeras were used to disrupt TLR signals selectively in distinct cellular compartments in the intestine. Results: MyD88-dependent activation of myeloid cells was required for the development of chronic intestinal inflammation. By contrast, although epithelial cell MyD88 signals were required for host survival, they were insufficient to induce intestinal inflammation in the absence of an MyD88-competent myeloid compartment. MyD88 expression by T cells was not required for their pathogenic and regulatory functions in the intestine. Conclusions: Cellular compartmentalization of MyD88 signals in the intestine allow the maintenance of host defense and prevent deleterious inflammatory responses. [ABSTRACT FROM AUTHOR]

Gut-associated lymphoid tissue (GALT) is a sensor region for luminal content and plays an important role in lymphoid maturation, activation and differentiation. It comprises isolated and aggregated lymphoid follicles, cryptopatches (CPs) and tertiary lymphoid tissue. Innate lymphoid cells (ILCs) play a central role within GALT. Prenatal GALT development is dependent on ILC lymphoid-inducer function. Postnatally, these cells rapidly respond to commensal and pathogenic intestinal bacteria, parasites and food components by polarized cytokine production [such as interleukin (IL)-22, IL-17 or IL-13] and further contribute to GALT formation and function. Here, we discuss how ILCs shape lymphoid intestinal microenvironments and act as amplifier cells for innate and adaptive immune responses. [ABSTRACT FROM AUTHOR]

Gastrointestinal GVHD is the leading cause of GVHD-associated mortality. Regulatory T cells (Tregs) can suppress GVHD [Edinger 2003], whilst a disrupted gut microbiota negatively impacts transplant outcomes [Taur 2014]. Regulatory T cells (Tregs) help to control gut inflammation, but how their positioning within tissues relates to their function is unclear. Gut solitary isolated lymphoid tissues (SILTs) are organised lymphoid aggregates containing Tregs and other immune cells. We aim to understand how the location, interaction and travel of Tregs within SILTs change from steady state to inflammation. We hypothesise that the induction of microbe-specific Tregs within SILTs and their subsequent egress into the surrounding tissue play a critical role in controlling inflammation at barrier surfaces. The Powrie group has expertise in quantitative gut tissue image analysis of complex cell populations. We characterised the location and immune cell composition of caecal and colonic SILTs using confocal imaging of CD4, B220 and Foxp3 markers in specific pathogen free (SPF) and germ-free (GF) C56BL/6J mice. In SPF mice, we observe structurally mature SILTs located in the distal colon, whilst less developed SILTs reside in the caecum and proximal colon. Flow cytometric analysis show differential expression of Treg numbers and subsets within these discrete gut regions. Mature SILTs are well organised, with a central B cell zone, and an outer T cell area containing Tregs. Interestingly, we observe increased numbers of SILTs in GF compared to SPF conditions, with a greater proportion of Tregs within SILTs. Tregs within SILTs are poised to respond to inflammation, and their positioning may play key roles in damage control. Using TCR transgenic mice and in-vivo live imaging, we are currently exploring antigen-specific Treg responses in microbiome-replete and germ-free settings. Understanding the functional role of SILTs in intestinal homeostasis could identify targets for novel therapies in intestinal GVHD. [ABSTRACT FROM AUTHOR]

BCG, the only licensed vaccine against tuberculosis (TB), provides geographically variable protection, an effect ascribed to exposure to environmental mycobacteria (EM). Here we show that altering the intestinal microbiota of mice by early-life infection with the commensal bacterium Helicobacter hepaticus ( Hh ) increases their susceptibility to challenge with Mycobacterium tuberculosis ( Mtb ). Furthermore Hh -infected mice immunised parenterally with the recombinant subunit vaccine, human adenovirus type 5 expressing the immunodominant antigen 85A of Mtb (Ad85A), display a reduced lung immune response and protection against Mtb challenge is also reduced. Expression of interleukin 10 (IL10) messenger RNA is increased in the colon of Hh infected mice. Treatment of Hh- infected Ad85A-immunised mice with anti-IL10 receptor antibody, following challenge with Mtb , restores the protective effect of the vaccine. These data show for the first time that alteration of the intestinal microbiota by addition of a single commensal organism can profoundly influence protection induced by a TB subunit vaccine via an IL10-dependent mechanism, a result with implications for the deployment of such vaccines in the field. [ABSTRACT FROM AUTHOR]

Abstract: Previously we have shown that intradermal (i.d.) immunization with a recombinant adenovirus expressing antigen 85A (Ad85A) induced a strong splenic CD8T cell response in BALB/c mice but a weak lung immune response and did not protect mice against challenge with Mycobacterium tuberculosis (Mtb). After moving to a new animal house, the same i.d. immunization induced a strong lung immune response and the mice were protected against Mtb challenge. Increased numbers of antigen 85A-specific CD8 cells were present in lung tissue but were not recoverable by bronchoalveolar lavage (BAL). Mycobacterial growth was inhibited 21 days after Mtb challenge. In contrast, the effects of intranasal (i.n.) immunization did not change between the animal houses; 85A-specific T cells were recovered by BAL and were able to inhibit Mtb growth early after challenge. The effect of alterations to the environment was investigated by administering BCG or Mycobacterium abscessus in the drinking water, which induced protection against Mtb challenge, while Mycobacterium smegmatis did not. However, when Ad85A was given i.d. at the same time as BCG or M. abscessus, but not M. smegmatis, the protection induced by Ad85A was abolished. Treatment of mice with a CD25 antibody during the challenge period, abolished the suppressive effect of oral mycobacterial administration, suggesting that regulatory T cells (T regs) were involved. These results showed that exposure to environmental microorganisms can alter the protective immune response to a parenterally administered subunit vaccine, a result with important implications for the use of such vaccines in humans. [Copyright &y& Elsevier]

Abstract: CD4+CD25+ regulatory T (TR) cells are a naturally occurring population of T cells that suppress the development of a variety of pathological immune responses. However, as human inflammatory diseases are usually not diagnosed until after the onset of clinical symptoms, it is of great interest to determine whether CD4+CD25+ TR cells can reverse established pathology. To examine this question we have utilized a murine model of human inflammatory bowel disease (IBD), where pathology is triggered by infection of immune deficient RAG−/− mice with the pathogenic bacterium Helicobacter hepaticus. Here we demonstrate that adoptively transferred CD4+CD25+ TR cells can cure established intestinal inflammation that is mediated by innate immune activation in H. hepaticus-infected RAG−/− mice. CD4+CD25+ TR cell-mediated amelioration of innate intestinal pathology was accompanied by a reversal in systemic innate immune activation, but did not involve any detectable anti-bacterial effects, as bacterial colonization levels were unchanged. Cure of established pathology was not achieved using subpopulations of CD4+CD25− T cells, further emphasizing the enhanced regulatory activity of CD4+CD25+ TR cells. [Copyright &y& Elsevier]