June L. Round

The gut microbiota shapes intestinal immune responses during health and disease

Immunological dysregulation is the cause of many non-infectious human diseases such as autoimmunity, allergy and cancer. The gastrointestinal tract is the primary site of interaction between the host immune system and microorganisms, both symbiotic and pathogenic. In this Review we discuss findings indicating that developmental aspects of the adaptive immune system are influenced by bacterial colonization of the gut. We also highlight the molecular pathways that mediate host–symbiont interactions that regulate proper immune function. Finally, we present recent evidence to support that disturbances in the bacterial microbiota result in dysregulation of adaptive immune cells, and this may underlie disorders such as inflammatory bowel disease. This raises the possibility that the mammalian immune system, which seems to be designed to control microorganisms, is in fact controlled by microorganisms.

A microbial symbiosis factor prevents intestinal inflammatory disease.

Humans are colonized by multitudes of commensal organisms representing members of five of the six kingdoms of life; however, our gastrointestinal tract provides residence to both beneficial and potentially pathogenic microorganisms. Imbalances in the composition of the bacterial microbiota, known as dysbiosis, are postulated to be a major factor in human disorders such as inflammatory bowel disease. We report here that the prominent human symbiont Bacteroides fragilis protects animals from experimental colitis induced by Helicobacter hepaticus, a commensal bacterium with pathogenic potential. This beneficial activity requires a single microbial molecule (polysaccharide A, PSA). In animals harbouring B. fragilis not expressing PSA, H. hepaticus colonization leads to disease and pro-inflammatory cytokine production in colonic tissues. Purified PSA administered to animals is required to suppress pro-inflammatory interleukin-17 production by intestinal immune cells and also inhibits in vitro reactions in cell cultures. Furthermore, PSA protects from inflammatory disease through a functional requirement for interleukin-10-producing CD4+ T cells. These results show that molecules of the bacterial microbiota can mediate the critical balance between health and disease. Harnessing the immunomodulatory capacity of symbiosis factors such as PSA might potentially provide therapeutics for human inflammatory disorders on the basis of entirely novel biological principles.

Inducible Foxp3+ regulatory T-cell development by a commensal bacterium of the intestinal microbiota.

To maintain intestinal health, the immune system must faithfully respond to antigens from pathogenic microbes while limiting reactions to self-molecules. The gastrointestinal tract represents a unique challenge to the immune system, as it is permanently colonized by a diverse amalgam of bacterial phylotypes producing multitudes of foreign microbial products. Evidence from human and animal studies indicates that inflammatory bowel disease results from uncontrolled inflammation to the intestinal microbiota. However, molecular mechanisms that actively promote mucosal tolerance to the microbiota remain unknown. We report herein that a prominent human commensal, Bacteroides fragilis, directs the development of Foxp3+ regulatory T cells (Tregs) with a unique “inducible” genetic signature. Monocolonization of germ-free animals with B. fragilis increases the suppressive capacity of Tregs and induces anti-inflammatory cytokine production exclusively from Foxp3+ T cells in the gut. We show that the immunomodulatory molecule, polysaccharide A (PSA), of B. fragilis mediates the conversion of CD4+ T cells into Foxp3+ Treg cells that produce IL-10 during commensal colonization. Functional Foxp3+ Treg cells are also produced by PSA during intestinal inflammation, and Toll-like receptor 2 signaling is required for both Treg induction and IL-10 expression. Most significantly, we show that PSA is not only able to prevent, but also cure experimental colitis in animals. Our results therefore demonstrate that B. fragilis co-opts the Treg lineage differentiation pathway in the gut to actively induce mucosal tolerance.

The Toll-Like Receptor 2 Pathway Establishes Colonization by a Commensal of the Human Microbiota

Signaling through innate immune receptors promotes commensal bacteria colonization of the gut. Mucosal surfaces constantly encounter microbes. Toll-like receptors (TLRs) mediate recognition of microbial patterns to eliminate pathogens. By contrast, we demonstrate that the prominent gut commensal Bacteroides fragilis activates the TLR pathway to establish host-microbial symbiosis. TLR2 on CD4+ T cells is required for B. fragilis colonization of a unique mucosal niche in mice during homeostasis. A symbiosis factor (PSA, polysaccharide A) of B. fragilis signals through TLR2 directly on Foxp3+ regulatory T cells to promote immunologic tolerance. B. fragilis lacking PSA is unable to restrain T helper 17 cell responses and is defective in niche-specific mucosal colonization. Therefore, commensal bacteria exploit the TLR pathway to actively suppress immunity. We propose that the immune system can discriminate between pathogens and the microbiota through recognition of symbiotic bacterial molecules in a process that engenders commensal colonization.

MicroRNA-155 Promotes Autoimmune Inflammation by Enhancing Inflammatory T Cell Development

Mammalian noncoding microRNAs (miRNAs) are a class of gene regulators that have been linked to immune system function. Here, we have investigated the role of miR-155 during an autoimmune inflammatory disease. Consistent with a positive role for miR-155 in mediating inflammatory responses, Mir155(-/-) mice were highly resistant to experimental autoimmune encephalomyelitis (EAE). miR-155 functions in the hematopoietic compartment to promote the development of inflammatory T cells including the T helper 17 (Th17) cell and Th1 cell subsets. Furthermore, the major contribution of miR-155 to EAE was CD4(+) T cell intrinsic, whereas miR-155 was also required for optimum dendritic cell production of cytokines that promoted Th17 cell formation. Our study shows that one aspect of miR-155 function is the promotion of T cell-dependent tissue inflammation, suggesting that miR-155 might be a promising therapeutic target for the treatment of autoimmune disorders.

Coordination of tolerogenic immune responses by the commensal microbiota

All mammals are born ignorant to the existence of micro-organisms. Soon after birth, however, every mammal begins a lifelong association with a multitude of microbes that lay residence on the skin, mouth, vaginal mucosa and gastrointestinal (GI) tract. Approximately 500-1000 different species of microbes have highly evolved to occupy these bodily niches, with the highest density and diversity occurring within the intestine. These organisms play a vital role in mammalian nutrient breakdown and provide resistance to colonization by pathogenic micro-organisms. More recently, however, studies have demonstrated that the microbiota can have a profound and long-lasting effect on the development of our immune system both inside and outside the intestine. While our immune system has evolved to recognize and eradicate foreign entities, it tolerates the symbiotic micro-organisms of the intestine. How and why this tolerance occurs has remained unclear. Here we present evidence that the commensal microbes of the intestine actively induce tolerant responses from the host that coordinate healthy immune responses. Potentially, disruption of this dialogue between the host and microbe can lead to the development of autoimmune diseases such as inflammatory bowel disease (IBD), rheumatoid arthritis (RA), or Type I diabetes (TID). As a wealth of publications have focused on the impact of the microbiota on intestinal immune responses and IBD, this chapter will focus on the extra-intestinal impacts of the microbiota from development to disease and integrate the known mechanisms by which the microbiota is able to actively communicate with its host to promote health.

Exosome-delivered microRNAs modulate the inflammatory response to endotoxin.

MicroRNAs regulate gene expression posttranscriptionally and function within the cells in which they are transcribed. However, recent evidence suggests that microRNAs can be transferred between cells and mediate target gene repression. We find that endogenous miR-155 and miR-146a, two critical microRNAs that regulate inflammation, are released from dendritic cells within exosomes and are subsequently taken up by recipient dendritic cells. Following uptake, exogenous microRNAs mediate target gene repression and can reprogramme the cellular response to endotoxin, where exosome-delivered miR-155 enhances while miR-146a reduces inflammatory gene expression. We also find that miR-155 and miR-146a are present in exosomes and pass between immune cells in vivo, as well as demonstrate that exosomal miR-146a inhibits while miR-155 promotes endotoxin-induced inflammation in mice. Together, our findings provide strong evidence that endogenous microRNAs undergo a functional transfer between immune cells and constitute a mechanism of regulating the inflammatory response.

Defining dysbiosis and its influence on host immunity and disease

Mammalian immune system development depends on instruction from resident commensal microorganisms. Diseases associated with abnormal immune responses towards environmental and self antigens have been rapidly increasing over the last 50 years. These diseases include inflammatory bowel disease (IBD), multiple sclerosis (MS), type I diabetes (T1D), allergies and asthma. The observation that people with immune mediated diseases house a different microbial community when compared to healthy individuals suggests that pathogenesis arises from improper training of the immune system by the microbiota. However, with hundreds of different microorganisms on our bodies it is hard to know which of these contribute to health and more importantly how? Microbiologists studying pathogenic organisms have long adhered to Kochs postulates to directly relate a certain disease to a specific microbe, raising the question of whether this might be true of commensal–host relationships as well. Emerging evidence supports that rather than one or two dominant organisms inducing host health, the composition of the entire community of microbial residents influences a balanced immune response. Thus, perturbations to the structure of complex commensal communities (referred to as dysbiosis) can lead to deficient education of the host immune system and subsequent development of immune mediated diseases. Here we will overview the literature that describes the causes of dysbiosis and the mechanisms evolved by the host to prevent these changes to community structure. Building off these studies, we will categorize the different types of dysbiosis and define how collections of microorganisms can influence the host response. This research has broad implications for future therapies that go beyond the introduction of a single organism to induce health. We propose that identifying mechanisms to re‐establish a healthy complex microbiota after dysbiosis has occurred, a process we will refer to as rebiosis, will be fundamental to treating complex immune diseases.

Dlgh1 coordinates actin polymerization, synaptic T cell receptor and lipid raft aggregation, and effector function in T cells.

Lipid raft membrane compartmentalization and membrane-associated guanylate kinase (MAGUK) family molecular scaffolds function in establishing cell polarity and organizing signal transducers within epithelial cell junctions and neuronal synapses. Here, we elucidate a role for the MAGUK protein, Dlgh1, in polarized T cell synapse assembly and T cell function. We find that Dlgh1 translocates to the immune synapse and lipid rafts in response to T cell receptor (TCR)/CD28 engagement and that LckSH3-mediated interactions with Dlgh1 control its membrane targeting. TCR/CD28 engagement induces the formation of endogenous Lck–Dlgh1–Zap70–Wiskott-Aldrich syndrome protein (WASp) complexes in which Dlgh1 acts to facilitate interactions of Lck with Zap70 and WASp. Using small interfering RNA and overexpression approaches, we show that Dlgh1 promotes antigen-induced actin polymerization, synaptic raft and TCR clustering, nuclear factor of activated T cell activity, and cytokine production. We propose that Dlgh1 coordinates TCR/CD28-induced actin-driven T cell synapse assembly, signal transduction, and effector function. These findings highlight common molecular strategies used to regulate cell polarity, synapse assembly, and transducer organization in diverse cellular systems.

Epistasis between microRNAs 155 and 146a during T cell-mediated antitumor immunity

An increased understanding of antitumor immunity is necessary for improving cell-based immunotherapies against human cancers. Here, we investigated the roles of two immune system-expressed microRNAs (miRNAs), miR-155 and miR-146a, in the regulation of antitumor immune responses. Our results indicate that miR-155 promotes and miR-146a inhibits interferon γ (IFNγ) responses by T cells and reduces solid tumor growth in vivo. Using a double-knockout (DKO) mouse strain deficient in both miR-155 and miR-146a, we have also identified an epistatic relationship between these two miRNAs. DKO mice had defective T cell responses and tumor growth phenotypes similar to miR-155(-/-) mice. Further analysis of the T cell compartment revealed that miR-155 modulates IFNγ expression through a mechanism involving repression of Ship1. Our work reveals critical roles for miRNAs in the reciprocal regulation of CD4(+) and CD8(+) T cell-mediated antitumor immunity and demonstrates the dominant nature of miR-155 during its promotion of immune responses.