Immune tolerance. Group 3 innate lymphoid cells mediate intestinal selection of commensal bacteria-specific CD4⁺ T cells.

The ovarian immune system ages while coping with the two main challenges of the aging ovary before menopause, the inflammatory stimulations due to repeated cycles and the increasing need for clearance of accumulating atretic follicles.

This article introduces a series of reviews covering Microbiome-Immune System Interplay appearing in Volume 279 of Immunological Reviews. Mucosal body surfaces, including the mucosa of the gut, lung, and urogenital tract as well as the skin and potentially other tissues are colonized by a diverse collection of eubacteria, archaea, protozoa, yeasts, and viruses. These taxonomically different organisms build a cooperative ecological community referred to as microbiota, whereas the term microbiome relates to the genomes and genes of the microbiota. The composition of the microbiota dynamically changes depending on external and internal influences such as diet, health state, age, and genetic and environmental factors.1, 2 The number of bacteria in our body roughly equals the number of our own "eukaryotic" cells3 and more than 1000 bacterial species have been reported in human feces out of which 150-300 typically are present in a given individual.1 The microbiome exerts far-reaching consequences on health and disease. It affects the development of cancer, metabolic diseases, type 1 diabetes, inflammation, and influences mood and behavior just to name a few examples. Already before birth the microbiome affects the fetus4 with lasting effects that carry into adolescence and adulthood; the value of microbiota-based diagnostic tests are being explored and first microbiota-directed therapies show exciting results. The critical importance of the microbiota had been realized long before the current activity in microbiota research. Already Louis Pasteur wondered if life is possible without bacteria. At the end of the 19th century, germ-free animal models were developed, providing a clear answer to Pasteur's question and opening the field to perform functional studies on the microbiome. Many exciting studies have been published during the early days of microbiota research. Yet, scientific and public attention to the microbiome has taken a big leap only in the last decade. In fact the number of reports retrieved from the Pubmed database with "microbiota" as the search term increased exponentially during the last years and the field continues to progress at dazzling speed. This development was spurred by the feasibility and cost effectiveness of next-generation sequencing (NGS), which allowed for the systematic description of microbial communities independent of conventional microbiological culture. First NGS-based descriptions of microbial communities used 16S rDNA sequencing to provide snapshots of "what's out there," and it is a bit more than 10 years since the first metagenomic analysis of the gut microbiota has been reported.5 Today, comprehensive 16S rDNA, metagenomic sequencing, and transcriptional profiling allow for in-depth characterization of microbial communities and a wealth of data has been collected linking microbiome data to disease states, nutrition, age, and other factors. Despite these impressive technical developments and exciting studies, most studies in human microbiome research remain correlative in nature. Depending on personal life style, genetic predisposition, and numerous external factors, the microbiome can exist in different states without any signs of disease.6, 7 Such natural variations, along with the technical challenges to standardized microbiome analysis, make it difficult to define a healthy microbiome. For example, in inflammatory bowel disease (IBD) an altered state of the microbiome, also known as dysbiosis, has been well documented. Numerous studies reported a reduction in species richness (diversity), imbalance between Firmicutes and Bacteriodetes, and loss of protective bacterial species, eg, Faecalibacterium prausnitzii, in IBD patients compared to healthy individuals, suggesting a role of the microbiome in the pathogenesis of IBD. However, whether these changes emerge as a consequence of the disease or indeed play a causative or perpetuating functional role remains controversial.8 Thus, today a major challenge in the field is to refine the experimental approaches to discriminate cause and consequence and understand the functionality of the microbiome. Causality of microbiome alterations can be established by microbiota transfer experiments. In IBD and Clostridium difficile-associated colitis patients, fecal microbiota transfer was successfully applied to treat disease,9, 10 establishing a causal relationship between alteration of the microbiota and disease. Many more examples have been reported in animal experiments including germ-free animals transferred with dysbiotic human microbiota. Causality should not be confused with functionality that establishes the molecular and cellular interaction between the microbiome and host. Functionality so far has been established only in few cases. Yet it will be key to exploit microbiome-based diagnostics and therapeutic interventions. This volume focuses on the interplay between the microbiome and the immune system, in particular the interaction of the gut microbiota and the intestinal immune system. Germ-free mice, ie, mice that lack any live microorganisms, are characterized by an underdeveloped immune system, an observation reported more than 50 years back.11 Germ-free mice have largely reduced numbers of plasma cells and T cells in their intestinal lamina propria. Peyer's patches, secondary lymphoid organs in the intestine that serve the induction of immune responses, are smaller and have minute germinal centers compared to colonized mice. Similarly, development of isolated lymphoid follicles, tiny lymphoid clusters that share some anatomical and functional properties with Peyer's patches, is stalled in germ-free mice. Yet other immune cell populations seem less affected. For example, eosinophils and macrophages are present at comparable numbers in germ-free and colonized mice and many of their functions seem unaffected by the lack of microbiota. Colonization of germ-free mice results in normalization of their immune status and most of the differences characterizing the deviation from the colonized state disappear in formerly germ-free animals within a few weeks. Thus, immune maturation is clearly driven by and a consequence of the microbiota. Curiously, in most cases, the type of colonizing microbes seems to play a minor role and many bacteria seem capable of triggering immune maturation in germ-free animals. However, the various interactions between the microbiota and the immune system that shape the situation that we are considering as healthy are only beginning to emerge and are discussed in various reviews in this volume. Many microbiome-mediated effects on the immune system seem linked to their metabolic activity. Microbiota metabolize dietary components and host factors to yield bioactive compounds which can affect immune cells either directly or indirectly (Figure 1A). As an example, indigestible polysaccharides are fermented to yield short-chain fatty acids (SCFA) that exert broad immunmodulatory functions, such as favoring differentiation and survival of regulatory T cells. Curiously, efficient Treg induction and a disease-modulating function of the microbiota required the presence of a complex consortium of Clostridia and could not be achieved by single bacterial species.12 SCFA, mostly acetate, propionate, and butyrate, differ with respect to their in vivo distribution and signaling function. Although butyrate is largely used as energy source by epithelial cells in the colon and enters systemic compartments only at low concentrations, acetate, depending of the diet and composition of the microbiota, reaches high micro-molar concentrations in blood. Besides metabolites that come from dietary components, the microbiota convert host factors such as primary bile salts that become deconjugated and metabolized to secondary bile salts. Primary and secondary bile salts display altered signaling functions and differential effects on the microbiota (reviewed by Schubert et al.13). Additionally, the microbiota de novo synthesizes compounds with distinct immune-modulatory functions and interactions of the microbiota and its constituents can modulate immune function (Figure 1B). Some functions seem to be linked to defined members of the microbiota, whereas other pathways seem served by a broad range of bacterial species. The filamentous bacteria (SFB) represent a particularly curious case. SFB constitute a group of bacteria in the family of Clostridiaceae that, in mice, colonize mostly in the ileum and associated Peyer's patches. SFB are present in some but not all animal facilities, and the presence or absence of SFB correlates with the frequency of TH17 T cells in the lamina propria and disease severity in various experimental models. Moreover, introduction of SFB to the microbiota of SFB-free animals triggers TH17 differentiation showing that the presence of a single bacterial species can indeed exert a major influence on immune development and disease susceptibility. The capacity of SFB to modulate the immune system seems linked to the production of serum amyloid A and various cytokines, yet how SFB are sensed, if and how they are different from other bacteria adhering to epithelial cells as well as the precise signaling pathways triggered by SFB are unresolved.14 Thomas Clavel and colleagues have compiled key findings on the interaction between defined members of the microbiota and the immune system, in particular emphasizing taxa that can be cultured.15 Interaction between the microbiome and the immune system affects numerous immune cell types of which in particular the interaction with innate lymphoid cells (ILC), unconventional lymphocytes, and conventional lymphocytes are discussed in three articles of this volume.16-18 ILC are particularly enriched in barrier tissues, ie, tissues typically directly exposed to the microbiota. The transcriptional and epigenetic landscape of ILC and potentially their function is affected by the microbiome.19 ILC have not been reported to produce effector molecules that directly target the microbiota but instead seem to regulate the microbiome indirectly through epithelial cells and by other mechanisms. Microbial metabolites of tryptophan trigger the aryl hydrocarbon receptor and thereby contribute to the expression of interleukin (IL)22. IL22 is a key cytokine in ILC functionality and induces the production of the regenerating islet-derived protein 3 γ (Reg3γ) by epithelial cells (Figure 1A and C).20 Reg3γ is a c-type lectin with antibacterial activity that affects the anatomical distribution and composition of the microbiota.21 Many more ILC-dependent effects on the microbiota are linked to their role in lymphoid organogenesis and innate and adaptive immune responses. The intricate interactions between the microbiome and ILC are discussed in this volume.16 Between innate immune cells and ILC on the one hand and conventional adaptive immune cells on the other, unconventional immune cells, including γδ T cells and NK cells, are another curious immune population characteristic of barrier tissues. Besides ILC, unconventional lymphocytes complement the gap between classical innate and adaptive immune cells and may have evolved to regulate the microbiota. Microbiota-derived sphingolipids restrict the accumulation of invariant natural killer cells (iNKT) during neonatal development22 (Figure 1B), whereas another type of unconventional lymphocytes, the mucosal associated invariant T cells, are absent from germ-free mice. The interesting interrelation of unconventional lymphocytes and the microbiota is summarized by Pasman and Kasper.17 Conventional adaptive immune responses and their reciprocal interactions with the microbiome are discussed in two articles.18, 23 Although multiple pathways are emerging of how the microbiome directly or indirectly shapes the immune system, there is still scarce information on how immune cells control the microbiota. It is, however, clear that a central role in this is taken by secretory antibodies. Polymeric Ig receptor transports multimeric Ig across the epithelial barrier. At the apical side, a fragment of the pIgR linked to the plasma cell-derived immunoglobulin is released into the lumen as secretory Ig (SIg). In the gut lumen, SIg binds to the microbiota (Figure 1C) and can affect their growth,24 result in aggregation and excretion by a process called immune exclusion, and can also anchor SIg-coated microorganism in the mucus layer or facilitate their sampling by M cells (summarized in Ref. 25). Frequently, this process is solely discussed in the context of immunoglobulin A and in fact IgA-producing plasma cells in the mucosa outnumber other isotypes. However, in contrast to mice, humans constitutively harbor a more numerous population of IgM-producing plasma cells and IgM is known to compensate for lack of IgA in IgA-deficient individuals. Thus, in addition to IgA, other antibody isotypes, as well as systemic microbiota-directed antibody responses need to be considered. McCoy et al.18 discussed the wider implication of systemic antibodies directed against the microbiota and the specificity of microbiota-activated lymphocytes. A comprehensive view of microbiome-immune interaction including systemic effects of the microbiome and colonization resistance is given by Pickard et al.23. Colonization resistance describes the capacity of the endogenous microbiota to exclude invading bacteria and to inhibit the overgrowth of indigenous minority bacteria within the intestinal tract. Colonization resistance is compromised by antibiotic administration. The ramifications of perturbed colonization resistance in hospital setting are summarized previously.26 The smallest entity of the microbiota is bacteriophages. Bacteriophages are present in all niches of the body, promote rapid exchange of genetic material and exert significant selective pressure on their bacterial hosts. However, a functional characterization of how bacteriophages affect the microbiome is largely missing. In this volume, Jeremy Barr discussed the intra-body phagosome and described a framework to study the functional impact of bacteriophages.27 Finally, the series of articles is concluded by Deines and colleagues. Discussing microbiome-host interactions in Hydra, the authors delineate fundamental principles in the organization and setup of a host plus microbiota metaorganism.28 This work was supported by Deutsche Forschungsgemeinschaft grant DFG PA921/4-1 to O. P. I apologize to all the authors publishing relevant work that I could not mention in this brief overview. In the intestine epithelial cells, chemical and immunological barriers separate the outside world—the microbiota, dietary components, and potential pathogens—from the inside of our body. This barrier is shaped by a close crosstalk between the microbiota and the immune system. Figure by Ana Izcue and Claudia Bentley. The author declares no conflict of interest.

The Aryl Hydrocarbon Receptor: A Sentinel Safeguarding the Survival of Immune Cells in the Gut

ILC3-derived acetylcholine promotes protease-driven allergic lung pathology

Single-Cell Analyses of Colon and Blood Reveal Distinct Immune Cell Signatures of Ulcerative Colitis and Crohn’s Disease

Background:Emerging evidence points to the critical role of the innate immune system in the pathogenesis of inflammatory bowel disease (IBD). Innate lymphoid cells (ILCs) are a recently described immune cell population that is commonly found, but not limited to, mucosal surfaces, that interacts dire

DICE: Transcriptomic Profiling and Analysis of Impact of Genetic Variation on Rare Immune Cell Populations

Transcription factor T-bet regulates intestinal inflammation mediated by innate lymphoid cells with the interleukin-7 receptor

Immunity at the Barriers

IBD Genetics: Focus on (Dys) Regulation in Immune Cells and the Epithelium

Innate lymphoid cell-derived cytokines in autoimmune diseases

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the intestine that encompasses Crohn’s disease (CD) and ulcerative colitis. The cause of IBD is unknown, but the evidence suggests that an aberrant immune response toward the commensal bacterial flora is responsible for disease in genetically susceptible individuals. Results from animal models of colitis and human studies indicate a role for innate lymphoid cells (ILC) in the pathogenesis of chronic intestinal inflammation in IBD. ILC are a population of lymphocytes that are enriched at mucosal sites, where they play a protective role against pathogens including extracellular bacteria, helminthes, and viruses. ILC lack an antigen-specific receptor, but can respond to environmental stress signals contributing to the rapid orchestration of an early immune response. Several subsets of ILC reflecting functional characteristics of T helper subsets have been described. ILC1 express the transcription factor T-bet and are characterized by secretion of IFNγ, ILC2 are GATA3+ and secrete IL5 and IL13 and ILC3 depend on expression of RORγt and secrete IL17 and IL22. However, ILC retain a degree of plasticity depending on exposure to cytokines and environmental factors. IL23 responsive ILC have been implicated in the pathogenesis of colitis in several innate murine models through the production of IL17, IFNγ, and GM-CSF. We have previously identified IL23 responsive ILC in the human intestine and found that they accumulate in the inflamed colon and small bowel of patients with CD. Other studies have confirmed accumulation of ILC in CD with increased frequencies of IFNγ-secreting ILC1 in both the intestinal lamina propria and the epithelium. Moreover, IL23 driven IL22 producing ILC have been shown to drive bacteria-induced colitis-associated cancer in mice. Interestingly, our data show increased ILC accumulation in patients with IBD and primary sclerosing cholangitis, who carry an increased risk of developing colorectal cancer. ILC may play an important amplifying role in IBD and IBD-associated cancer, through secretion of inflammatory cytokines and interaction with other immune and non-immune cells. Here, we will review the evidence indicating a role for ILC in the pathogenesis of chronic intestinal inflammation.

Innate lymphoid cells (ILCs) are highly plastic and predominantly mucosal tissue-resident cells that contribute to both homeostasis and inflammation depending on the microenvironment. The discovery of naïve-like ILCs suggests an ILC differentiation process that is akin to naïve T cell differentiation. Delineating the mechanisms that underlie ILC differentiation in tissues is crucial for understanding ILC biology in health and disease. Here, we showed that tonsillar ILCs expressing CD45RA lacked proliferative activity, indicative of cellular quiescence. CD62L distinguished two subsets of CD45RA+ ILCs. CD45RA+CD62L+ ILCs (CD62L+ ILCs) resembled circulating naïve ILCs because they lacked the transcriptional, metabolic, epigenetic, and cytokine production signatures of differentiated ILCs. CD45RA+CD62L- ILCs (CD62L- ILCs) were epigenetically similar to CD62L+ ILCs but showed a transcriptional, metabolic, and cytokine production signature that was more akin to differentiated ILCs. CD62L+ and CD62L- ILCs contained uni- and multipotent precursors of ILC1s/NK cells and ILC3s. Differentiation of CD62L+ and CD62L- ILCs led to metabolic reprogramming including up-regulation of genes associated with glycolysis, which was needed for their effector functions after differentiation. CD62L- ILCs with preferential differentiation capacity toward IL-22-producing ILC3s accumulated in the inflamed mucosa of patients with inflammatory bowel disease. These data suggested distinct differentiation potential of CD62L+ and CD62L- ILCs between tissue microenvironments and identified that manipulation of these cells is a possible approach to restore tissue-immune homeostasis.

Innate Lymphoid Cells and Celiac Disease: Current Perspective.

Simple SummaryInnate lymphoid cells (ILCs), a family of innate immune cells including natural killers (NKs), play a multitude of roles in first-line cancer control, in escape from immunity and in cancer progression. In this review, we summarize preclinical and clinical data on ILCs and NK cells concerning their phenotype, function and clinical applications in cellular therapy trials. We also describe how single-cell transcriptome sequencing has been used and forecast how it will be used to better understand ILC and NK involvement in cancer control and progression as well as their therapeutic potential.The role of innate lymphoid cells (ILCs), including natural killer (NK) cells, is pivotal in inflammatory modulation and cancer. Natural killer cell activity and count have been demonstrated to be regulated by the expression of activating and inhibitory receptors together with and as a consequence of different stimuli. The great majority of NK cell populations have an anti-tumor activity due to their cytotoxicity, and for this reason have been used for cellular therapies in cancer patients. On the other hand, the recently classified helper ILCs are fundamentally involved in inflammation and they can be either helpful or harmful in cancer development and progression. Tissue niche seems to play an important role in modulating ILC function and conversion, as observed at the transcriptional level. In the past, these cell populations have been classified by the presence of specific cellular receptor markers; more recently, due to the advent of single-cell RNA sequencing (scRNA-seq), it has been possible to also explore them at the transcriptomic level. In this article we review studies on ILC (and NK cell) classification, function and their involvement in cancer. We also summarize the potential application of NK cells in cancer therapy and give an overview of the most recent studies involving ILCs and NKs at scRNA-seq, focusing on cancer. Finally, we provide a resource for those who wish to start single-cell transcriptomic analysis on the context of these innate lymphoid cell populations.