Gut complement induced by the microbiota combats pathogens and spares commensals.

Canonically, the complement system is known for its rapid response to remove microbes in the bloodstream. However, relatively little is known about a functioning complement system on intestinal mucosal surfaces. Herein, we report the local synthesis of complement component 3 (C3) in the gut, primarily by stromal cells. C3 is expressed upon commensal colonization and is regulated by the composition of the microbiota in healthy humans and mice, leading to an individual host's specific luminal C3 levels. The absence of membrane attack complex (MAC) components in the gut ensures that C3 deposition does not result in the lysis of commensals. Pathogen infection triggers the immune system to recruit neutrophils to the infection site for pathogen clearance. Basal C3 levels directly correlate with protection against enteric infection. Our study reveals the gut complement system as an innate immune mechanism acting as a vigilant sentinel that combats pathogens and spares commensals.

The porcine large intestine contains developmentally distinct submucosal lymphoid clusters and mucosal isolated lymphoid follicles.

Gut-associated lymphoid tissues (GALT) serve as key priming sites for intestinal adaptive immune responses. Most of our understanding of GALT function and development arises from studies in mice. However, the diversity, structure and cellular composition of GALT differs markedly between mammalian species and the developmental window in which distinct GALT structures develop in large mammals remains poorly understood. Given the importance of pigs as models of human disease, as well as their role in livestock production, we adapted a recently developed protocol for the isolation of human GALT to assess the diversity, development and immune composition of large intestinal GALT in neonatal and adult pigs. We demonstrate that the large intestine of adult pigs contains two major GALT types; multifollicular submucosal GALT that we term submucosal lymphoid clusters (SLC) which develop prenatally, and as yet undescribed mucosal isolated lymphoid follicles (M-ILF), which arise after birth. Using confocal laser microscopy and flow cytometry, we additionally assess the microanatomy and lymphocyte composition of SLC and M-ILF, compare them to jejunal Peyer's patches (PP), and describe the maturation of these structures. Collectively, our results provide a deeper understanding of the diversity and development of GALT within the porcine large intestine.

Colitis and Colorectal Carcinogenesis: The Focus on Isolated Lymphoid Follicles.

Gut-associated lymphoid tissue is one of the most diverse and complex immune compartments in the human body. The subepithelial compartment of the gut consists of immune cells of innate and adaptive immunity, non-hematopoietic mesenchymal cells, and stem cells of different origins, and is organized into secondary (and even tertiary) lymphoid organs, such as Peyer's patches, cryptopatches, and isolated lymphoid follicles. The function of isolated lymphoid follicles is multifaceted; they play a role in the development and regeneration of the large intestine and the maintenance of (immune) homeostasis. Isolated lymphoid follicles are also extensively associated with the epithelium and its conventional and non-conventional immune cells; hence, they can also function as a starting point or maintainer of pathological processes such as inflammatory bowel diseases or colorectal carcinogenesis. These relationships can significantly affect both physiological and pathological processes of the intestines. We aim to provide an overview of the latest knowledge of isolated lymphoid follicles in colonic inflammation and colorectal carcinogenesis. Further studies of these lymphoid organs will likely lead to an extended understanding of how immune responses are initiated and controlled within the large intestine, along with the possibility of creating novel mucosal vaccinations and ways to treat inflammatory bowel disease or colorectal cancer.

Suppression of plasmacytoid dendritic cell migration to colonic isolated lymphoid follicles abrogates the development of colitis.

BACKGROUND: Dendritic cells (DCs) play a pivotal role in maintaining immunological homeostasis by orchestrating innate and adaptive immune responses via migration to inflamed sites and the lymph nodes (LNs). Plasmacytoid DCs (pDCs) have been reported to accumulate in the colon of inflammatory bowel disease (IBD) patients and dextran sulfate sodium (DSS)-induced colitis mice. However, the role of pDCs in the progression of colonic inflammation remains unclear. METHODS: 80 compounds in natural medicines were searched for inhibitors of pDC migration using bone marrow-derived pDCs (BMpDCs) and conventional DCs (BMcDCs). BALB/c mice were given 3% DSS in the drinking water to induce acute colitis. Compounds, which specifically inhibited pDC migration, were administrated into DSS-induced colitis mice. FINDINGS: Astragaloside IV (As-IV) and oxymatrine (Oxy) suppressed BMpDC migration but not BMcDC migration. In DSS-induced colitis mice, the number of pDCs was markedly increased in the colonic lamina propria (LP), and the expression of CCL21 was obviously observed in colonic isolated lymphoid follicles (ILFs). As-IV and Oxy reduced symptoms of colitis and the accumulation of pDCs in colonic ILFs but not in the colonic LP. Moreover, in a BMpDC adoptive transfer model, BMpDC migration to colonic ILFs was significantly decreased by treatment with As-IV or Oxy. INTERPRETATION: pDCs accumulated in the colon of colitis mice, and As-IV and Oxy ameliorated colitis by suppressing pDC migration to colonic ILFs. Accordingly, the selective inhibition of pDC migration may be a potential therapeutic approach for treating colonic inflammatory diseases.

Group 3 Innate Lymphoid Cells Program a Distinct Subset of IL-22BP-Producing Dendritic Cells Demarcating Solitary Intestinal Lymphoid Tissues.

Solitary intestinal lymphoid tissues such as cryptopatches (CPs) and isolated lymphoid follicles (ILFs) constitute steady-state activation hubs containing group 3 innate lymphoid cells (ILC3) that continuously produce interleukin (IL)-22. The outer surface of CPs and ILFs is demarcated by a poorly characterized population of CD11c+ cells. Using genome-wide single-cell transcriptional profiling of intestinal mononuclear phagocytes and multidimensional flow cytometry, we found that CP- and ILF-associated CD11c+ cells were a transcriptionally distinct subset of intestinal cDCs, which we term CIA-DCs. CIA-DCs required programming by CP- and ILF-resident CCR6+ ILC3 via lymphotoxin-ß receptor signaling in cDCs. CIA-DCs differentially expressed genes associated with immunoregulation and were the major cellular source of IL-22 binding protein (IL-22BP) at steady state. Mice lacking CIA-DC-derived IL-22BP exhibited diminished expression of epithelial lipid transporters, reduced lipid resorption, and changes in body fat homeostasis. Our findings provide insight into the design principles of an immunoregulatory checkpoint controlling nutrient absorption.

Intestinal B cells in the red-eared slider turtle, Trachemys scripta: Anatomical distribution and implications for ecological interactions with pathogenic microbes.

Disease is a significant threat in the global decline of reptile species. Many aquatic reptiles live in habitats with high levels of opportunistic microbial pathogens, yet little is known about their immune system. Gut-associated lymphoid tissue is vital for protection against ingested pathogens and maintenance of normal gut microbiota. In mammals, gut mucosal immunity is well-characterized and mucosal surfaces are coated in protective antibodies. However, reptiles lack lymph nodes and Peyer's patches, which are the major sites of mammalian B cell responses. The presence or distribution of mucosal B cells in reptiles is unknown. In this study, we first set out to determine if B cells could be detected in intestinal tissues of red-eared slider turtles, Trachemys scripta. Using whole-mount immunochemistry and a primary antibody to turtle antibody light chains, we identified widely distributed B cell aggregates within the small intestine of hatchling turtles. These aggregates appeared similar to isolated lymphoid follicles (ILFs) in mammals and the frequency was much higher in distal intestinal sections than in proximal sections. To determine if these structures were inducible in the presence of microbes, we introduced an enteric Salmonella species through oral gavage. Analysis of intestinal tissues revealed that hatchlings exposed to Salmonella exhibited significantly more of these aggregates when compared with those that did not receive bacteria. These studies provide the first evidence for B cell-containing ILF-like structures in reptiles and provide novel information about gut immunity in nonmammalian vertebrates that could have important implications for ecological interactions with pathogens.

Development and Function of Secondary and Tertiary Lymphoid Organs in the Small Intestine and the Colon.

The immune system of the gut has evolved a number of specific lymphoid structures that contribute to homeostasis in the face of microbial colonization and food-derived antigenic challenge. These lymphoid organs encompass Peyer's patches (PP) in the small intestine and their colonic counterparts that develop in a programed fashion before birth. In addition, the gut harbors a network of lymphoid tissues that is commonly designated as solitary intestinal lymphoid tissues (SILT). In contrast to PP, SILT develop strictly after birth and consist of a dynamic continuum of structures ranging from small cryptopatches (CP) to large, mature isolated lymphoid follicles (ILF). Although the development of PP and SILT follow similar principles, such as an early clustering of lymphoid tissue inducer (LTi) cells and the requirement for lymphotoxin beta (LTß) receptor-mediated signaling, the formation of CP and their further maturation into ILF is associated with additional intrinsic and environmental signals. Moreover, recent data also indicate that specific differences exist in the regulation of ILF formation between the small intestine and the colon. Importantly, intestinal inflammation in both mice and humans is associated with a strong expansion of the lymphoid network in the gut. Recent experiments in mice suggest that these structures, although they resemble large, mature ILF in appearance, may represent de novo-induced tertiary lymphoid organs (TLO). While, so far, it is not clear whether intestinal TLO contribute to the exacerbation of inflammatory pathology, it has been shown that ILF provide the critical microenvironment necessary for the induction of an effective host response upon infection with enteric bacterial pathogens. Regarding the importance of ILF for intestinal immunity, interfering with the development and maturation of these lymphoid tissues may offer novel means for manipulating the immune response during intestinal infection or inflammation.

The bilateral responsiveness between intestinal microbes and IgA.

The immune system has developed strategies to maintain a homeostatic relationship with the resident microbiota. IgA is central in holding this relationship, as the most dominant immunoglobulin isotype at the mucosal surface of the intestine. Recent studies report a role for IgA in shaping the composition of the intestinal microbiota and exploit strategies to characterise IgA-binding bacteria for their inflammatory potential. We review these findings here, and place them in context of the current understanding of the range of microorganisms that contribute to the IgA repertoire and the pathways that determine the quality of the IgA response. We examine why only certain intestinal microbes are coated with IgA, and discuss how understanding the determinants of this specific responsiveness may provide insight into diseases associated with dysbiosis.

Re-thinking the functions of IgA(+) plasma cells.

The intestinal mucosa harbors the largest population of antibody (Ab)-secreting plasma cells (PC) in the human body, producing daily several grams of immunoglobulin A (IgA). IgA has many functions, serving as a first-line barrier that protects the mucosal epithelium from pathogens, toxins and food antigens (Ag), shaping the intestinal microbiota, and regulating host-commensal homeostasis. Signals induced by commensal colonization are central for regulating IgA induction, maintenance, positioning and function and the number of IgA(+) PC is dramatically reduced in neonates and germ-free (GF) animals. Recent evidence demonstrates that the innate immune effector molecules tumor necrosis factor α (TNFα) and inducible nitric oxide synthase (iNOS) are required for IgA(+) PC homeostasis during the steady state and infection. Moreover, new functions ascribed to PC independent of Ab secretion continue to emerge, suggesting that PC, including IgA(+) PC, should be re-examined in the context of inflammation and infection. Here, we outline mechanisms of IgA(+) PC generation and survival, reviewing their functions in health and disease.

Transition from an autoimmune-prone state to fatal autoimmune disease in CCR7 and RORγt double-deficient mice is dependent on gut microbiota.

Autoimmunity is associated with a strong genetic component, but onset and persistence of clinically apparent autoimmune diseases often require an additional environmental trigger. The balance between immunity and tolerance is regulated by numerous molecular factors including nuclear hormone and homeostatic chemokine receptors. The nuclear hormone receptor RORγt and the chemokine receptor CCR7 are both essentially involved in functional lymphoid organogenesis and maintenance of lymphocyte homeostasis. Lack of one or the other impairs thymic T cell development and alters T cell homeostasis. Mice deficient for both, Ccr7(-/-)Rorγt(-/-), succumbed early to acute destructive inflammation, characterized by massive recruitment of inflammatory leukocytes, pro-inflammatory cytokine and autoantibody production, and wasting disease. Antibiotic-treatment of mice before disease onset reduced the overall gut microflora and abrogated the development of fatal mucosal inflammation. Hence, commensal bacteria and a confined tissue-specific inflammatory milieu serve as complementary trigger to initiate the lethal pathophysiologic process in Ccr7(-/-)Rorγt(-/-) mice.

Role of the Aryl Hydrocarbon Receptor in Controlling Maintenance and Functional Programs of RORγt(+) Innate Lymphoid Cells and Intraepithelial Lymphocytes.

Mucosal retinoic receptor-related orphan receptor (ROR)γt-expressing innate lymphoid cells (ILC) play an important role in the defense against intestinal pathogens and in promoting epithelial homeostasis and adaptation, thereby effectively protecting the vertebrate host against intestinal inflammatory disorders. The functional activity of RORγt(+) ILC is under the control of environmental cues. However, the molecular sensors for such environmental signals are largely unknown. Recently, the aryl hydrocarbon receptor (AhR) has emerged as a master regulator for the postnatal maintenance of intestinal RORγt(+) ILC and intraepithelial lymphocytes. AhR is a highly conserved transcription factor whose activity is regulated by environmental and dietary small molecule ligands. Here, we review the role of AhR signaling for the maintenance of intestinal immune cells and its impact on the immunological protection against intestinal infections and debilitating chronic inflammatory disorders.

Isolated Lymphoid Follicles are Dynamic Reservoirs for the Induction of Intestinal IgA.

IgA is one of the most important molecules in the regulation of intestinal homeostasis. Peyer's patches have been traditionally recognized as sites for the induction of intestinal IgA responses, however more recent studies demonstrate that isolated lymphoid follicles (ILFs) can perform this function as well. ILF development is dynamic, changing in response to the luminal microbial burden, suggesting that ILFs play an important role providing an expandable reservoir of compensatory IgA inductive sites. However, in situations of immune dysfunction, ILFs can over-develop in response to uncontrollable enteric flora, resulting in ILF hyperplasia. The ability of ILFs to expand and respond to help control the enteric flora makes this dynamic reservoir an important arm of IgA inductive sites in intestinal immunity.