Emma Slack

Toll-like receptor expression in murine DC subsets: lack of TLR7 expression by CD8α+ DC correlates with unresponsiveness to imidazoquinolines

Toll‐like receptors (TLR) recognize microbial and viral patterns and activate dendritic cells (DC). TLR distribution among human DC subsets is heterogeneous: plasmacytoid DC (PDC) express TLR1, 7 and 9, while other DC types do not express TLR9 but express other TLR. Here, we report that mRNA for most TLR is expressed at similar levels by murine splenic DC sub‐types, including PDC, but that TLR3 is preferentially expressed by CD8α+ DC while TLR5 and TLR7 are selectively absent from the same subset. Consistent with the latter, TLR7 ligand activates CD8α– DC and PDC, but not CD8α+ DC as measured by survival ex vivo, up‐regulation of surface markers and production of IL‐12p40. These data suggest that the dichotomy in TLR expression between plasmacytoid and non‐plasmacytoid DC is not conserved between species. However, lack of TLR7 expression could restrict the involvement of CD8α+ DC in recognition of certain mouse pathogens.

Intestinal Bacterial Colonization Induces Mutualistic Regulatory T Cell Responses

Mammals harbor a dense commensal microbiota in the colon. Regulatory T (Treg) cells are known to limit microbe-triggered intestinal inflammation and the CD4+ T cell compartment is shaped by the presence of particular microbes or bacterial compounds. It is, however, difficult to distinguish whether these effects reflect true mutualistic immune adaptation to intestinal colonization or rather idiosyncratic immune responses. To investigate truly mutualistic CD4+ T cell adaptation, we used the altered Schaedler flora (ASF). Intestinal colonization resulted in activation and de novo generation of colonic Treg cells. Failure to activate Treg cells resulted in the induction of T helper 17 (Th17) and Th1 cell responses, which was reversed by wild-type Treg cells. Efficient Treg cell induction was also required to maintain intestinal homeostasis upon dextran sulfate sodium-mediated damage in the colon. Thus, microbiota colonization-induced Treg cell responses are a fundamental intrinsic mechanism to induce and maintain host-intestinal microbial T cell mutualism.

Myeloid C-type lectins in innate immunity

C-type lectins expressed on myeloid cells comprise a family of proteins that share a common structural motif, and some act as receptors in pathogen recognition. But just as the presence of leucine-rich repeats alone is not sufficient to define a Toll-like receptor, the characterization of C-type lectin receptors in innate immunity requires the identification of accompanying signaling motifs. Here we focus on the known signaling pathways of myeloid C-type lectins and on their possible functions as autonomous activating or inhibitory receptors involved in innate responses to pathogens or self.

Reversible microbial colonization of germ-free mice reveals the dynamics of IgA immune responses.

A Gut Feeling The mammalian gut is colonized by many nonpathogenic, commensal microbes. In order to prevent the body from mounting inappropriate immune responses to these microbes, plasma cells in the gut produce large amounts of immunoglobulin A (IgA) specific for commensal bacteria. Because of the difficulties of uncoupling IgA production from microbial colonization, how commensal bacteria shape the gut IgA response is not well understood. Hapfelmeier et al. (p. 1705; see the Perspective by Cerutti) have now devised a way to get around this problem by developing a reversible system of gut bacterial colonization in mice. Commensal-specific IgA responses were able to persist for long periods of time in the absence of microbial colonization and required the presence of high microbial loads in the gut for their induction. IgA responses upon bacterial reexposure did not resemble the synergistic prime-boost effect seen in classical immunological memory responses but rather exhibited an additive effect that matched the current bacterial content present in the gut. The body thus constantly adapts the commensal-specific immune response to the microbial species present in the gut, which contrasts with the systemic immune response, which persists in the absence of pathogenic microbes. Immunoglobulin responses against nonpathogenic bacteria in the gut are specific for the resident microbial flora. The lower intestine of adult mammals is densely colonized with nonpathogenic (commensal) microbes. Gut bacteria induce protective immune responses, which ensure host-microbial mutualism. The continuous presence of commensal intestinal bacteria has made it difficult to study mucosal immune dynamics. Here, we report a reversible germ-free colonization system in mice that is independent of diet or antibiotic manipulation. A slow (more than 14 days) onset of a long-lived (half-life over 16 weeks), highly specific anticommensal immunoglobulin A (IgA) response in germ-free mice was observed. Ongoing commensal exposure in colonized mice rapidly abrogated this response. Sequential doses lacked a classical prime-boost effect seen in systemic vaccination, but specific IgA induction occurred as a stepwise response to current bacterial exposure, such that the antibody repertoire matched the existing commensal content.

Innate and Adaptive Immunity Cooperate Flexibly to Maintain Host-Microbiota Mutualism

Maintaining Mutual Ignorance Our gut is colonized by trillions of bacteria that do not activate the immune system because of careful compartmentalization. Such compartmentalization means that our immune system is “ignorant” of these microbes and thus it has been proposed that loss of compartmentalization might result in an immune response to the colonizing bacteria. Microorganisms are sensed by cells that express pattern recognition receptors, such as Toll-like receptors, which recognize patterns specific to those microbes. Slack et al. (p. 617) show that Toll-like receptor–dependent signaling is required to maintain compartmentalization of bacteria to the gut of mice. In the absence of Toll-dependent signaling, intestinal bacteria disseminated throughout the body and the mice mounted a high-titer antibody response against them. This antibody response was of great functional importance because, despite the loss of systemic ignorance to intestinal microbes, the mice were tolerant of the bacteria. Thus, in the absence of innate immunity, the adaptive immune system can compensate so that host and bacterial mutualism can be maintained. Mouse immune systems interact to ensure tolerance to nonpathogenic bacteria in the gut. Commensal bacteria in the lower intestine of mammals are 10 times as numerous as the body’s cells. We investigated the relative importance of different immune mechanisms in limiting the spread of the intestinal microbiota. Here, we reveal a flexible continuum between innate and adaptive immune function in containing commensal microbes. Mice deficient in critical innate immune functions such as Toll-like receptor signaling or oxidative burst production spontaneously produce high-titer serum antibodies against their commensal microbiota. These antibody responses are functionally essential to maintain host-commensal mutualism in vivo in the face of innate immune deficiency. Spontaneous hyper-activation of adaptive immunity against the intestinal microbiota, secondary to innate immune deficiency, may clarify the underlying mechanisms of inflammatory diseases where immune dysfunction is implicated.

The Microbiota Mediates Pathogen Clearance from the Gut Lumen after Non-Typhoidal Salmonella Diarrhea

Many enteropathogenic bacteria target the mammalian gut. The mechanisms protecting the host from infection are poorly understood. We have studied the protective functions of secretory antibodies (sIgA) and the microbiota, using a mouse model for S. typhimurium diarrhea. This pathogen is a common cause of diarrhea in humans world-wide. S. typhimurium (S. tm att, sseD) causes a self-limiting gut infection in streptomycin-treated mice. After 40 days, all animals had overcome the disease, developed a sIgA response, and most had cleared the pathogen from the gut lumen. sIgA limited pathogen access to the mucosal surface and protected from gut inflammation in challenge infections. This protection was O-antigen specific, as demonstrated with pathogens lacking the S. typhimurium O-antigen (wbaP, S. enteritidis) and sIgA-deficient mice (TCRβ−/−δ−/−, JH −/−, IgA−/−, pIgR−/−). Surprisingly, sIgA-deficiency did not affect the kinetics of pathogen clearance from the gut lumen. Instead, this was mediated by the microbiota. This was confirmed using ‘L-mice’ which harbor a low complexity gut flora, lack colonization resistance and develop a normal sIgA response, but fail to clear S. tm att from the gut lumen. In these mice, pathogen clearance was achieved by transferring a normal complex microbiota. Thus, besides colonization resistance ( = pathogen blockage by an intact microbiota), the microbiota mediates a second, novel protective function, i.e. pathogen clearance. Here, the normal microbiota re-grows from a state of depletion and disturbed composition and gradually clears even very high pathogen loads from the gut lumen, a site inaccessible to most “classical” immune effector mechanisms. In conclusion, sIgA and microbiota serve complementary protective functions. The microbiota confers colonization resistance and mediates pathogen clearance in primary infections, while sIgA protects from disease if the host re-encounters the same pathogen. This has implications for curing S. typhimurium diarrhea and for preventing transmission.

The Liver May Act as a Firewall Mediating Mutualism Between the Host and Its Gut Commensal Microbiota

The liver forms a firewall that protects against vascular-borne gut microbes and is commonly impaired in liver disease. Breaching Barriers Premature death from chronic liver disease is a rising global trend. Opportunistic bacterial infections caused by beneficial microbes that have breached the gut and its immune barrier often lead to death in liver cirrhosis patients. Balmer et al. now show that the liver forms a second vascular barrier for eliminating commensal bacteria that have escaped from the gut. In animal models of liver disease and gut dysfunction and in patients with nonalcoholic steatohepatitis, the liver is unable to capture escaped gut commensal bacteria, which then leak into the systemic circulation, resulting in a robust host nonmucosal immune response and the breakdown of mutualism between the host and its gut microbiota. Mutualism breakdown is an important complication of liver disease. A prerequisite for establishment of mutualism between the host and the microbial community that inhabits the large intestine is the stringent mucosal compartmentalization of microorganisms. Microbe-loaded dendritic cells trafficking through lymphatics are arrested at the mesenteric lymph nodes, which constitute the firewall of the intestinal lymphatic circulation. We show in different mouse models that the liver, which receives the intestinal venous blood circulation, forms a vascular firewall that captures gut commensal bacteria entering the bloodstream during intestinal pathology. Phagocytic Kupffer cells in the liver of mice clear commensals from the systemic vasculature independently of the spleen through the liver’s own arterial supply. Damage to the liver firewall in mice impairs functional clearance of commensals from blood, despite heightened innate immunity, resulting in spontaneous priming of nonmucosal immune responses through increased systemic exposure to gut commensals. Systemic immune responses consistent with increased extraintestinal commensal exposure were found in humans with liver disease (nonalcoholic steatohepatitis). The liver may act as a functional vascular firewall that clears commensals that have penetrated either intestinal or systemic vascular circuits.

Syk-dependent ERK activation regulates IL-2 and IL-10 production by DC stimulated with zymosan

Zymosan is a particulate yeast preparation that elicits high levels of IL‐2 and IL‐10 from dendritic cells (DC) and engages multiple innate receptors, including the Syk‐coupled receptor dectin‐1 and the MyD88‐coupled receptor TLR2. Here, we show that induction of IL‐2 and IL‐10 by zymosan requires activation of ERK MAP kinase in murine DC. Surprisingly, ERK activation in response to zymosan is completely blocked in Syk‐deficient DC and unaffected by MyD88 deficiency. Conversely, ERK activation in response to the TLR2 agonist Pam3Cys is completely MyD88 dependent and unaffected by Syk deficiency. The inability of TLR2 ligands in zymosan to couple to ERK may explain the Syk dependence of the IL‐2 and IL‐10 response in DC and emphasises the importance of Syk‐coupled pattern recognition receptors such as dectin‐1 in the detection of yeasts. Furthermore, the lack of receptor compensation observed here suggests that responses induced by complex innate stimuli cannot always be predicted by the signalling pathways downstream of individual receptors.

The functional interactions of commensal bacteria with intestinal secretory IgA.

Purpose of review The aim of this article is to describe the immune geography of IgA induction by commensal intestinal bacteria and the underlying mechanisms of cytokine and costimulatory signalling between dendritic cells, B cells and T cells. Recent findings Intestinal dendritic cells sample commensal intestinal bacteria that penetrate the epithelial layer and induce IgA+ B cells to seed the mucosa with IgA plasma cells. Constitutive secretion of retinoic acid by intestinal dendritic cells directs the specificity of the IgA class switch and homing receptor expression in Peyers patch B cells. In-vivo experiments have shown that TGF-β is a vital cytokine for IgA induction in vivo, and the tumour necrosis factor family members BAFF and APRIL provide key costimulatory signals. After transport through the epithelial layer secretory IgA limits penetration of commensal bacteria back through the epithelium and shapes the density of different bacterial species in the intestinal lumen. Summary Production of IgA is an important adaptation to the presence of commensal intestinal bacteria and induction of the response is compartmentalized within the intestinal mucosal immune system. This compartmentalization allows a vigorous mucosal immune response to commensals without needing the systemic immune system to be tolerant of these organisms.

The habitat, double life, citizenship, and forgetfulness of IgA

Summary:  Immunoglobulin A (IgA) is the main secretory immunoglobulin of mucous membranes and is powerfully induced by the presence of commensal microbes in the intestine. B cells undergo class switch recombination to IgA in the mucosa‐associated lymphoid tissues, particularly mesenteric lymph nodes (MLNs) and Peyer’s patches, through both T‐dependent and T‐independent pathways. IgA B cells primed in the mucosa traffic from the intestinal lymphoid structures, initially through the lymphatics and then join the bloodstream, to home back to the intestinal mucosa as IgA‐secreting plasma cells. Once induced, anti‐bacterial IgA can be extremely long‐lived but is replaced if there is induction of additional IgA specificities by other microbes. The mucosal immune system is anatomically separated from the systemic immune system by the MLNs, which act as a firewall to prevent penetration of live intestinal bacteria to systemic sites. Dendritic cells sample intestinal bacteria and induce B cells to switch to IgA. In contrast, intestinal macrophages are adept at killing extracellular bacteria and are able to clear bacteria that have crossed the mucus and epithelial barriers. There is both a continuum between innate and adaptive immune mechanisms and compartmentalization of the mucosal immune system from systemic immunity that function to preserve host microbial mutualism.