Highlights • Intestinal macrophages express CD64 and are constantly replenished by blood monocytes. • Intestinal macrophages are anti-inflammatory, sessile, and maintain local homeostasis. • ...Intestinal DCs derive from committed DC precursors in an flt3L-dependent manner. • Four subsets of intestinal DCs exist, identified by CD103 and CD11b expression. • DCs migrate to lymph nodes to prime naïve T cells. Subsets have discrete functions.
Heterogeneity between different macrophage populations has become a defining feature of this lineage. However, the conserved factors defining macrophages remain largely unknown. The transcription ...factor ZEB2 is best described for its role in epithelial to mesenchymal transition; however, its role within the immune system is only now being elucidated. We show here that Zeb2 expression is a conserved feature of macrophages. Using Clec4f-cre, Itgax-cre, and Fcgr1-cre mice to target five different macrophage populations, we found that loss of ZEB2 resulted in macrophage disappearance from the tissues, coupled with their subsequent replenishment from bone-marrow precursors in open niches. Mechanistically, we found that ZEB2 functioned to maintain the tissue-specific identities of macrophages. In Kupffer cells, ZEB2 achieved this by regulating expression of the transcription factor LXRα, removal of which recapitulated the loss of Kupffer cell identity and disappearance. Thus, ZEB2 expression is required in macrophages to preserve their tissue-specific identities.
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•ZEB2 is highly expressed across the macrophage lineage•ZEB2 preserves the tissue-specific identities of macrophages across tissues•ZEB2 deficient macrophages are outcompeted by WT counterparts•LXRα is crucial for Kupffer cell identity and is maintained by ZEB2
Scott et al. demonstrate that ZEB2 is critical for maintaining the tissue identities of macrophages. Loss of ZEB2 results in tissue-specific changes in different macrophage populations and their subsequent disappearance. In Kupffer cells, ZEB2 maintains LXRα expression, loss of which reproduces the change in Kupffer cell identity and their disappearance.
Macrophages in the healthy intestine are highly specialized and usually respond to the gut microbiota without provoking an inflammatory response. A breakdown in this tolerance leads to inflammatory ...bowel disease (IBD), but the mechanisms by which intestinal macrophages normally become conditioned to promote microbial tolerance are unclear. Strong epidemiological evidence linking disruption of the gut microbiota by antibiotic use early in life to IBD indicates an important role for the gut microbiota in modulating intestinal immunity. Here, we show that antibiotic use causes intestinal macrophages to become hyperresponsive to bacterial stimulation, producing excess inflammatory cytokines. Re-exposure of antibiotic-treated mice to conventional microbiota induced a long-term, macrophage-dependent increase in inflammatory T helper 1 (T
1) responses in the colon and sustained dysbiosis. The consequences of this dysregulated macrophage activity for T cell function were demonstrated by increased susceptibility to infections requiring T
17 and T
2 responses for clearance (bacterial
and helminth
infections), corresponding with increased inflammation. Short-chain fatty acids (SCFAs) were depleted during antibiotic administration; supplementation of antibiotics with the SCFA butyrate restored the characteristic hyporesponsiveness of intestinal macrophages and prevented T cell dysfunction. Butyrate altered the metabolic behavior of macrophages to increase oxidative phosphorylation and also promoted alternative macrophage activation. In summary, the gut microbiota is essential to maintain macrophage-dependent intestinal immune homeostasis, mediated by SCFA-dependent pathways. Oral antibiotics disrupt this process to promote sustained T cell-mediated dysfunction and increased susceptibility to infections, highlighting important implications of repeated broad-spectrum antibiotic use.
The murine serous cavities contain a rare and enigmatic population of short‐lived F4/80loMHCII+ macrophages but what regulates their development, survival, and fate is unclear. Here, we show that ...mature F4/80loMHCII+ peritoneal macrophages arise after birth, but that this occurs largely independently of colonization by microbiota. Rather, microbiota specifically regulate development of a subpopulation of CD11c+ cells that express the immunoregulatory cytokine RELM‐α, are reliant on the transcription factor EGR2, and develop independently of the growth factor CSF1. Furthermore, we demonstrate that intrinsic expression of RELM‐α, a signature marker shared by CD11c+ and CD11c– F4/80loMHCII+ cavity macrophages, regulates survival and differentiation of these cells in the peritoneal cavity in a sex‐specific manner. Thus, we identify a previously unappreciated diversity in serous cavity F4/80loMHCII+ macrophages that is regulated by microbiota, and describe a novel sex and site‐specific function for RELM‐α in regulating macrophage endurance that reveals the unique survival challenge presented to monocyte‐derived macrophages by the female peritoneal environment.
Monocyte‐derived antigen presenting cells in the peritoneal cavity are heterogenous, with two discrete populations distinguished by expression of CD11c, dependence upon microbiota, CSF1R, and the transcription factor EGR2, and distinct sex‐dependent contribution to resident CD102+ macrophages. These results help us understand the diverse factors regulating monocyte fate in serous cavities.
To understand the contribution of mononuclear phagocytes (MNP), which include monocyte-derived intestinal macrophages, to the pathogenesis of inflammatory bowel disease (IBD), it is necessary to ...identify functionally-different MNP populations. We aimed to characterise intestinal macrophage populations in patients with IBD. We developed 12-parameter flow cytometry protocols to identify and human intestinal MNPs. We used these protocols to purify and characterize colonic macrophages from colonic tissue from patients with Crohn's disease (CD), ulcerative colitis (UC), or non-inflamed controls, in a cross-sectional study. We identify macrophage populations (CD45
CD64
HLA-DR
) and describe two distinct subsets, differentiated by their expression of the mannose receptor, CD206. CD206
macrophages expressed markers consistent with a mature phenotype: high levels of CD68 and CD163, higher transcription of IL-10 and lower expression of TREM1. CD206
macrophages appear to be less mature, with features more similar to their monocytic precursors. We identified and purified macrophage populations from human colon. These appear to be derived from a monocytic precursor with high CCR2 and low CD206 expression. As these cells mature, they acquire expression of IL-10, CD206, CD63, and CD168. Targeting the newly recruited monocyte-derived cells may represent a fruitful avenue to ameliorate chronic inflammation in IBD.
Type 2 immunity is activated in response to both allergens and helminth infection. It can be detrimental or beneficial, and there is a pressing need to better understand its regulation. The ...immunosuppressive cytokine IL-10 is known as a T helper 2 (Th2) effector molecule, but it is currently unclear whether IL-10 dampens or promotes Th2 differentiation during infection. Here we show that helminth infection in mice elicits IL-10 expression in both the intestinal lamina propria and the draining mesenteric lymph node, with higher expression in the infected tissue. In vitro, exogenous IL-10 enhanced Th2 differentiation in isolated CD4
T cells, increasing expression of GATA3 and production of IL-5 and IL-13. The ability of IL-10 to amplify the Th2 response coincided with its suppression of IFNγ expression and in vivo we found that, in intestinal helminth infection, IL-10 receptor expression was higher on Th1 cells in the small intestine than on Th2 cells in the same tissue, or on any Th cell in the draining lymph node. In vivo blockade of IL-10 signalling during helminth infection resulted in an expansion of IFNγ
and Tbet
Th1 cells in the small intestine and a coincident decrease in IL-13, IL-5 and GATA3 expression by intestinal T cells. These changes in Th2 cytokines correlated with reduced expression of type 2 effector molecules, such as RELMα, and increased parasite egg production. Together our data indicate that IL-10 signalling promotes Th2 differentiation during helminth infection at least in part by regulating competing Th1 cells in the infected tissue.
T-helper 2 (Th2) cell responses defend against parasites. Although dendritic cells (DCs) are vital for the induction of T-cell responses, the DC subpopulations that induce Th2 cells in the intestine ...are unidentified. Here we show that intestinal Th2 responses against Trichuris muris worms and Schistosoma mansoni eggs do not develop in mice with IRF-4-deficient DCs (IRF-4
CD11c-cre). Adoptive transfer of conventional DCs, in particular CD11b-expressing DCs from the intestine, is sufficient to prime S. mansoni-specific Th2 responses. Surprisingly, transferred IRF-4-deficient DCs also effectively prime S. mansoni-specific Th2 responses. Egg antigens do not induce the expression of IRF-4-related genes. Instead, IRF-4
CD11c-cre mice have fewer CD11b
migrating DCs and fewer DCs carrying parasite antigens to the lymph nodes. Furthermore, CD11b
CD103
DCs induce Th2 responses in the small intestine, whereas CD11b
CD103
DCs perform this role in the colon, revealing a specific functional heterogeneity among intestinal DCs in inducing Th2 responses.
The use of bacteria as an alternative cancer therapy has been reinvestigated in recent years. SL7207: an auxotrophic Salmonella enterica serovar Typhimurium aroA mutant with immune‐stimulatory ...potential has proven a promising strain for this purpose. Here, we show that systemic administration of SL7207 induces melanoma tumor growth arrest in vivo, with greater survival of the SL7207‐treated group compared to control PBS‐treated mice. Administration of SL7207 is accompanied by a change in the immune phenotype of the tumor‐infiltrating cells toward pro‐inflammatory, with expression of the TH1 cytokines IFN‐γ, TNF‐α, and IL‐12 significantly increased. Interestingly, Ly6C+MHCII+ monocytes were recruited to the tumors following SL7207 treatment and were pro‐inflammatory. Accordingly, the abrogation of these infiltrating monocytes using clodronate liposomes prevented SL7207‐induced tumor growth inhibition. These data demonstrate a previously unappreciated role for infiltrating inflammatory monocytes underlying bacterial‐mediated tumor growth inhibition. This information highlights a possible novel role for monocytes in controlling tumor growth, contributing to our understanding of the immune responses required for successful immunotherapy of cancer.
Salmonella Typhimurium can target tumors and stall their growth in mouse models, a process associated with tumor infiltration by inflammatory monocytes. Clodronate liposomes, which abrogate phagocytic cells, inhibit Salmonella‐mediated tumor growth inhibition, suggesting a role for these cells in Salmonella’s antitumor properties.
FoxP3
regulatory T cells (Tregs) control inflammation and maintain mucosal homeostasis, but their functions during infection are poorly understood. Th1, Th2, and Th17 cells can be identified by ...master transcription factors (TFs) T-bet, GATA3, and RORγT; Tregs also express these TFs. While T-bet
Tregs can selectively suppress Th1 cells, it is unclear whether distinct Treg populations can alter Th bias. To address this, we used Salmonella enterica serotype Typhimurium to induce nonlethal colitis. Following infection, we observed an early colonic Th17 response within total CD4 T cells, followed by a Th1 bias. The early Th17 response, which contains both Salmonella-specific and non-Salmonella-specific cells, parallels an increase in T-bet
Tregs. Later, Th1 cells and RORγT
Tregs dominate. This reciprocal dynamic may indicate that Tregs selectively suppress Th cells, shaping the immune response. Treg depletion 1-2 days post-infection shifted the early Th17 response to a Th1 bias; however, Treg depletion 6-7 days post-infection abrogated the Th1 bias. Thus, Tregs are necessary for the early Th17 response, and for a maximal Th1 response later. These data show that Tregs shape the overall tissue CD4 T cell response and highlight the potential for subpopulations of Tregs to be used in targeted therapeutic approaches.
infection is a globally important cause of gastroenteritis and systemic disease and is a useful tool to study immune responses in the intestine. Although mechanisms leading to immune responses ...against
have been extensively studied, questions remain about how bacteria travel from the intestinal mucosa to the mesenteric lymph nodes (MLN), a key site for Ag presentation. In this study, we used a mouse model of infection with
serovar Typhimurium (STM) to identify changes in intestinal immune cells induced during early infection. We then used fluorescently labeled STM to identify interactions with immune cells from the site of infection through migration in lymph to the MLN. We show that viable STM can be carried in the lymph by any subset of migrating dendritic cells but not by macrophages. Moreover, approximately half of the STM in lymph are not associated with cells at all and travel autonomously. Within the MLN, STM associates with dendritic cells and B cells but predominantly with MLN-resident macrophages. In conclusion, we describe the routes used by STM to spread systemically in the period immediately postinfection. This deeper understanding of the infection process could open new avenues for controlling it.
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