Daigo Hashimoto

Tissue-resident macrophages self-maintain locally throughout adult life with minimal contribution from circulating monocytes.

Despite accumulating evidence suggesting local self-maintenance of tissue macrophages in the steady state, the dogma remains that tissue macrophages derive from monocytes. Using parabiosis and fate-mapping approaches, we confirmed that monocytes do not show significant contribution to tissue macrophages in the steady state. Similarly, we found that after depletion of lung macrophages, the majority of repopulation occurred by stochastic cellular proliferation inxa0situ in a macrophage colony-stimulating factor (M-Csf)- and granulocyte macrophage (GM)-CSF-dependent manner but independently of interleukin-4. We also found that after bone marrow transplantation, host macrophages retained the capacity to expand when the development of donor macrophages was compromised. Expansion of host macrophages was functional and prevented the development of alveolar proteinosis in mice transplanted with GM-Csf-receptor-deficientxa0progenitors. Collectively, these results indicate that tissue-resident macrophages and circulating monocytes should be classified as mononuclear phagocyte lineages that are independently maintained in the steady state.

Origin of the Lamina Propria Dendritic Cell Network

CX(3)CR1(+) and CD103(+) dendritic cells (DCs) in intestinal lamina propria play a key role in mucosal immunity. However, the origin and the developmental pathways that regulate their differentiation in the lamina propria remain unclear. We showed that monocytes gave rise exclusively to CD103(-)CX(3)CR1(+) lamina propria DCs under the control of macrophage-colony-stimulating factor receptor (M-CSFR) and Fms-like thyrosine kinase 3 (Flt3) ligands. In contrast, common DC progenitors (CDP) and pre-DCs, which give rise to lymphoid organ DCs but not to monocytes, differentiated exclusively into CD103(+)CX(3)CR1(-) lamina propria DCs under the control of Flt3 and granulocyte-macrophage-colony-stimulating factor receptor (GM-CSFR) ligands. CD103(+)CX(3)CR1(-) DCs but not CD103(-)CX(3)CR1(+) DCs in the lamina propria constitutively expressed CCR7 and were the first DCs to transport pathogenic Salmonella from the intestinal tract to the mesenteric lymph nodes. Altogether, these results underline the diverse origin of the lamina propria DC network and identify mucosal DCs that arise from pre-DCs as key sentinels of the gut immune system.

The origin and development of nonlymphoid tissue CD103+ DCs

CD103+ dendritic cells (DCs) in nonlymphoid tissues are specialized in the cross-presentation of cell-associated antigens. However, little is known about the mechanisms that regulate the development of these cells. We show that two populations of CD11c+MHCII+ cells separated on the basis of CD103 and CD11b expression coexist in most nonlymphoid tissues with the exception of the lamina propria. CD103+ DCs are related to lymphoid organ CD8+ DCs in that they are derived exclusively from pre-DCs under the control of fms-like tyrosine kinase 3 (Flt3) ligand, inhibitor of DNA protein 2 (Id2), and IFN regulatory protein 8 (IRF8). In contrast, lamina propria CD103+ DCs express CD11b and develop independently of Id2 and IRF8. The other population of CD11c+MHCII+ cells in tissues, which is CD103−CD11b+, is heterogenous and depends on both Flt3 and MCSF-R. Our results reveal that nonlymphoid tissue CD103+ DCs and lymphoid organ CD8+ DCs derive from the same precursor and follow a related differentiation program.

Bone marrow CD169 + macrophages promote the retention of hematopoietic stem and progenitor cells in the mesenchymal stem cell niche

Hematopoietic stem cells (HSCs) reside in specialized bone marrow (BM) niches regulated by the sympathetic nervous system (SNS). Here, we have examined whether mononuclear phagocytes modulate the HSC niche. We defined three populations of BM mononuclear phagocytes that include Gr-1hi monocytes (MOs), Gr-1lo MOs, and macrophages (MΦ) based on differential expression of Gr-1, CD115, F4/80, and CD169. Using MO and MΦ conditional depletion models, we found that reductions in BM mononuclear phagocytes led to reduced BM CXCL12 levels, the selective down-regulation of HSC retention genes in Nestin+ niche cells, and egress of HSCs/progenitors to the bloodstream. Furthermore, specific depletion of CD169+ MΦ, which spares BM MOs, was sufficient to induce HSC/progenitor egress. MΦ depletion also enhanced mobilization induced by a CXCR4 antagonist or granulocyte colony-stimulating factor. These results highlight two antagonistic, tightly balanced pathways that regulate maintenance of HSCs/progenitors in the niche during homeostasis, in which MΦ cross talk with the Nestin+ niche cell promotes retention, and in contrast, SNS signals enhance egress. Thus, strategies that target BM MΦ hold the potential to augment stem cell yields in patients that mobilize HSCs/progenitors poorly.

Deciphering the transcriptional network of the dendritic cell lineage

Although much progress has been made in the understanding of the ontogeny and function of dendritic cells (DCs), the transcriptional regulation of the lineage commitment and functional specialization of DCs in vivo remains poorly understood. We made a comprehensive comparative analysis of CD8+, CD103+, CD11b+ and plasmacytoid DC subsets, as well as macrophage DC precursors and common DC precursors, across the entire immune system. Here we characterized candidate transcriptional activators involved in the commitment of myeloid progenitor cells to the DC lineage and predicted regulators of DC functional diversity in tissues. We identified a molecular signature that distinguished tissue DCs from macrophages. We also identified a transcriptional program expressed specifically during the steady-state migration of tissue DCs to the draining lymph nodes that may control tolerance to self tissue antigens.

Microbiota-Dependent Crosstalk Between Macrophages and ILC3 Promotes Intestinal Homeostasis

Introduction The gastrointestinal tract is colonized by an extraordinarily large number of commensal microbes and is constantly exposed to ingested antigens and potential pathogens. Regulation of intestinal tolerance thus represents the main task of the immune system of the gut mucosa. Accumulated evidence suggests that gut commensals contribute to the maintenance of intestinal homeostasis, partly through their ability to control the differentiation of effector T lymphocytes in the mucosa and to modulate inflammatory responses through the induction of regulatory T cells (Tregs) and interleukin-10 (IL-10) production. Tissue-resident mononuclear phagocytes (MNPs), including macrophages (MPs) and dendritic cells (DCs), are specifically equipped to detect a wide range of microbial signals and to capture, process, and present extracellular antigenic material to T lymphocytes. MNPs have been shown to contribute to the maintenance of intestinal immune tolerance through the induction or expansion of Tregs in the intestine. Despite their key role in microbial sensing and immune tolerance, the cellular and molecular cues that translate microbial signals into immunoregulatory MNPs in the intestine are not completely understood. ILC3 translate microbial cues into immunoregulatory signals in the intestine. Microbial cues sensed by intestinal MPs lead to IL-1β release. IL-1β engages IL-1R on ILC3, promoting Csf2 release. ILC3-derived Csf2 triggers DC and MP production of regulatory molecules (i.e., RA and IL-10), which, in turn, promotes the induction and expansion of regulatory T cells. Csf2-primed DCs and MPs promote Treg homeostasis locally and in mesenteric lymph nodes. ILC3 translate microbial cues into immunoregulatory signals in the intestine. Microbial cues sensed by intestinal MPs lead to IL-1β release. IL-1β engages IL-1R on ILC3, promoting Csf2 release. ILC3-derived Csf2 triggers DC and MP production of regulatory molecules (i.e., RA and IL-10), which, in turn, promotes the induction and expansion of regulatory T cells. Csf2-primed DCs and MPs promote Treg homeostasis locally and in mesenteric lymph nodes. Rationale The cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF), recently renamed Csf2, is a key determinant of myeloid lineage differentiation and is required for the optimal function of tissue MNPs. Recent results from our laboratory revealed that although Csf2-deficient mice have normal numbers of lymphoid tissue-resident DCs, they display significantly reduced numbers of steady-state nonlymphoid tissue-resident DCs in the small intestine, including the lamina propria CD103+CD11b+ DC subset implicated in the induction of lamina propria Tregs. These results prompted us to further explore the contribution of Csf2 to intestinal immune homeostasis in vivo. We used detailed profiling studies and functional immune assays of the MNP and lymphocyte compartment in the gut, as well as genetically engineered mice that lack Csf2 or the transducer Myd88 specifically in MNPs or lymphocytes, to explore the role of MNPs in the maintenance of immune homeostasis in the gut. Results Our results revealed a crosstalk between IL-1β–secreting MPs and Csf2-producing RORγt+ type 3 innate lymphoid cells (ILC3) in the intestinal mucosa. Microbiota-driven IL-1β production by MPs promoted the release of Csf2 by ILC3, which in turn controlled DCs and MPs to maintain colonic Treg homeostasis. Ablation of Csf2 reduced DC and MP numbers and impaired their ability to produce regulatory factors such as retinoic acid (RA) and IL-10, leading to disrupted Treg homeostasis in the large intestine. Conversely, administration of Csf2 cytokine increased Treg frequency in the gut. Most notably, cell type–specific ablation of IL-1 receptor (IL-1R)–dependent signaling in RORγt+ ILC3 abrogated oral tolerance to dietary antigens and compromised intestinal immune homeostasis in vivo. Although the reduction in Treg numbers was mostly observed in the large intestine, adoptive transfer studies in Csf2–/– mice revealed impaired Treg differentiation both in the small and large intestine, suggesting that Csf2-dependent MNP immunoregulatory functions control Treg induction in both tissues. Conclusion This study established the commensal-driven MNP-ILC-Csf2 axis as a key regulator of intestinal T cell homeostasis in the mouse intestine. Disturbance of this axis radically altered MNP effector function, resulting in impaired oral tolerance to dietary antigens. These results represent an important advance in our understanding of how commensal microbes can regulate host intestinal immunity and may inform the design of novel immunotherapies for patients with inflammatory intestinal diseases with impaired GM-CSF function. Gut Immune Tolerance With the constant assault of food antigens and its billions of resident microbes, the gut is an important site of immune tolerance. By studying specific intestinal immune cell populations in genetically modified mice, Mortha et al. (10.1126/science.1249288, published online 13 March; see the Perspective by Aychek and Jung) found that gut macrophages produce the cytokine interleukin-1 (IL-1) in response to signals derived from the microbiota. IL-1 acts on type 3 innate lymphoid cells in the intestine, which then produce the cytokine, colony-stimulating factor 2 (Csf2). Csf-2, in turn, induces myeloid cells (including dendritic cells and macrophages) to produce regulatory factors like retinoic acid and interleukin-10, which support the conversion and expansion of regulatory T cells, a population of cells known to be critical for maintaining immune tolerance in the gut. Myeloid cells, innate lymphoid cells, and the cytokines they secrete cooperate to maintain immune tolerance in the gut. [Also see Perspective by Aychek and Jung] The intestinal microbiota and tissue-resident myeloid cells promote immune responses that maintain intestinal homeostasis in the host. However, the cellular cues that translate microbial signals into intestinal homeostasis remain unclear. Here, we show that deficient granulocyte-macrophage colony-stimulating factor (GM-CSF) production altered mononuclear phagocyte effector functions and led to reduced regulatory T cell (Treg) numbers and impaired oral tolerance. We observed that RORγt+ innate lymphoid cells (ILCs) are the primary source of GM-CSF in the gut and that ILC-driven GM-CSF production was dependent on the ability of macrophages to sense microbial signals and produce interleukin-1β. Our findings reveal that commensal microbes promote a crosstalk between innate myeloid and lymphoid cells that leads to immune homeostasis in the intestine.

Dendritic cell and macrophage heterogeneity in vivo.

Macrophage and dendritic cell (DC) are hematopoietic cells found in all tissues in the steady state that share the ability to sample the environment but have distinct function in tissue immunity. Controversies remain on the best way to distinguish macrophages from DCs inxa0vivo. In this Perspective, we discuss how recent discoveries in the origin of the DC and macrophage lineage help establish key functional differences between tissue DC and macrophage subsets. We also emphasize the need to further understand the functional heterogeneity of the tissue DC and macrophage lineages to better comprehend the complex role of these cells in tissue homeostasis and immunity.

GM-CSF controls nonlymphoid tissue dendritic cell homeostasis but is dispensable for the differentiation of inflammatory dendritic cells.

GM-CSF (Csf-2) is a critical cytokine for the inxa0vitro generation of dendritic cells (DCs) and is thought to control the development of inflammatory DCs and resident CD103(+) DCs in some tissues. Here we showed that in contrast to the current understanding, Csf-2 receptor acts in the steady state to promote the survival and homeostasis of nonlymphoid tissue-resident CD103(+) and CD11b(+) DCs. Absence of Csf-2 receptor on lung DCs abrogated the induction of CD8(+) Txa0cell immunity after immunization with particulate antigens. In contrast, Csf-2 receptor was dispensable for the differentiation and innate function of inflammatory DCs during acute injuries. Instead, inflammatory DCs required Csf-1 receptor for their development. Thus, Csf-2 is important in vaccine-induced CD8(+) Txa0cell immunity through the regulation of nonlymphoid tissue DC homeostasis rather than control of inflammatory DCs inxa0vivo.

CD169+ macrophages provide a niche promoting erythropoiesis under homeostasis and stress

The role of macrophages in erythropoiesis was suggested several decades ago with the description of “erythroblastic islands” in the bone marrow (BM) composed of a central macrophage surrounded by developing erythroblasts. However, the in vivo role of macrophages in erythropoiesis under homeostasis or disease remains unclear. Specific depletion of CD169+ macrophages markedly reduced erythroblasts in the BM but did not result in overt anemia under homeostasis likely due to concomitant alterations in RBC clearance. However, CD169+ macrophage depletion significantly impaired erythropoietic recovery from hemolytic anemia, acute blood loss and myeloablation. Furthermore, macrophage depletion normalized the erythroid compartment in a JAK2V617F-driven murine model of polycythemia vera (PV), suggesting that erythropoiesis in PV, unexpectedly, remains under the control of macrophages in the BM and splenic microenvironments. These data indicate that CD169+ macrophages promote late erythroid maturation and that modulation of the macrophage compartment represents a novel strategy to treat erythropoietic disorders.A role for macrophages in erythropoiesis was suggested several decades ago when erythroblastic islands in the bone marrow, composed of a central macrophage surrounded by developing erythroblasts, were described. However, the in vivo role of macrophages in erythropoiesis under homeostatic conditions or in disease remains unclear. We found that specific depletion of CD169+ macrophages markedly reduced the number of erythroblasts in the bone marrow but did not result in overt anemia under homeostatic conditions, probably because of concomitant alterations in red blood cell clearance. However, CD169+ macrophage depletion significantly impaired erythropoietic recovery from hemolytic anemia, acute blood loss and myeloablation. Furthermore, macrophage depletion normalized the erythroid compartment in a JAK2V617F-driven mouse model of polycythemia vera, suggesting that erythropoiesis in polycythemia vera remains under the control of macrophages in the bone marrow and splenic microenvironments. These results indicate that CD169+ macrophages promote late erythroid maturation and that modulation of the macrophage compartment may be a new strategy to treat erythropoietic disorders.

Crosstalk between Muscularis Macrophages and Enteric Neurons Regulates Gastrointestinal Motility

Intestinal peristalsis is a dynamic physiologic process influenced by dietary and microbial changes. It is tightly regulated by complex cellular interactions; however, our understanding of these controls is incomplete. A distinct population of macrophages is distributed in the intestinal muscularis externa. We demonstrate that, in the steady state, muscularis macrophages regulate peristaltic activity of the colon. They change the pattern of smooth muscle contractions by secreting bone morphogenetic protein 2 (BMP2), which activates BMP receptor (BMPR) expressed by enteric neurons. Enteric neurons, in turn, secrete colony stimulatory factor 1 (CSF1), a growth factor required for macrophage development. Finally, stimuli from microbial commensals regulate BMP2 expression by macrophages and CSF1 expression by enteric neurons. Our findings identify a plastic, microbiota-driven crosstalk between muscularis macrophages and enteric neurons that controls gastrointestinal motility. PAPERFLICK: