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 (T_regs) 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 T_regs 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 T_reg 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 T_reg homeostasis locally and in mesenteric lymph nodes.

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 T_regs. 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.

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 T_reg 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 T_reg homeostasis in the large intestine. Conversely, administration …