Project description:In the present study, we demonstrate that NOD2 receptor triggering by muramyl dipeptide converts blood inflammatory Ly6Chigh monocytes into patrolling Ly6Clow monocytes. Muramyl dipeptide administration to Nr4a1-/- mice, which lack Ly6Clow monocytes, and to Ly6Clow-depleted mice leads to the emergence of blood patrolling monocytes expressing phenotypic profile of typical Ly6Clow monocytes, including high expression of CX3CR1 and LFA1. Furthermore using intravital microscopy in live animal models of inflammatory diseases, we observed that converted Ly6Chigh monocytes are capable of patrolling the endothelium of blood vessels and their presence contributes to reduce the inflammatory response following muramyl dipeptide injection.
Project description:Monocytes are central for atherosclerotic vascular inflammation. The human non-classical, patrolling subtype, which expresses high levels of CD16 and fractalkine receptor CX3CR1, strongly associates with cardiovascular events. Compared to classical CD14+ monocytes or transwell cocultures, CD16+ monocytes enhanced endothelial STAT1 and NFκB p65 phosphorylation, upregulated expression of CX3CL1 and IL-1β, numerous CCL and CXCL chemokines and molecules promoting leukocyte patrolling and adhesion such as ICAM1 and VCAM1.
Project description:Mononuclear phagocytes are a diverse cell family that occupy all tissues and assume numerous functions to support tissue and systemic homeostasis. Our ability to investigate the roles of individual subsets is limited by an absence of approaches to ablate gene function within specific sub-populations. Using Nr4a1-dependent Ly6Clow monocytes as a representative cell type we show that enhancer deletion addresses these limitations. Combining ChIP-Seq and molecular approaches we identify a single, conserved, sub-domain within the Nr4a1 enhancer that is essential for Ly6Clow monocyte development. Mice lacking this enhancer lack Ly6Clow monocytes but retain Nr4a1 gene expression in macrophages during steady state and in response to LPS. Nr4a1 is a key negative regulator of inflammatory gene expression and decoupling these processes allows Ly6Clow monocytes to be studied without confounding influences. Enhancer targeting possesses greater specificity than cre recombinase-mediated gene deletion, providing a route to generate loss-of-function models in closely related cell types.
Project description:Mononuclear phagocytes are a diverse cell family that occupy all tissues and assume numerous functions to support tissue and systemic homeostasis. Our ability to investigate the roles of individual subsets is limited by an absence of approaches to ablate gene function within specific sub-populations. Using Nr4a1-dependent Ly6Clow monocytes as a representative cell type we show that enhancer deletion addresses these limitations. Combining ChIP-Seq and molecular approaches we identify a single, conserved, sub-domain within the Nr4a1 enhancer that is essential for Ly6Clow monocyte development. Mice lacking this enhancer lack Ly6Clow monocytes but retain Nr4a1 gene expression in macrophages during steady state and in response to LPS. Nr4a1 is a key negative regulator of inflammatory gene expression and decoupling these processes allows Ly6Clow monocytes to be studied without confounding influences. Enhancer targeting possesses greater specificity than cre recombinase-mediated gene deletion, providing a route to generate loss-of-function models in closely related cell types.
Project description:In rare instances, pediatric SARS-CoV2 infection results in a novel immunodysregulation syndrome termed multisystem inflammatory syndrome in children (MIS-C). We compared MIS-C immunopathology with that of severe COVID-19. MIS-C does not result in pneumocyte damage but is associated with vascular endothelitis, gastrointestinal epithelial injury and endotoxemia. In MIS-C, IFN dominates the inflammatory signature in contrast to type I interferons in severe COVID-19. While HLA-DRlo classical monocytes dominate severe COVID-19, patrolling monocyte activation characterize MIS-C. TIM-3 expression on activated CD4/CD8 T cells and on CD57+ NK cells is a distinctive MIS-C feature. In all MIS-C patients, single cell TCR profiling demonstrates a skewed TCR repertoire enriched for TRBV11-2 and a superantigenic signature which is absent in severe COVID-19. Using NicheNet, we confirm IFN as a central cytokine in the communication between activated T cells, NK cells and patrolling monocytes. Rapid normalization of IFN, TIM-3 loss on T cells and patrolling monocytes contraction upon treatment highlight their potential role in MIS-C immunopathogenesis.
Project description:We sought to evaluate in an unbiased way the heterogeneity of lung interstitial macrophages and their relationship with alveolar macrophages, lung Ly-6Chi classical monocytes and Ly-6Clo patrolling monocytes, by single cell RNA-Seq.
Project description:Extracellular vesicles (EVs) secreted by tumor cells are able to establish a pre-metastatic niche in distant organs, or on the contrary, exert anti-tumor activity. The mechanisms directing distinct EV functions are unknown. Using the B-16V transplantation mouse melanoma model we demonstrate that EVs from B-16V cells mobilize Ly6Clow patrolling monocytes and inhibit lung metastasis. Mechanistically, the formation of anti-tumor-EVs was dependent on the chaperone BAG6 and the acetylation of p53 by the BAG6/CBP/p300-acetylase complex, followed by the recruitment of components of the endosomal sorting complexes required for transport (ESCRT) via a P(S/T)AP double motif of BAG6. By contrast, deficiency of BAG6 led to the release of a distinct vesicle subtype with pro-tumorigenic activity, which recruited neutrophils to the pre-metastatic niche. In humans, BAG6 expression decreases in late-stage melanoma patients, correlating with an increase of the mRNA for the metastasis driver alpha-catulin in EVs, as observed in BAG6-deficient mouse EVs. We conclude that the BAG6/CBP/p300-p53 axis is a key pathway directing EV-formation and function.
Project description:Targeted gene expression on Lewis lung carcinoma (LLC) sorted MHC-IIlow macrophages (M2), Ly6Chigh inflammatory (IM) and Ly6Clow residential (RM) monocytes to investigate the effect of anti-PD-L1 mAb on specific myeloid subsets in the lung tumor microenvironment performed using the nCounter Myeloid Innate Immunity Panel by Nanostring .
