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Phagocytes, including neutrophils and macrophages, have been suggested to function in a cooperative way in the initial phase of inflammatory responses, but their interaction and integration in the resolution of inflammation and tissue repair remain unclear. Here we show that neutrophils have crucial functions in liver repair by promoting the phenotypic conversion of pro-inflammatory Ly6ChiCX3CR1lo monocytes/macrophages to pro-resolving Ly6CloCX3CR1hi macrophages. Intriguingly, reactive oxygen species (ROS), expressed predominantly by neutrophils, are important mediators that trigger this phenotypic conversion to promote liver repair. Moreover, this conversion is prevented by the depletion of neutrophils via anti-Ly6G antibody, genetic deficiency of granulocyte colony-stimulating factor, or genetic deficiency of NADPH oxidase 2 (Nox2). By contrast, adoptive transfer of WT rather than Nox2−/− neutrophils rescues the impaired phenotypic conversion of macrophages in neutrophil-depleted mice. Our findings thus identify an intricate cooperation between neutrophils and macrophages that orchestrate resolution of inflammation and tissue repair., Neutrophils and macrophages are both involved in the initiation of inflammation, but whether and how they may participate in inflammation resolution is unclear. Here the authors show that neutrophils may mediate the conversion of macrophage into a pro-resolution phenotype via reactive oxygen species production to promote liver repair.

Background & Aims Production of neutralizing antibodies against hepatitis B surface antigen (HBsAg) is dysregulated in patients with persistent hepatitis B virus (HBV) infection. We investigated mechanisms by which this immune response to the virus is disrupted and whether it can be restored to promote clearance of HBV. Methods Immune-competent C57BL/6N and C57BL/6J, as well as mice deficient in follicular helper T cells (Tfh-cell-deficient), B cells, or Foxp3+ T-regulatory cells (Treg cell deficient), were given hydrodynamic injections of pAAV/HBV1.2 plasmids. Some mice were given injections of sorted Tfh cells, pan-B cells, Treg cells, or a blocking antibody against CTLA4. Production of antibodies against HBsAg and clearance of HBV were assessed by flow cytometry, enzyme-linked immunosorbent assay, polymerase chain reaction, and immunohistochemical analyses. We obtained blood samples from patients with HBV infection and isolated Treg cells. We measured the ability of Treg cells to suppress production of interleukin 21 (IL21) in CD4+ T cells. Results Immune-competent C57BL/6N and C57BL/6J mice transfected with the plasmid encoding HBV had features of viral clearance and viral persistence observed in humans. A Tfh-cell response to HBsAg was required for clearance of HBV and was suppressed by Treg cells in mice with persistent HBV infection. Depletion of Treg cells or inhibition of Treg-cell function (with blocking antibody against CTLA4) restored the Tfh-cell response against HBsAg and clearance of HBV in mice. Impaired Tfh-cell response to HBsAg was observed in blood from patients with chronic HBV infection, responsiveness was restored by depletion of Treg cells or blocking antibody against CTLA4. Conclusions In studies of HBV-infected mice and blood from patients with chronic HBV infection, we found a Tfh-cell response to HBsAg of to be required for HBV clearance, and that this response was blocked by Treg cells. Inhibiting Treg-cell activity using neutralizing antibody against CTLA4 restored the ability of Tfh cells to clear HBV infection; this approach might be developed for treatment of patients with chronic HBV infection.

Kupffer cells (KCs) are the resident macrophages of the liver, and they respond to and counteract metabolic stresses, such as those imposed by high-fat diet feeding in mouse models. However, little is known regarding the role of these cells in maintaining metabolic homeostasis under metabolically normal conditions. In this study, we found that depletion of KCs in vivo led to enhanced lipolysis in adipose tissue by increasing the expression of FGF21, a metabolic regulator, in hepatocytes. IL-1β secreted from KCs contributed to the suppression of FGF21 expression in hepatocytes. FGF21 overexpression led to a lean phenotype and enhanced lipolysis in mice. KC depletion resulted in a lack of IL-1β signaling in the liver, leading to elevated expression of FGF21 in hepatocytes. FGF21 promoted lipolysis in adipose tissue and led to hyperlipidemia and decreased body weight. The secretion of IL-1β in KCs was mediated by bacterial products. Antibiotic treatment also led to enhanced lipolysis. Therefore, the current study identified a physiological role of KCs in the regulation of adipose lipolysis.

Macrophages have been suggested to contribute to constructing a cancer stem cell (CSC) niche. However, whether and how macrophages regulate the activity of CSCs through juxtacrine signaling are poorly understood. Here we report LSECtin, a transmembrane protein highly expressed on tumor-associated macrophages (TAMs), enhances stemness of breast cancer cells (BCCs). We identified BTN3A3, a B7 family member with previously unknown functions as the receptor for LSECtin on BCCs responsible for stemness-promoting effect of LSECtin. In mice bearing human tumor xenografts, either macrophage-specific ablation of LSECtin or silencing of BTN3A3 in BCCs decreased CSC frequency and tumor growth. Admixture of LSECtin-positive macrophages increased the tumorigenic activity of BCCs dependent on BTN3A3. Disruption of the LSECtin-BTN3A3 axis with BTN3A3-Fc or anti-BTN3A3 mAb has a therapeutic effect on breast cancer. These findings define a juxtacrine signaling mechanism by which TAMs promote cancer stemness. Targeting this axis in the CSC niche may provide potential therapies to breast cancer.

Epithelial barrier disruption is a major cause of inflammatory bowel disease (IBD); however, the cellular and molecular regulation of intestinal epithelial homeostasis remains largely undefined. Here, we show that the C-type lectin receptor LSECtin (Clec4g) on macrophages is required for protection against dextran sulfate sodium-induced colitis. Mechanistically, LSECtin promotes apoptotic cell clearance by macrophages and induces the production of antiinflammatory/tissue repair factors in an engulfment-dependent manner, which in turn stimulates epithelial cell proliferation. Deletion of LSECtin results in defective engulfment by colon macrophages, leading to aberrant proresolving factor production and impaired intestinal epithelium repair. Collectively, our findings suggest that LSECtin-dependent corpse clearance by macrophages can direct intestinal regeneration and maintenance of the mucosal barrier after injury.

Accumulating evidence has indicated that natural killer cells (NK cells) play an important role in immune responses generated in the liver. However, the underlying molecular basis for local immune regulation is poorly understood. Mice were intraperitoneally injected with polyinosinic-polycytidylic acid (PolyI:C) at a dose of 20 mg/kg body wt. The percentage and absolute number of NK cells in the liver were analysed with flow cytometry. LSECtin knockout mice and LSECtin cDNA plasmids were used for analyze the role of LSECtin in hepatic NK cell regulation in vivo. Here, we show that the C-type lectin LSECtin, a member of the DC-SIGN family, is a novel liver regulator for NK cells. LSECtin could bind to NK cells in a carbohydrate-dependent manner and could regulate the number of hepatic NK cells. In the NK cell-mediated acute liver injury model induced with PolyI:C, the exogenous expression of LSECtin accelerated NK cell-induced liver injury, whereas the absence of LSECtin ameliorated this condition. Our results reveal that LSECtin is a novel, liver-specific NK cell regulator that may be a target for the treatment of inflammatory diseases in the liver., Highlights • Liver-specific C-type lectin, LSECtin, binds to NK cells. • LSECtin expression in vivo accelerates inflammatory liver injury induced by polyI:C. • Absence of LSECtin in vivo decreased the member of hepatic NK cells. • LSECtin is not only an activated T cells but also a NK cells modulator in the liver.

Fatal Ebola virus infection is characterized by a systemic inflammatory response similar to septic shock. Ebola glycoprotein (GP) is involved in this process through activating dendritic cells (DCs) and macrophages. However, the mechanism is unclear. Here, we showed that LSECtin (also known as CLEC4G) plays an important role in GP-mediated inflammatory responses in human DCs. Anti-LSECtin mAb engagement induced TNF-α and IL-6 production in DCs, whereas silencing of LSECtin abrogated this effect. Intriguingly, as a pathogen-derived ligand, Ebola GP could trigger TNF-α and IL-6 release by DCs through LSECtin. Mechanistic investigations revealed that LSECtin initiated signaling via association with a 12-kDa DNAX-activating protein (DAP12) and induced Syk activation. Mutation of key tyrosines in the DAP12 immunoreceptor tyrosine-based activation motif abrogated LSECtin-mediated signaling. Furthermore, Syk inhibitors significantly reduced the GP-triggered cytokine production in DCs. Therefore, our results demonstrate that LSECtin is required for the GP-induced inflammatory response, providing new insights into the EBOV-mediated inflammatory response., Author Summary Ebola virus (EBOV), a highly virulent pathogen, causes a severe hemorrhagic fever syndrome. The fatal infection is characterized by a systemic inflammatory response similar to septic shock. Ebola glycoprotein (GP) is thought to contribute to disease pathogenesis, as high amounts of shed GP from virus-infected cells are detected in patients, and activate macrophages and dendritic cells (DCs) to produce proinflammatory cytokines. Here, we show that LSECtin plays an important role in GP-mediated inflammatory responses in human DCs. LSECtin is a DAP12-coupled receptor able to initiate specific signaling events in human DCs. LSECtin interacts with Ebola GP and results in DAP12 phosphorylation. LSECtin knockdown impairs the production of proinflammatory cytokines induced by Ebola GP. Thus, this study suggests that LSECtin may contribute to Ebola GP-mediated pathogenicity.

Liver sinusoidal endothelial cell lectin (LSECtin) was recently reported to suppress intrahepatic T cell immunity and to limit immune-mediated liver injury. However, its role in the outcome and pathogenesis of viral infection has not yet been elucidated. Using a mouse model infected with a hepatotropic adenovirus, we found that the absence of LSECtin led to a higher frequency of intrahepatic effector CTLs. These cells produced higher levels of antiviral cytokines and cytotoxic factors and exhibited an increased expression of the transcription factors T-bet and Runx3. This phenotype observed in the LSECtin-knockout cells mediated a more efficient virus-specific cytotoxity compared with that of wild-type cells. As a consequence, LSECtin deficiency significantly accelerated liver adenovirus clearance. In contrast, LSECtin upregulation in the liver delayed viral clearance; this delayed clearance was accompanied by the downregulation of the antiviral activity of CTLs. We further constructed an immunocompetent mouse model of acute hepatitis B viral infection to demonstrate that LSECtin significantly delayed the clearance of hepatitis B virus from blood and infected hepatocytes by limiting the frequency of hepatitis B virus–specific IFN-γ–producing cells. Consistent with this function, LSECtin was upregulated in the liver of mouse models of viral hepatitis. Taken together, our results suggest that LSECtin may facilitate the reduction of liver inflammation at the cost of delaying virus clearance and that this effect might be hijacked by the virus as an escape mechanism.

Background & Aims The liver is an organ with paradoxic immunologic properties and is known for its tolerant microenvironment, which holds important implications for hepatic diseases. The molecular basis for this local immune suppression, however, is poorly understood. In this study, we aimed to determine the role of liver sinusoidal endothelial cell lectin (LSECtin), a recently identified member of the dendritic cell-specific ICAM-3 grabbing nonintegrin (DC-SIGN) family, in the regulation of hepatic T-cell immune response. Methods The regulation of T-cell effector function by LSECtin was determined by co-stimulated T cells with anti-CD3/CD28 monoclonal antibody and LSECtin protein, or co-culture of T-cell receptor transgenic T cells with mouse LSECs in vitro. We generated LSECtin knockout mice and prepared recombinant LSECtin protein and complementary DNA plasmids to analyze the role of LSECtin in hepatic T-cell immune regulation in vivo. Results We showed that LSECtin specifically recognized activated T cells and negatively regulated their immune responses. In mice with T-cell–mediated acute liver injury, the lack of LSECtin accelerated the disease owing to an increased T-cell immune response, whereas the exogenous administration of recombinant LSECtin protein or plasmid ameliorated the disease via down-regulation of T-cell immunity. Conclusions Our results reveal that LSECtin is a novel regulator of T cells and expose a crucial mechanism for hepatic T-cell immune suppression, perhaps opening up a new approach for treatment of inflammatory diseases in the liver.