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VSIG4, a newly identified co-inhibitory molecule belonging to the B7-related family, is exclusively expressed on tissue-resident macrophages and is involved in the suppression of T cell proliferation and cytokine production. We sought to characterize the role of VSIG4 in anti-tumor immunity in the tumor microenvironment, focusing on VSIG4-expressing tumor-associated macrophages (TAMs). We found that VSIG4-expressing TAMs negatively regulated antigen-specific T cell proliferation and cytokine production through direct inhibition via cell cycle arrest, but not apoptosis, as well as through their arginase 1 activity. Furthermore, VSIG4-expressing TAMs suppress tumor-specific CD8

Our previous study demonstrated that G-CSF treatment increased the expression of TLR2 in donor grafts; this contributed to rapid engraftment after allogeneic hematopoietic stem cell transplantation (HSCT) in mice. In the current study, we investigated the effects of upregulated TLR2 expression in G-CSF-mobilized donor grafts on acute graft-versus-host disease (GVHD). We found that TLR2 was highly expressed on myeloid cell populations but not T and B cells from the spleens of G-CSF-treated donor mice. After transplantation, the mortality and disease severity in recipients were not significantly different between G-CSF-treated TLR2-/- and wt donor grafts. Although endogenous TLR2 ligand was detected in the serum of both recipients, T cells from TLR2-/- and wt donors have the same ability regarding alloreactivity. Moreover, the blockade of TLR2 signaling in recipients by administering anti-TLR2 blocking antibody after BMT did not lead to a significant difference in acute GVHD compared with control IgG treatment. However, the hematopoietic ability of G-CSF-mobilized lin−c-kit+ HSCs from TLR2-/- donor grafts was lower than that from wt donor grafts. Our results demonstrate that upregulated TLR2 expression in G-CSF-mobilized donor grafts has no effect on acute GVHD, suggesting that TLR2 is a valuable target for increasing HSCT efficiency in order to enhance engraftment without exacerbating acute GVHD.

Aim B7-H1, which belongs to the B7 family of costimulatory molecules, is implicated in the ability of tumors to evade the host immune response. The development of evasion mechanisms within the tumor microenvironment may be responsible for poor therapeutic responses. In this manuscript, we report that the 15-deoxy-δ 12,14 -prostaglandin J 2 (15d-PGJ 2 ), peroxisome proliferator-activated receptor gamma (PPARγ) activator leads to the downregulation of the cancer-associated expression of B7-H1 in response to interferon-gamma (IFN-γ) and the associated signaling cascades. Main methods The expression of B7-H1 from IFN-γ-induced B16F10 melanoma cells was measured with flow cytometric analysis. The regulatory mechanisms of 15d-PGJ 2 on cellular signaling pathways were examined using Western blot and electrophoretic mobility shift assays. Key findings The flow cytometric analysis revealed that the B7-H1 costimulatory molecule is significantly upregulated in B16F10 melanoma cells by stimulation with IFN-γ. However, 15d-PGJ 2 strongly downregulates B7-H1 expression in IFN-γ-stimulated B16F10 melanoma cells. Furthermore, the significant damping effect of 15d-PGJ 2 on B7-H1 expression involves the inhibition of the tyrosine phosphorylation of Janus kinase (Jak) and signal transducer(s) and activator(s) of transcription (STAT) and, thereby, the interferon regulatory factor-1 (IRF-1) trans-activation of STAT. These effects of 15d-PGJ 2 were not abrogated by the PPARγ antagonist GW9662, indicating that they occur through a PPARγ-independent mechanism. Significance In this study, we demonstrate that 15d-PGJ 2 suppresses the IFN-γ-elicited expression of B7-H1 by the inhibition of IRF-1 transcription via the Jak/STAT signaling pathway through a PPARγ-independent mechanism in mouse melanoma cells.

Tumor cells have been used as the tumor antigen sources for developing cancer vaccines. Due to their low immunogenicity, tumor antigens are combined with various adjuvants to enhance immunogenicity of cancer vaccines. Among them, a natural killer T cell (NKT)-ligand, α-galactosylceramide (αGC) has been reported as a powerful adjuvant showing therapeutic effects in solid tumors as well as hematological malignancies including lymphoma. In this study, we applied αGC-based tumor cell vaccine in mouse multiple myeloma model. The αGC-loaded MOPC315BM myeloma cell vaccine efficiently retarded tumor growth, induced regression of established tumors, and protected surviving mice from tumor rechallenge. Therapeutic responses were associated with induction of strong humoral immune responses, including myeloma-specific antibodies, and cellular immune responses, including myeloma-specific CD8(+) cytotoxic T lymphocytes and memory T cells. In addition, regulatory T cells were significantly decreased in mice that received the αGC-loaded myeloma cell vaccine. Thus, our results demonstrated that αGC-loaded myeloma vaccine efficiently promoted NKT-dependent anti-tumor immunity in a mouse model. These findings are informative for improving the efficacy of tumor-cell-based immunotherapy for patients with MM and other CD1d-expressing tumors.

The immunoregulatory effects of granulocyte colony-stimulating factor (G-CSF) on allogeneic peripheral blood cell transplantation (PBCT) have been demonstrated to reduce acute graft-versus-host disease (GVHD). However, the underlying mechanism is still not clear. In this study, we focused on the direct effects of G-CSF on donor CD4(+) T cell responses after transplantation. We observed that lethally irradiated B6D2F1 recipient mice that are transplanted with CD4(+) T cells from G-CSF-treated B6 donors showed mild attenuations in severity and mortality compared with recipients transplanted with PBS-treated CD4(+) T cells. Notably, skin GVHD was significantly reduced, but no such reduction was observed in other organs. Although there was no difference with respect to alloreactive expansion or Foxp3(+) Treg induction, the use of G-CSF-treated CD4(+) T cells significantly reduced the numbers of IL-17-producing and RORγt-expressing cells in the secondary lymphoid organs of allogeneic recipients after transplantation compared with the use of the control cells. Finally, we found that the suppressor of cytokine signaling-3 (SOCS3) expression in G-CSF-treated donor CD4(+) T cells was much higher than that in control CD4(+) T cells. Our results demonstrate that the inhibition of Th17 cell differentiation by SOCS3 induction is associated with the immunoregulatory role of G-CSF in CD4(+) T cell-mediated acute GVHD.

Tryptophan-derived metabolites, initiated by indoleamine 2,3-dioxygenase (IDO), preferentially induce activated T cell death, which is an important mechanism in IDO-mediated T cell suppression. However, the mechanism of this phenomenon remains unclear. We found that 3-hydroxyanthranilic acid (3-HAA), the most potent metabolite, selectively eliminated activated T cells, which were stimulated with the bacterial superantigen staphylococcal enterotoxin A (SEA), but not resting T cells, by inducing apoptosis. We observed 3-HAA-induced depletion of intracellular glutathione (GSH) in activated T cells. When GSH levels were maintained by addition of N-acetylcysteine (NAC) and GSH, 3-HAA-mediated T cell death was completely inhibited. This was associated with extrusion of GSH from activated T cells without increased reactive oxygen species (ROS) formation. Finally, we showed that administration of 3-HAA in mice after allogeneic bone marrow transplantation reduced acute graft-versus-host disease (GVHD) lethality by inhibition of alloreactive T cell expansion through intracellular GSH depletion. Our data suggest that direct depletion of intracellular GSH is the major mechanism of 3-HAA-mediated activated T cell death.

While programmed death-1 (PD-1), a co-inhibitory member of CD28 immunoglobulin superfamily plays negative roles in effector functions of T cells and B cells, little is known about the function of PD-1 expressed on innate immune cells. In this study, we demonstrate that IL-12 production was greatly suppressed in LPS-stimulated RAW264.7 cells upon PD-1 engagement with B7-H1.Fc fusion protein, and was restored in the presence of antagonistic anti-PD-1 mAb. PD-1-mediated suppression of IL-12 production in LPS-stimulated RAW264.7 cells was mediated by inhibition of Janus N-terminal-linked kinase (JNK) signaling pathway, and to a lesser extent, phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway through the recruitment of SHP-2 to PD-1 cytoplasmic tail. B7-H1.Fc-mediated PD-1 engagement also downregulates the expression of co-stimulatory molecules such as CD80, CD86, MHC class I and II proteins in LPS-stimulated RAW264.7 cells. Furthermore, the endocytic activity is enhanced but the allostimulatory capacity is suppressed in LPS-treated RAW264.7 cells upon PD-1 engagement. Taken together, our results reveal a novel function of macrophage PD-1 in the negative regulation of IL-12 synthesis and differentiation into dendritic cell-like cells.

The aim of the present work is focused on protective effects of an edible red alga, Laurencia undulata ethanolic (EtOH) extracts (LU) containing a large amount of polyphenols against OVA-induced murine allergic airway reactions using in vivo histological and cytokine assay. Mice sensitized and challenged with ovalbumin (OVA) showed typical asthmatic reactions as follows: an increase in the number of eosinophil in bronchoalveolar lavage fluid; a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; the development of airway hyperresponsiveness; the detection of TNF-α and Th2 cytokines, such as IL-4 and IL-5 in the bronchoalveolar lavage (BAL) fluid; and detection of allergen-specific IgE in the serum. The successive intraperitoneal administration of LU before the last airway OVA-challenge resulted in a significant inhibition of all asthmatic reactions. These results suggest that L. undulata polyphenolic extracts possess therapeutic potential for combating bronchial asthma associated with allergic diseases.

A potent anti-inflammatory activity was detected from methanol extracts prepared from roots of Rhodiola sacra. In the preliminary test, methanol extracts from the roots of R. sacra could potently attenuate histological change in the lung tissue after lipopolysaccharide (LPS)-treated inflammatory stimulation. The active compound has been identified as 3-(3,4-dihydroxy-phenyl)-acrylic acid phenethyl ester (caffeic acid phenethyl ester, CAPE). In vivo study on protective effect of R. sacra CAPE against inflammatory response by endotoxin shock in the LPS-induced inflammatory mice showed significantly attenuated the intravenous LPS-induced increase in plasma TNF-α and IL-1β concentration. R. sacra CAPE significantly inhibited LPS-induced nuclear transcription factor-κB (NF-κB) activation. It also down-regulates enhanced matrix metalloproteinase-9 (MMP-9) activity triggered by LPS in mouse lung. In the present study, the results indicate that the intake of R. sacra CAPE could provide health benefits, reducing the magnitude of the inflammatory process triggered by endotoxin shock and the production of inflammatory mediators.

Ecklonia cava (EC) is a brown alga that evidences radical scavenging activity, bactericidal activity, tyrosinase inhibitory activity, and protease inhibitory activity. However, its anti-allergic effects remain poorly understood. In the current study, we attempted to determine whether pretreatment with EC induces a significant inhibition of asthmatic reactions in a mouse asthma model. Mice sensitized and challenged with ovalbumin (OVA) evidenced typical asthmatic reactions, as follows: an increase in the number of eosinophils in bronchoalveolar lavage fluid; a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; the development of airway hyperresponsiveness; the detection of tumor necrosis factor-alpha (TNF-α) and Th2 cytokines, including IL-4 and IL-5 in the bronchoalveolar lavage (BAL) fluid; and the detection of allergen-specific immunoglobulin E (IgE) in the serum. However, the administration of EC extract prior to the final airway OVA challenge resulted in a significant inhibition of all asthmatic reactions. We also demonstrated that EC extracts treatment resulted in significant reductions on matrix metalloproteinase-9 (MMP-9) and Suppressor of cytokine signaling-3 (SOCS-3) expression and a reduction in the increased eosinophil peroxidase (EPO) activity. The treatment of animals with EC extracts resulted in a significant reduction in the concentrations of the Th2 cytokine (IL-4 and IL-5) in the airways, without any concomitant increase in the concentration of Th1 cytokines. These findings indicate that EC extracts may prove useful as an adjuvant therapy for allergic airway reactions via the inhibition of the Th2 response. Accordingly, this study may provide evidence that EC extract performs a critical function in the amelioration of the pathogenetic process of asthma in mice.

Caffeic acid phenethyl ester (CAPE) is a biologically active ingredient of propolis, which has several interesting biological properties, including antioxidant and anti-inflammatory; however, its anti-allergic effects are poorly understood. The objective of this study was to determine whether treatment with CAPE results in significant inhibition of asthmatic reactions in a mouse model. Mice sensitized and challenged with ovalbumin (OVA) had the following typical asthmatic reactions: an increase in the number of eosinophils in bronchoalveolar lavage (BAL) fluid; a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; the development of airway hyperresponsiveness (AHR); the presence of tumor necrosis factor-alpha (TNF-alpha) and Th2 cytokines, including IL-4 and IL-5, in the BAL fluid; and the presence of allergen-specific IgE in the serum. Five successive intraperitoneal administrations of CAPE before the last airway OVA challenge resulted in significant inhibition of characteristic asthmatic reactions. We determined that increased generation of reactive oxygen species (ROS) by inhalation of OVA was diminished via the administration of CAPE in BAL fluid, as well as nuclear factor-kappaB (NF-kappaB) DNA binding activity. These findings indicate that oxidative stress may have a crucial function in the pathogenesis of bronchial asthma, and that CAPE may be useful as an adjuvant therapy for the treatment of bronchial asthma.

Caffeic acid phenethyl ester (CAPE) is an active component of honeybee propolis extracts. It has several positive effects, including anti-inflammatory, anti-oxidation, anti-cancer, anti-bacterial, anti-viral, anti-fungal, and immunomodulatory effects. In particular, the suppressive effect of NF-kappaB may disrupt a component of allergic induction. The principal objective of this experimental study was to evaluate the effects of CAPE on the active systemic anaphylaxis induced by ovalbumin (OVA) challenge in mice. Mice were intraperitoneally sensitized and intravenously challenged with OVA. Histopathological analysis, nuclear factor (NF)-kappaB activation, and the plasma levels of histamine and total IgE after allergen challenge were evaluated. After challenges, all of the sham-treated mice developed anaphylactic symptoms, increased plasma levels of histamine and OVA-specific IgE, marked vascular leakage, NF-kappaB activation, platelet-activating factor (PAF) production, and histological changes including pulmonary edema and hemorrhage in the renal medullae within 20 min. By way of contrast, a reduction in the plasma levels of histamine and OVA-specific IgE and an inhibition of NF-kappaB activation and PAF release were observed in the CAPE-treated mice. In addition, a significant prevention of hemoconcentration and OVA-induced pathological changes were noted. These results indicate that CAPE demonstrates an anti-allergic effect, which may be the result of its protective effects against IgE-mediated allergy.

Water-soluble proteoglycan (WSPG) of Agaricus blazei Murill has been known to stimulate the non-specific complements and humoral immune functions to act as polyclonal activators of B cells and to inhibit tumor growth and metastasis. However, little is known about its immunomodulating effects on murine bone marrow (BM)-derived dendritic cells (DC). In the present study, we examined the maturation process of murine BM-DC. BM cells were cultured in the presence of IL-4 and GM-CSF and the generated immature DC were stimulated with WSPG or LPS (WSPG-DC and LPS-DC, respectively) for 24 h. WSPG significantly enhanced the expression of co-stimulatory molecules (CD80 and CD86) and major histocompatibility complex (MHC) II, as did LPS. IL-12p70 production in WSPG-DC was also identical to that in LPS-DC. The antigen-uptake capacity of WSPG-DC was determined by FITC-labeled dextran uptake. WSPG-DC lost dextran uptake capacity comparable to LPS-DC. The antigen-presenting capacity of WSPG-DC as analyzed by allogeneic T cell proliferation was significantly increased in comparison with immature DC, was identical to LPS-DC, and induced higher levels of IL-2 in responding T cells. These results indicate the immunomodulatory properties of WSPG, which might be therapeutically useful in the control of cancers and immunodeficient diseases through the up-regulation of DC maturation.