論文

Background: Suppressor of cytokine signaling 1 (SOCS1) is a negative regulator of cytokine signaling whose expression is induced in the rat heart by cardiotrophin-1 (CT-1). Sepsis-induced myocardial depression results from the expression of inducible nitric oxide synthase (iNOS) evoked by inflammatory cytokines. Methods and Results: The effect of CT-1 on lipopolysaccharide (LPS)-induced cardiac dysfunction was examined in a rat model of sepsis. In the absence of CT-1, LPS (1 mg/kg ip) elicited a reduction of systolic function and dilation of the ventricular cavity within 3-6 h after administration. These physiological effects were accompanied by increased ventricular phosphorylation of signal transducers and activators of transcription (STAT) 1 and STAT3, activation of nuclear factor-• B• and expression of iNOS mRNA. Notably, administration of CT-1 (20• g •/kg iv) immediately prior to LPS significantly inhibited all of these LPS-induced changes. To determine whether SOCS1 expression in cardiomyocytes is sufficient to inhibit LPS- and cytokine-induced expression of iNOS mRNA, the effects of forced expression of SOCS1 in cultured neonatal cardiomyocytes were investigated using an adenovirus-mediated transfection system. Forced expression of SOCS1 significantly inhibited iNOS transcription induced by LPS, tumor necrosis factor-• •or interferon-•.• Conclusions: CT-1-mediated expression of SOCS1 in cardiomyocytes may be a useful target for preventing sepsis-induced myocardial depression.

Lipopolysaccharide (LPS) activates macrophages through toll-like receptor (TLR) 4. Although the mechanism of the TLR signaling pathway has been well documented, the mechanism of the negative regulation in response to LPS, particularly LPS tolerance, is still poorly understood. In this study we identified and characterized a novel interferon- and LPS-inducible gene, FLN29, which contains a TRAF6-related zinc finger motif and TRAF family member-associated NF-κB activator-related sequences. The induction of FLN29 was dependent on STAT1. The forced expression of FLN29 in macrophage-like RAW cells resulted in the suppression of TLR-mediated NF-κB and mitogen-activated protein kinase activation, while a reduced expression of FLN29 by small interfering RNA partly cancelled the down-regulation of LPS signaling. Furthermore, we demonstrated that NF-κB activation induced by TRAF6 and TAB2 was impaired by co-expression of FLN29, suggesting FLN29 may regulate the downstream of TRAF6. Taken together, FLN29 is a new negative feedback regulator of TLR signaling. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.

Immune and inflammatory systems are controlled by multiple cytokines, including interleukins (ILs) and interferons. Many of these cytokines exert their biological functions through Janus tyrosine kinases (JAKs) and STAT transcription factors. The CIS (cytokine-inducible SH2 protein) and SOCS (suppressors of cytokine signaling) are a family of intracellular proteins, several of which have emerged as key physiological regulators of cytokine-mediated homeostasis, including innate and adaptive immunity. We investigated the roles of suppressors of cytokine signaling (SOCSs) in regulating dendritic cell (DC) maturation and function. We showed that SOCS1-deficient DCs induced stronger Th1-type responses both in vitro and in vivo. SOCS1-deficient DCs induced higher interferon-γ (IFNγ) production from naive T cells than wild-type DCs in vitro. Lymph node T cells also produced higher amount of IFNγ when SOCS1-deficient bone marrow-derived (BM) DCs were transferred in vivo. Moreover, SOCS1-/- BMDCs raised more effective anti-tumor immunity than wild-type BMDCs. On the other hand, SOCS3-deficeint BMDCs expressed lower levels of MHC, co-stimulators and CD40 in response to LPS. SOCS3-deficient DCs induced lower T cell responses. Thus, SOCS1 and SOCS3 reciprocally regulate DC maturation. © 2005.

The closure of skin wounds is essential for resistance against microbial pathogens, and keratinecyte migration is an important step in skin wound healing. Cathelicidin hCAP18/LL-37 is an innate antimicrobial peptide that is expressed in the skin and acts to eliminate microbial pathogens. Because hCAP18/LL-37 is up-regulated at skin wound sites, we hypothesized that LL-37 induces keratinocyte migration. In this study, we found that 1 μg/ml LL-37 induced the maximum level of keratinocyte migration in the Boyden chamber assay. In addition, LL-37 phosphorylated the epidermal growth factor receptor (EGFR) after 10 min, which suggests that LL-37-induced keratinocyte migration occurs via EGFR transactivation. To test this assumption, we used inhibitors that block the sequential steps of EGFR transactivation, such as OSU8-1, CRM197, anti-EGFR no. 225 Ab, and AG1478. All of these inhibitors completely blocked LL-37-induced keratinocyte migration, which indicates that migration occurs via HB-EGF-mediated EGFR transactivation. Furthermore, CRM197, anti-EGFR no. 225, and AG1478 blocked the LL-37-induced phosphorylation of STAT3, and transfection with a dominant-negative mutant of STAT3 abolished LL-37-induced keratinocyte migration, indicating the involvement of the STAT3 pathway downstream of EGFR transactivation. Finally, we tested whether the suppressor of cytokine signaling (SOCS)/cytokine-inducible Src homology 2-containing protein (CIS) family of negative regulators of STAT3 regulates LL-37-induced keratinocyte migration. Transfection with SOCS1/Jak2 binding protein or SOCS3/CIS3 almost completely abolished LL-37-induced keratinocyte migration. In conclusion, LL-37 induces keratinocyte migration via heparin-binding-EGF-mediated transactivation of EGFR, and SOCS1/Jak2 binding and SOCS3/CIS3 negatively regulate this migration. The results of this study suggest that LL-37 closes skin wounds by the induction of keratinocyte migration. Copyright © 2005 by The American Association of Immunologists, Inc.

The suppressor of cytokine signaling (SOCS) 1 is a negative regulator in multiple cytokine-related aspects to maintain immunological homeostasis. Here, we studied a role of SOCS1 on dendritic cell (DC) maturation in the mice lacking either TCRα chain or CD28 in SOCS1-deficient background, and found that the SOCS1 could restore acute phase of inflammatory response in SOCS1-deficient mice. The CD11c+ CD8- DC population in freshly isolated splenic DCs from normal mice highly expressed SOCS1. However, in SOCS1-deficient environment, the proportion of CD8α+ DCs (CD8 DCs) noticeably increased without affecting the cell number of conventional and plasmacytoid DC populations. This population revealed the CD11cdull CD8α+ CD11b- CD45RA- B220- phenotype, which is a minor population in normal mice. Localization of the abnormal CD8 DCs in splenic microenvironments was mainly restricted to deep within red pulp. The CD8 DCs secrete a large amount of IFN-γ, IL-12 and B lymphocyte stimulator/B cell activation factor of the tumor necrosis factor family in response to LPS and CpG stimulation. This is responsible for the development of DC-mediated systemic autoimmunity in the old age of SOCS1-deficient mice. Moreover, the CD8 DC subsets expressed more indoleamine 2,3-dioxygenase and IL-10, and hence inhibit the allogeneic proliferative T cell response and antigen-induced Th1 responses. Therefore, SOCS1 expression during DC maturation plays a role in surveillance in controlling the aberrant expansion of abnormal DC subset to maintain homeostasis of immune system. © The Japanese Society for Immunology. 2005. All rights reserved.

Caveolin-1 (Cav-1) has been suggested to function as a negative regulator of mitogen-stimulated proliferation and the Ras-p42/44 ERK (MAP kinase) pathway in a variety of cell types. However, the molecular basis of this suppression has not been clarified. Spred/Sprouty family proteins are also negative regulators of the ERK pathway by interacting with Raf-1. The Spred/Sprouty family proteins contain a cysteine-rich (CR) domain at the C-terminus, which is thought to be palmitoylated like Cav-1 and necessary for membrane anchoring. In this study, we demonstrated that Spred-1 localized in cholesterol-rich membrane raft/caveola fractions and interacted with Cav-1. To clarify the biological effect of Cav-1/Spred-1 interaction, we used hematopoietic cells that lacked expression of caveolins but expressed Spred-1. Forced expression of Cav-1 suppressed SCF- and IL-3-induced proliferation and ERK activation. Furthermore, forced expression of exogenous Spred-1 in Cav-1-expressing cells further suppressed proliferation and ERK activation. These data suggest that Spred-1 inhibits ERK activation in collaboration with Cav-1. © Blackwell Publishing Limited.

WSX-1 (IL-27R) is a class I cytokine receptor with homology to gp130 and IL-12 receptors and is typically expressed on CD4+ T lymphocytes. Although previous reports have clarified that IL-27/WSX-1 signaling plays critical roles in both Th1 differentiation and attenuation of cell activation and proinflammatory cytokine production during some bacterial or protozoan infections, little is known about the importance of WSX-1 in cytokine-mediated diseases of allergic origin. To this aim, we took advantage of WSX-1-deficient (WSX-1-/-) mice and induced experimental asthma, in which Th2 cytokines are central modulators of the pathology. OVA-challenged WSX-1 -/- mice showed marked enhancement of airway responsiveness with goblet cell hyperplasia, pulmonary eosinophil infiltration, and increased serum IgE levels compared with wild-type mice. Production of Th2 cytokines, which are largely responsible for the pathogenesis of asthma, was augmented in the lung or in the culture supernatants of peribronchial lymph node CD4+ T cells from WSX-1-/- mice compared with those from wild-type mice. Surprisingly, IFN-γ production was also enhanced in WSX-1-/- mice, albeit at a low concentration. The cytokine overproduction, thus, seems independent from the Th1-promoting property of WSX-1. These results demonstrated that IL-27/WSX-1 also plays an important role in the down-regulation of airway hyper-reactivity and lung inflammation during the development of allergic asthma through its suppressive effect on cytokine production. Copyright © 2005 by The American Association of Immunologists, Inc.

WSX-1 is a component of the IL-27R. Analyses of WSX-1 knockout (WSX-1-/-) mice have shown that IL-27/WSX-1 signaling is essential for the proper development of Th1 responses and that WSX-1 can suppress cellular activation and pro-inflammatory cytokine production. We have generated transgenic mouse lines over-expressing the WSX-1 gene under the control of the T cell-specific CD2 promoter (WSX-1 Tg mice). Unexpectedly, like activated CD4+ T cells from WSX-1-/- mice, activated CD4+ T cells from WSX-1 Tg mice showed increased proliferation, augmented IL-2 production and up-regulated surface expression of activation markers. IL-27-mediated tyrosine phosphorylation of STAT1 was also enhanced in WSX-1 Tg CD4+ T cells, but STAT3 activation was normal. Exogenous IL-27 supported the proliferation of wild-type CD4+ T cells but suppressed that of WSX-1 Tg cells. WSX-1 over-expression increased IFN-γ production in Th1-polarized CD4+ T cells, but also promoted Th2 cytokine production under Th 1-polarizing conditions. Importantly, WSX-1 over-expression failed to suppress Th2 cytokine production under Th 2-polarizing conditions. Cytokine hyperproduction was also observed in vivo in WSX-1 Tg mice injected with Con A. Our data suggest that WSX-1 plays a pivotal role in regulating T cell responsiveness to TCR stimulation and that the correct balance of STAT1/STAT3 activation downstream of IL-27R engagement is crucial for the physiological function of IL-27. © The Japanese Society for Immunology 2005. All rights reserved.

We report here that loss of the Sprouty2 gene (also known as Spry2) in mice resulted in enteric nerve hyperplasia, which led to esophageal achalasia and intestinal pseudo-obstruction. Glial cell line-derived neurotrophic factor (GDNF) induced hyperactivation of ERK and Akt in enteric nerve cells. Anti-GDNF antibody administration corrected nerve hyperplasia in Sprouty2-deficient mice. We show Sprouty2 to be a negative regulator of GDNF for the neonatal development or survival of enteric nerve cells. © 2005 Nature Publishing Group.

Neuromedin U (NMU) is a neuropeptide that is expressed in the gastrointestinal tract and central nervous system. NMU interacts with two G protein-coupled receptors, NMU-R1 and NMU-R2. Whereas NMU-R2 localizes predominantly to nerve cells, NMU-R1 is expressed in peripheral tissues including lymphocytes and monocytes, suggesting a role of NMU in immunoregulation. However, the functions of NMU in peripheral tissues have not been clarified. In this study, using NMU-deficient mice, we first demonstrated that NMU plays an important role in mast cell-mediated inflammation. Complete Freund's adjuvant-induced mast cell degranulation as well as edema and neutrophil infiltration, which occurred weakly in mast cell-deficient WBB6F 1 - W/Wv mice, did not occur in NMU-deficient mice. Moreover, intraplantar injection of NMU into paws induced early inflammatory responses such as mast cell degranulation, vasodilation, and plasma extravasation in WT mice but not in WBB6F1 - W/Wv mice. NMU-R1 was highly expressed in primary mast cells, and NMU induced Ca 2+ mobilization and degranulation in peritoneal mast cells. These data indicate that NMU promotes mast cell-mediated inflammation; therefore, NMU receptor antagonists could be a novel target for pharmacological inhibition of mast cell-mediated inflammatory diseases. © The Rockefeller University Press.

Immune and inflammatory systems are controlled by multiple cytokines, including interleukins and interferons. Many of these cytokines exert their biological functions through Janus kinases and signal transducers and activators of transcription factors. The cytokine-inducible SH2 protein (CIS) and suppressors of cytokine signaling (SOCS) are members of a family of intracellular proteins, several of which have emerged as key physiological regulators of cytokine-mediated homeostasis, including innate and adaptive immunity. This article focuses on the molecular mechanism of the action of CIS/SOCS family proteins and their roles in immune regulation and inflammatory diseases. © Copyright 2005 by Humana Press Inc.