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Ko Okumura

Publications and source records attributed to Ko Okumura.

At least 19 recordsLinked to original sources
PubMed2007

TACI regulates IgA production by APRIL in collaboration with HSPG.

Transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) is a member of the tumor necrosis factor (TNF) receptor family that serves as a receptor for B-cell activating factor of the TNF family (BAFF) and as a proliferation-inducing ligand (APRIL). Although TACI is reported to function as a positive or negative regulator for B-cell responses, its roles remain elusive. Experiments using TACI siRNA into B cells indicated that TACI positively regulated APRIL-induced IgA production in collaboration with heparan sulfate proteoglycans (HSPG). Furthermore, TACI negatively regulated BAFF-induced B-cell proliferation and production of IgA and IgG. In addition, B cells treated with heparitinase to denature HSPG showed that HSPG is essential for APRIL-induced B-cell responses such as B-cell proliferation, IgG and IgA production, induction of activation-induced cytidine deaminase (AID), and noncanonical NF-kappaB2. In contrast, phosphorylation of physiological AID kinase, protein kinase A (PKA), was dependent on TACI. Importantly, coligation of TACI and HSPG by specific antibodies, but not by TACI or HSPG ligation itself, could induce the phosphorylation of PKA and IgA production instead of APRIL. Our findings indicate that simultaneous binding of TACI and HSPG on B cells with APRIL is crucial for IgA production.

B-Cell Activating Factor↗
PubMed2007

Antimicrobial peptides human beta-defensin (hBD)-3 and hBD-4 activate mast cells and increase skin vascular permeability.

Antimicrobial peptides human beta-defensins (hBD) are mainly produced by epithelia of several organs including skin, and participate in innate immunity by killing invading pathogens. Besides their microbicidal activities, hBD activate several inflammatory and immune cells. Since hBD are generated by tissues where mast cells are present, we hypothesized that these peptides could activate mast cells. In this study, we demonstrated that both hBD-3 and hBD-4 induced mast cell degranulation, prostaglandin D2 production, intracellular Ca2+ mobilization and chemotaxis. Furthermore, hBD-3- and hBD-4-induced activation of mast cells was suppressed by pertussis toxin and U-73122, inhibitors for G protein and phospholipase C, respectively. We further revealed that hBD-3 and hBD-4 increased vascular permeability in the skin, which was dependent on the presence of mast cells, because hBD-3 and hBD-4 failed to enhance vascular permeability in mast cell-deficient Ws/Ws rats. We also demonstrated that hBD-3 and hBD-4 induced phosphorylation of MAPK p38 and ERK1/2, which were further required for hBD-mediated mast cell activation, as evidenced by the inhibitory effects of p38 and ERK1/2 inhibitors on mast cell degranulation. Together, these findings suggest the key role of hBD in inflammatory responses by recruiting and activating mast cells, and increasing vascular permeability.

Animals↗
PubMed2007

TACI attenuates antibody production costimulated by BAFF-R and CD40.

B cell activating factor of the TNF family (BAFF), plays critical roles in B cell survival, activation, differentiation, and antibody (Ab) production. BAFF binds to three receptors: BAFF-R, transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI) and B cell maturation antigen. While BAFF-R is the primary receptor for B cell costimulation by BAFF, TACI is reported to serve as a positive or negative regulator for B cell responses depending on conditions. To determine the real role of TACI in B cell responses, we examined the functional relationship between TACI and BAFF-R in Ab production from human peripheral blood B cells using agonistic mAb. BAFF-R and CD40 enhanced IgG secretion and B cell proliferation, which were inhibited by TACI. Although TACI induced mild B cell apoptosis, its extent did not correlate with that of TACI-mediated inhibition of IgG secretion. In addition, TACI inhibited B-lymphocyte-induced maturation protein-1 expression, IgG secretion from previously IgG-negative selected B cells, and activation-induced cytidine deaminase expression enhanced by BAFF-R and CD40. Importantly, BAFF-R and CD40 enhanced B cell responsiveness to TACI-mediated suppression. Thus, BAFF may attenuate T cell-independent and -dependent B cell responses by TACI.

Adjuvants, Immunologic↗
PubMed2006

Antibodies to T-cell Ig and mucin domain-containing proteins (Tim)-1 and -3 suppress the induction and progression of murine allergic conjunctivitis.

The T cell Ig and mucin domain-containing proteins (Tim) regulate Th1- and Th2-mediated immune responses. We investigated the ability of Abs blocking Tim-1 or Tim-3 ligand-binding activity to prevent and treat murine experimental allergic conjunctivitis (EC), a Th2-mediated disease. Treatment with either Ab during the induction phase of EC in actively immunized wild-type mice suppressed EC and upregulated Th1 and Th2 immune responses. In contrast, both Abs exacerbated EC in actively immunized IFN-gamma-knockout mice. Thus, both anti-Tim Abs suppress the pathogenic immune responses generated in the induction phase by upregulating systemic IFN-gamma production. Treatment of actively immunized mice and passively immunized mice with either anti-Tim Ab just prior to RW challenge also suppressed EC. Thus, treatment with anti-Tim-1 or anti-Tim-3 Ab can suppress both the induction and progression of EC, which could indicate potential preventive and/or therapeutic approaches for allergic diseases such as allergic conjunctivitis.

Animals↗
PubMed2006

TGF-beta type I receptor kinase inhibitor down-regulates rheumatoid synoviocytes and prevents the arthritis induced by type II collagen antibody.

Rheumatoid arthritis (RA) is characterized by hypertrophic synovial tissues comprising excessively proliferating synovial fibroblasts and infiltrating inflammatory cells. Transforming growth factor-beta (TGF-beta) is a multifunctional cytokine that regulates cell growth, inflammation and angiogenesis by acting on various cell types. In RA synovial tissues, TGF-beta is expressed at high levels. However, the precise role of TGF-beta in RA remains unclear. We herein demonstrated a causal link between the TGF-beta-induced RA synovial cell proliferation and induction of platelet-derived growth factor (PDGF)-AA. In addition, TGF-beta induced IL-6 and vascular endothelial growth factor (VEGF) production by RA synovial fibroblasts associated with nuclear factor-kappa B activation. These effects of TGF-beta on RA synovial fibroblasts were suppressed by TGF-beta type I receptor kinase inhibitor HTS466284. Furthermore, HTS466284 significantly prevented anti-collagen type II antibody-induced arthritis in mice according to the clinical manifestations, histology, tumor necrosis factor-alpha, PDGF and VEGF expression and 5-bromo-2'-deoxyuridine incorporation. These in vitro and in vivo results suggest that TGF-beta plays a role in the development of synovial hyperplasia consisting of synovial cell proliferation, inflammation and angiogenesis. The blockade of TGF-beta signaling may thus become an additional strategy for the treatment of RA.

Animals↗
PubMed2006

An antiapoptotic protein, c-FLIPL, directly binds to MKK7 and inhibits the JNK pathway.

Inhibition of NF-kappaB activation increases susceptibility to tumor necrosis factor (TNF)alpha-induced cell death, concurrent with caspases and prolonged c-Jun N-terminal kinase (JNK) activation, and reactive oxygen species (ROS) accumulation. However, the detailed mechanisms are unclear. Here we show that cellular FLICE-inhibitory protein (c-FLIP) is rapidly lost in NF-kappaB activation-deficient, but not wild-type fibroblasts upon TNFalpha stimulation, indicating that NF-kappaB normally maintains the cellular levels of c-FLIP. The ectopic expression of the long form of c-FLIP (c-FLIPL) inhibits TNFalpha-induced prolonged JNK activation and ROS accumulation in NF-kappaB activation-deficient fibroblasts. Conversely, TNFalpha induces prolonged JNK activation and ROS accumulation in c-Flip-/- fibroblasts. Moreover, c-FLIPL directly interacts with a JNK activator, MAP kinase kinase (MKK)7, in a TNFalpha-dependent manner and inhibits the interactions of MKK7 with MAP/ERK kinase kinase 1, apoptosis-signal-regulating kinase 1, and TGFbeta-activated kinase 1. This stimuli-dependent interaction of c-FLIPL with MKK7 might selectively suppress the prolonged phase of JNK activation. Taken that ROS promote JNK activation and activation of the JNK pathway may promote ROS accumulation, c-FLIPL might block this positive feedback loop, thereby suppressing ROS accumulation.

Animals↗
PubMed2006

B7-H1-induced apoptosis as a mechanism of immune privilege of corneal allografts.

The programmed death-1 (PD-1) costimulatory pathway has been demonstrated to play a role in the regulation of immune responses and peripheral tolerance. We investigated the role of this pathway in establishing an immune privilege status of corneal allografts in mice. B7-H1, but not B7-DC or PD-1, was expressed constitutively in the eye, i.e., cornea, iris-ciliary body, and retina. After corneal allografting, PD-1(+)CD4(+) T cells infiltrated and adhered with B7-H1(+) corneal endothelium. Blockade of PD-1 or B7-H1, but not B7-DC, led to accelerated corneal allograft rejection. In B7-H1-expressing corneal allografts, apoptosis of the infiltrating PD-1(+)CD4(+) or CD8(+) T cells was observed, after which there was allograft acceptance. In contrast, B7-H1 blockade suppressed apoptosis of infiltrating PD-1(+) T cells, which led to allograft rejection. In vitro, destruction of corneal endothelial cells by alloreactive T cells was enhanced when the cornea was pretreated with anti-B7-H1 Ab. This is the first demonstration that the constitutive expression of B7-H1 plays a critical role in corneal allograft survival. B7-H1 expressed on corneal endothelial cells maintains long-term acceptance of the corneal allografts by inducing apoptosis of effector T cells within the cornea.

Animals↗
PubMed2006

Involvement of TNF-like weak inducer of apoptosis in the pathogenesis of collagen-induced arthritis.

TNF-like weak inducer of apoptosis (TWEAK) is a type II membrane protein belonging to the TNF family that regulates apoptotic cell death, cellular proliferation, angiogenesis, and inflammation. However, the role of TWEAK in the pathogenesis of rheumatoid arthritis (RA) remains unclear. In this study, we have investigated the effect of neutralizing anti-TWEAK mAb on the development of collagen-induced arthritis (CIA), a well-established murine model of RA. Administration of anti-TWEAK mAb significantly ameliorated paw swelling, synovial hyperplasia, and infiltration of inflammatory cells. The levels of proinflammatory chemokines such as MCP-1 and MIP-2 in serum and knee joints were reduced by this treatment. Consistently, recombinant TWEAK enhanced the proliferation of MCP-1 and MIP-2 production by synovial cells from CIA mice in vitro. Histological examination also revealed that the treatment with anti-TWEAK mAb suppressed the development of small vessels in synovial tissues. These results indicated anti-inflammatory and antiangiogenic effects of the TWEAK blockade in CIA, which may be also beneficial for the treatment of RA.

Animals↗
PubMed2006

Antimicrobial peptides human beta-defensins stimulate epidermal keratinocyte migration, proliferation and production of proinflammatory cytokines and chemokines.

Besides their microbicidal functions, human beta-defensins (hBD) and LL-37 activate different immune and inflammatory cells, and their expression is enhanced in inflamed skin and cutaneous wound sites. To protect against pathogens, the skin produces antimicrobial peptides including hBDs and LL-37. Therefore, the aim of our study was to investigate whether hBDs participate in cutaneous inflammation and wound healing by inducing keratinocyte migration, proliferation, and production of proinflammatory cytokines/chemokines. We found that hBD-2, -3, and -4 but not hBD-1 stimulated human keratinocytes to increase their gene expression and protein production of IL-6, IL-10, IP-10, monocyte chemoattractant protein-1, macrophage inflammatory protein-3alpha, and RANTES. This stimulatory effect was markedly suppressed by pertussis toxin and U-73122, inhibitors for G protein and phospholipase C, respectively. We also demonstrated that hBDs elicited intracellular Ca2+ mobilization, and increased keratinocyte migration, and proliferation. In addition, these peptides induced phosphorylation of EGFR, signal transducer and activator of transcription (STAT)1, and STAT3, which are intracellular signaling molecules involved in keratinocyte migration and proliferation. In our study, inhibition of these molecules significantly reduced hBD-mediated keratinocyte migration and proliferation. In conclusion, this study provides evidence that human antimicrobial peptides may be involved in skin immunity through stimulating cytokine/chemokine production, and participate in wound healing by promoting keratinocyte migration and proliferation.

Antimicrobial Cationic Peptides↗
PubMed2006

Preferential involvement of Tim-3 in the regulation of hepatic CD8+ T cells in murine acute graft-versus-host disease.

Tim-3, a member of the T cell Ig mucin (TIM) family regulates effector Th1 responses. We examined Tim-3 and its ligand expression as well as the effects of anti-Tim-3 mAb treatment in a murine model of acute graft-vs-host disease (aGVHD). In mice with aGVHD, Tim-3 expression was markedly up-regulated on splenic and hepatic CD4+ and CD8+ T cells, dendritic cells (DCs), and macrophages, and this was especially dramatic in hepatic CD8+ T cells. Both donor- and host-derived CD8+ T cells induced similar levels of Tim-3. Tim-3 ligand expression was also up-regulated in splenic T cells, DCs, and macrophages, but not in the hepatic lymphocytes. The administration of anti-Tim-3 mAbs accelerated aGVHD, as demonstrated by body weight loss, reduction in total splenocyte number, and infiltration of lymphocytes in the liver. IFN-gamma expression by splenic and hepatic CD4+ and CD8+ T cells was significantly augmented by anti-Tim-3 mAb treatment. In addition, the cytotoxicity against host alloantigen by donor CD8+ T cells was enhanced. These results demonstrate that the anti-Tim-3 treatment in aGVHD augmented the activation of effector T cells expressing IFN-gamma or exerting cytotoxicity. Our results suggest that Tim-3 may play a crucial role in the regulation of CD8+ T cells responsible for the maintenance of hepatic homeostasis and tolerance.

Animals↗
PubMed2006

Application of immunoreaction enhancer solutions to an enzyme-linked immunosorbent assay for antigen-specific IgE in mice immunized with recombinant major mite allergens or ovalbumin.

BACKGROUND: Weak signals for allergen-specific IgE are a problem in murine models for the study of allergies. It has been reported that the removal of IgG from murine sera enhances signal intensity. Very recently, buffer solutions designed to enhance signals in immunoassays have been developed and made commercially available. METHODS: Sera from mice immunized either with a recombinant form of one of the major mite allergens Der p 1, Der f 1 and Der f 2, or with ovalbumin adsorbed to alum were used for the assays. Total IgE was measured by a sandwich enzyme-linked immunosorbent assay (ELISA). Allergen-specific IgE was assayed using plates coated with the allergens after the removal of IgG from sera with protein G-coupled sepharose beads in wells of other plates or with the use of commercially available enhancer solutions without the removal of IgG. IgE binding was detected with horseradish peroxidase-conjugated anti-murine IgE monoclonal antibody as the secondary antibody. RESULTS: Significant levels of total IgE were produced after the immunizations. The in-well pretreatment of diluted sera (1/10 dilution) with protein G-coupled beads enhanced the signals for allergen-specific IgE. The use of the enhancer solutions for dilution of the sera and secondary antibody and prolonged incubation remarkably enhanced the signals at a more extensive dilution of sera (1/200 or less) without the removal of IgG. CONCLUSIONS: An ELISA simply modified with the use of immunoreaction enhancer solutions has advantages in terms of signal intensity and ease of handling for the detection of allergen-specific murine IgE and would be useful for the study of allergies with murine models.

Animals↗
PubMed2006

Crucial commitment of proteolytic activity of a purified recombinant major house dust mite allergen Der p1 to sensitization toward IgE and IgG responses.

The major proteolytic allergen derived from the house dust mite Dermatophagoides pteronyssinus, Der p1, is one of the most clinically relevant allergens worldwide. In the present study, we evaluate the contribution of the proteolytic activity and structure of a highly purified rDer p 1 to immune responses. Mice were i.p. immunized with three forms of rDer p 1 adsorbed to Alum: one enzymatically active, one treated with an irreversible cysteine protease-specific inhibitor, E-64, and one heat denatured. Immunization with E-64-treated or heat-denatured rDer p 1 elicited much less production of serum total IgE and not only rDer p 1-specific IgE but also IgGs compared with immunization with active rDer p 1. Assays for Ab-binding and its inhibition and structural analyses indicated that E-64-treated rDer p 1 retained its global structure and conformational B cell epitopes. A proliferative response and production of IL-5 by spleen cells restimulated with rDer p 1 were observed on immunization with the active rDer p 1 but not E-64-treated rDer p 1. The cells from mice immunized with heat-denatured rDer p 1 exhibited the highest levels of proliferation and production of IL-5 and IFN-gamma. The results indicate that the proteolytic activity of the highly purified rDer p 1 crucially commits to the sensitization process, including both IgE and IgG responses. Additionally, we demonstrated immunogenic differences by functional or structural manipulations of the rDer p 1. The findings have implications for sensitization to this relevant allergen in humans and for the design of modified allergen-vaccines for future allergen-specific immunotherapy.

Animals↗
PubMed2006

Prolonged MHC class II expression and CIITA transcription in human keratinocytes.

A transcriptional cofactor, the MHC class II transactivator (CIITA), is a master regulator of MHC class II gene expression. CIITA expression is restricted in MHC class II-positive cells and is regulated by 4 (human) or 3 (mouse) promoters. To clarify the usage of human CIITA promoters in keratinocytes, we analyzed CIITA mRNA levels in IFN-gamma-stimulated normal human keratinocytes (NHK) by real-time PCR using promoter-specific primers. When the amount of total CIITA mRNA in NHK was quantified at 6h after IFN-gamma-stimulation, the amount of CIITA mRNA detected in NHK did not differ from that seen in the B cell line Raji or the IFN-gamma-stimulated macrophage cell line THP-1. Quantitative real-time PCR using promoter-specific primers showed that type IV CIITA mRNA was strongly transcribed and that type III CIITA mRNA was weakly transcribed in stimulated NHK, while no transcripts from pI and pII were detected. Although type IV mRNA in THP-1 was transiently transcribed by IFN-gamma-stimulation, transcription of type IV in IFN-gamma-stimulated keratinocytes was prolonged. This difference subsequently caused significantly higher expression at 72 h of MHC class II on NHK, compared with THP-1. This is the first report to quantitatively analyze each type of CIITA transcript in NHK.

Cell Line↗
PubMed2006

Amelioration of bone loss in collagen-induced arthritis by neutralizing anti-RANKL monoclonal antibody.

Receptor activator of NF-kappaB (RANK) and its ligand (RANKL) are pivotal regulators of osteoclast differentiation. RANK and RANKL also mediate T cell/dendritic cell (DC) interaction. Previous study has shown that RANK/RANKL interaction induces prolonged DC survival and antigen presentation. In the present study, we have newly established a hybridoma which produces neutralizing anti-RANKL monoclonal antibody (IK22-5). By treating collagen-induced arthritis (CIA) mice with IK22-5, we have investigated the role of RANKL in the pathogenesis of CIA. Although IK22-5 had no effect on immune responses or inflammation, it ameliorated bone loss at the site of inflammation. Histological analyses revealed that osteoclast formation was impaired at the site of joint inflammation in IK22-5-treated CIA mice. These results suggest the utility of anti-RANKL mAb for the prevention of osteoporosis associated with joint inflammation in RA.

Animals↗
PubMed2006

Eradication of established tumors in mice by a combination antibody-based therapy.

Tumor-cell apoptosis is the basis of many cancer therapies, and tumor-specific T cells are the principal effectors of successful antitumor immunotherapies. Here we show that induction of tumor-cell apoptosis by an agonistic monoclonal antibody to DR5, the apoptosis-inducing receptor for TNF-related apoptosis-inducing ligand (TRAIL), combined with T-cell activation by agonistic monoclonal antibodies to the costimulatory molecules CD40 and CD137, potently and rapidly stimulated tumor-specific effector CD8+ T cells capable of eradicating preestablished tumors. Primary fibrosarcomas initiated with the carcinogen 3-methylcholanthrene (MCA), multiorgan metastases and a primary tumor containing as many as 90% tumor cells resistant to DR5-specific monoclonal antibody were rejected without apparent toxicity or induction of autoimmunity. This combination therapy of three monoclonal antibodies (trimAb) rapidly induced tumor-specific CD8+ T cells producing interferon (IFN)-gamma in the tumor-draining lymph node, consistent with a crucial requirement for CD8+ T cells and IFN-gamma in the tumor rejection process. These results in mice indicate that a rational monoclonal antibody-based therapy that both causes tumor-cell apoptosis through DR5 and activates T cells may be an effective strategy for cancer immunotherapy in humans.

Animals↗