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Biomedical subjects

Miyuki Azuma

Publications and source records attributed to Miyuki Azuma.

At least 19 recordsLinked to original sources
PubMed2006

Overexpression of B7-H1 (PD-L1) significantly associates with tumor grade and postoperative prognosis in human urothelial cancers.

PURPOSE: The programmed death-1 (PD-1)/B7-H1 (also called PD-L1) pathway negatively regulates T cell activation and has been suggested to play an important role in regulating antitumor host immunity. To investigate the clinical significance of B7-H1 expression to the tumor grade and postoperative prognosis of patients with urothelial cancer, we analyzed the relationship between B7-H1 expression and various clinicopathological features and postoperative prognosis. EXPERIMENTAL DESIGN: Sixty-five urothelial cancer cases were examined. B7-H1 expression in tumors and the numbers and phenotypes of tumor-infiltrating lymphocytes were evaluated by immunohistochemistry and flow cytometry. RESULTS: A substantial expression of B7-H1 was observed in all urothelial cancers investigated. Tumor specimens from patients with higher WHO grade or primary tumor classifications showed significantly higher percentages of tumor-associated B7-H1. Tumor-associated B7-H1 expression was significantly associated with a high frequency of postoperative recurrence and poor survival rate. Furthermore, multivariate analysis indicated that tumor-associated B7-H1 was more significant prognostic factor than WHO grade. CONCLUSIONS: Our results demonstrate that the aberrant expression of B7-H1 in urothelial cancer is associated with aggressive tumors, suggesting a regulatory role of tumor-associated B7-H1 in antitumor immunity. Therefore, the manipulation of tumor-associated B7-H1 may become a beneficial target for immunotherapy in human urothelial cancer.

Antigens, CD↗
PubMed2006

Insulin-induced remission in new-onset NOD mice is maintained by the PD-1-PD-L1 pathway.

The past decade has seen a significant increase in the number of potentially tolerogenic therapies for treatment of new-onset diabetes. However, most treatments are antigen nonspecific, and the mechanism for the maintenance of long-term tolerance remains unclear. In this study, we developed an antigen-specific therapy, insulin-coupled antigen-presenting cells, to treat diabetes in nonobese diabetic mice after disease onset. Using this approach, we demonstrate disease remission, inhibition of pathogenic T cell proliferation, decreased cytokine production, and induction of anergy. Moreover, we show that robust long-term tolerance depends on the programmed death 1 (PD-1)-programmed death ligand (PD-L)1 pathway, not the distinct cytotoxic T lymphocyte-associated antigen 4 pathway. Anti-PD-1 and anti-PD-L1, but not anti-PD-L2, reversed tolerance weeks after tolerogenic therapy by promoting antigen-specific T cell proliferation and inflammatory cytokine production directly in infiltrated tissues. PD-1-PD-L1 blockade did not limit T regulatory cell activity, suggesting direct effects on pathogenic T cells. Finally, we describe a critical role for PD-1-PD-L1 in another powerful immunotherapy model using anti-CD3, suggesting that PD-1-PD-L1 interactions form part of a common pathway to selectively maintain tolerance within the target tissues.

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

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

Multipotency of CD11bhighGr-1+ immature myeloid cells accumulating in oral squamous cell carcinoma-bearing mice.

We demonstrated the accumulation of CD11b(high)Gr-1(+) cells in a murine model of squamous cell carcinoma (SCC). Inoculation of NR-S1K cells derived from oral SCC induced a rapid and clear accumulation of CD11b(high)Gr-1(+) cells in secondary lymphoid organs as well as in peripheral blood. Phenotypic and morphological analyses revealed that these CD11b(high)Gr-1(+) cells were not lymphoid lineage cells, mature dendritic cells, macrophages, or granulocytes. Although the freshly isolated CD11b(high) cells lacked antigen-presenting capacity, they acquired a potent antigen-presenting capacity that included the induction of MHC class II after culture with GM-CSF and IL-4 in vitro. These results suggest that CD11b(high) cells that accumulate in tumor-bearing hosts are immature myeloid cells, but have considerable potential to differentiate into potent antigen-presenting cells under appropriate culture conditions. The use of in vitro differentiated CD11b(high) cells may be a potential strategy for obtaining patient-matched dendritic cells for tumor immunotherapy.

Animals↗
PubMed2006

The existence of CD11c+ sentinel and F4/80+ interstitial dendritic cells in dental pulp and their dynamics and functional properties.

Dental caries and pulpitis are the most common bacterial infections in humans. However, the immune responses against bacterial stimulation in dental pulp that is bounded by special hard tissues are poorly understood. We examined the initial immune responses in mouse dental pulp after cusp trimming and acid treatment. Using fluorescence immunohistochemistry, two distinct cell populations were identified in the intact pulp; CD11c+F4/80- and CD11c-F4/80+ cells. CD11c+F4/80- cells were localized in the pulp-dentin (P-D) border of the central pulp beneath the dental fissure, whereas CD11c-F4/80+ cells with dendritic morphology were distributed in the perivascular region of the inner pulp and the sub-odontoblastic layer. CD11c+F4/80- cells, but not CD11c-F4/80+ cells, constitutively expressed toll-like receptors 2 and 4 and CD205, and migrated to the P-D border of the treated side within 2 h after the treatment. In parallel, some of the F4/80+ cells migrated to the inner pulp of the treated side, increased in size and enhanced CD86 expression. At 24 h, the CD86+ cells with high fluorescence intensity had disappeared entirely from the pulp. Concurrently, CD86high cells expressing intermediate levels of CD11c and high levels of MHC class II and F4/80, assessed by using flow cytometry, increased significantly in the regional lymph nodes, suggesting migration of these cells from the dental pulp. Our results are the first to demonstrate the existence of at least two types of dendritic cells (DCs) in dental pulp. The CD11c+ sentinel and F4/80+ interstitial DCs might have distinct territories and unique roles in responding to external stimuli via the dentinal tubules.

Animals↗
PubMed2006

Expression and function of the B and T lymphocyte attenuator (BTLA/CD272) on human T cells.

Co-signal receptors provide crucial activating or attenuating signals for T cells. The B and T lymphocyte attenuator (BTLA/CD272) is a third member of co-inhibitory receptors, which belongs to the CD28 immunoglobulin-superfamily. Using monoclonal antibodies (mAbs) against human BTLA, we show that BTLA is constitutively expressed on most CD4+ and CD8+ T cells and its expression progressively decreases upon T cell activation. Polarized Th1 and Th2 cells contained both BTLA-positive and BTLA-negative populations, but the extended culture diminished BTLA expression. Cross-linking BTLA with an agonistic mAb inhibited T cell proliferation and the production of the cytokines IFN-gamma and IL-10 in response to anti-CD3 stimulation. BTLA-mediated inhibition of T cell activation occurred during both primary CD4+ T cell responses and secondary CD4+ and CD8+ T cell responses, suggesting that BTLA ligation sends a constitutive "off" signal to T cells and thus might play an important role in the maintenance of T cell tolerance.

Antibodies, Monoclonal↗
PubMed2006

Differential role of programmed death-ligand 1 [corrected] and programmed death-ligand 2 [corrected] in regulating the susceptibility and chronic progression of experimental autoimmune encephalomyelitis.

Programmed death-1 (PD-1) is a negative costimulatory molecule, and blocking the interaction of PD-1 with its ligands, PD-L1 (B7-H1) and PD-L2 (B7-DC), enhances autoimmune disease in several animal models. We have studied the role of PD-1 ligands in disease susceptibility and chronic progression in experimental autoimmune encephalomyelitis (EAE). In BALB/c mice immunized with myelin oligodendrocyte glycoprotein (MOG) peptide 35-55, PD-L1 but not PD-L2 blockade significantly increased EAE incidence. In B10.S mice immunized with myelin proteolipid protein (PLP) peptide 139-151, both PD-L1 and PD-L2 blockade markedly enhanced EAE severity. In prediabetic NOD mice immunized with PLP48-70, PD-L2 blockade worsened EAE but did not induce diabetes, whereas PD-L1 blockade precipitated diabetes but did not worsen EAE, suggesting different regulatory roles of these two ligands in EAE and diabetes. B6 mice immunized with MOG35-55 developed chronic persistent EAE, and PD-L2 blockade in the chronic phase exacerbated EAE, whereas PD-L1 blockade did not. In contrast, SJL/J mice immunized with PLP139-151 developed chronic relapsing-remitting EAE, and only PD-L1 blockade during remission precipitated EAE relapse. The strain-specific effects of PD-1 ligand blockade did not correlate with the expression of PD-L1 and PD-L2 on dendritic cells and macrophages in lymphoid tissue, or on inflammatory cells in the CNS. However, EAE enhancement is correlated with less prominent Th2 cytokine induction after specific PD-1 ligand blockade. In conclusion, PD-L1 and PD-L2 differentially regulate the susceptibility and chronic progression of EAE in a strain-specific manner.

Animals↗
PubMed2005

Predominant expression of B7-H1 and its immunoregulatory roles in oral squamous cell carcinoma.

We examined cell surface expression of five B7 costimulatory molecules (B7-H1, B7-DC, B7h, CD80 and CD86) in human oral squamous cell carcinoma (SCC) lines. Most human SCC cell lines expressed various levels of B7-H1 and B7-DC. Their expression was further upregulated by interferon (IFN)-gamma stimulation. Immunohistochemical staining revealed substantial and predominant expression of B7-H1 on human primary oral SCC. A murine SCC line, NR-S1, neither expressed B7-H1 nor B7-DC, but induced B7-H1 by IFN-gamma stimulation in culture and the inoculation in vivo. Although NR-S1 tumors grew progressively in immunocompetent syngeneic mice, the administration of blocking anti-B7-Hl or anti-PD-1 mAb significantly inhibited the tumor growth, suggesting the negative regulation of host immune responses by the PD-1:B7-H1 pathway. Our results demonstrate that B7-H1 is predominantly induced on oral SCC within the B7 family molecules. A successful inhibition of tumor growth by blockade of the PD-1:B7-H1 pathway may implicate a new approach for immunotherapy of oral SCC.

Animals↗
PubMed2005

Blockade of B7-H1 on macrophages suppresses CD4+ T cell proliferation by augmenting IFN-gamma-induced nitric oxide production.

PD-1 is an immunoinhibitory receptor that belongs to the CD28/CTLA-4 family. B7-H1 (PD-L1) and B7-DC (PD-L2), which belong to the B7 family, have been identified as ligands for PD-1. Paradoxically, it has been reported that both B7-H1 and B7-DC co-stimulate or inhibit T cell proliferation and cytokine production. To determine the role of B7-H1 and B7-DC in T cell-APC interactions, we examined the contribution of B7-H1 and B7-DC to CD4+ T cell activation by B cells, dendritic cells, and macrophages using anti-B7-H1, anti-B7-DC, and anti-PD-1 blocking mAbs. Anti-B7-H1 mAb and its Fab markedly inhibited the proliferation of anti-CD3-stimulated naive CD4+ T cells, but enhanced IL-2 and IFN-gamma production in the presence of macrophages. The inhibition of T cell proliferation by anti-B7-H1 mAb was abolished by neutralizing anti-IFN-gamma mAb. Coculture of CD4+ T cells and macrophages from IFN-gamma-deficient or wild-type mice showed that CD4+ T cell-derived IFN-gamma was mainly responsible for the inhibition of CD4+ T cell proliferation. Anti-B7-H1 mAb induced IFN-gamma-mediated production of NO by macrophages, and inducible NO synthase inhibitors abrogated the inhibition of CD4+ T cell proliferation by anti-B7-H1 mAb. These results indicated that the inhibition of T cell proliferation by anti-B7-H1 mAb was due to enhanced IFN-gamma production, which augmented NO production by macrophages, suggesting a critical role for B7-H1 on macrophages in regulating IFN-gamma production by naive CD4+ T cells and, hence, NO production by macrophages.

Animals↗
PubMed2005

A critical role for the programmed death ligand 1 in fetomaternal tolerance.

Fetal survival during gestation implies that tolerance mechanisms suppress the maternal immune response to paternally inherited alloantigens. Here we show that the inhibitory T cell costimulatory molecule, programmed death ligand 1 (PDL1), has an important role in conferring fetomaternal tolerance in an allogeneic pregnancy model. Blockade of PDL1 signaling during murine pregnancy resulted in increased rejection rates of allogeneic concepti but not syngeneic concepti. Fetal rejection was T cell- but not B cell-dependent because PDL1-specific antibody treatment caused fetal rejection in B cell-deficient but not in RAG-1-deficient females. Blockade of PDL1 also resulted in a significant increase in the frequency of IFN-gamma-producing lymphocytes in response to alloantigen in an ELISPOT assay and higher IFN-gamma levels in placental homogenates by ELISA. Finally, PDL1-deficient females exhibited decreased allogeneic fetal survival rates as compared with littermate and heterozygote controls and showed evidence of expansion of T helper type 1 immune responses in vivo. These results provide the first evidence that PDL1 is involved in fetomaternal tolerance.

Animals↗
PubMed2005

Analysis of the role of negative T cell costimulatory pathways in CD4 and CD8 T cell-mediated alloimmune responses in vivo.

Negative costimulatory signals mediated via cell surface molecules such as CTLA-4 and programmed death 1 (PD-1) play a critical role in down-modulating immune responses and maintaining peripheral tolerance. However, their role in alloimmune responses remains unclear. This study examined the role of these inhibitory pathways in regulating CD28-dependent and CD28-independent CD4 and CD8 alloreactive T cells in vivo. CTLA-4 blockade accelerated graft rejection in C57BL/6 wild-type recipients and in a proportion of CD4(-/-) but not CD8(-/-) recipients of BALB/c hearts. The same treatment led to prompt rejection in CD28(-/-) and a smaller proportion of CD4(-/-)CD28(-/-) mice with no effect in CD8(-/-)CD28(-/-) recipients. These results indicate that the CTLA-4:B7 pathway provides a negative signal to alloreactive CD8(+) T cells, particularly in the presence of CD28 costimulation. In contrast, PD-1 blockade led to accelerated rejection of heart allografts only in CD28(-/-) and CD8(-/-)CD28(-/-) recipients. Interestingly, PD-1 ligand (PD-L1) blockade led to accelerated rejection in wild-type mice and in all recipients lacking CD28 costimulation. This effect was accompanied by expansion of IFN-gamma-producing alloreactive T cells and enhanced generation of effector T cells in rejecting allograft recipients. Thus, the PD-1:PD-L1 pathway down-regulates alloreactive CD4 T cells, particularly in the absence of CD28 costimulation. The differential effects of PD-1 vs PD-L1 blockade support the possible existence of a new receptor other than PD-1 for negative signaling through PD-L1. Furthermore, PD-1:PD-L1 pathway can regulate alloimmune responses independent of an intact CD28/CTLA-4:B7 pathway. Harnessing physiological mechanisms that regulate alloimmunity should lead to development of novel strategies to induce durable and reproducible transplantation tolerance.

Animals↗
PubMed2005

Expression of B7-H1 and B7-DC on the airway epithelium is enhanced by double-stranded RNA.

Viral infection in the airway provokes various immune responses, including Th1 and Th2 responses, which are partly initiated by double-stranded RNA (dsRNA), a viral product for its replication. B7-H1 (PD-L1) and B7-DC (PD-L2) are B7-family molecules that bind to programmed death-1 (PD-1) on lymphocytes and are implicated in peripheral tolerance. We investigated the effect of dsRNA on the expression of B7-H1 and B7-DC on airway epithelial cell lines. B7-H1 and B7-DC were constitutively expressed on the cells, and their expression was profoundly upregulated by stimulation with an analog of viral dsRNA, polyinosinic-polycytidylic acid. B7-H1 and B7-DC were also upregulated by stimulation with IFN-gamma, IL-13, and the supernatant from T cell clones. A relatively high concentration of dexamethasone (1 microM) was required to suppress the upregulation of B7-H1 or B7-DC. These results suggest that epithelial B7-H1 and B7-DC play a role in virus-associated immune responses in the airways.

Antigens, CD↗
PubMed2005

Clinical significance of programmed death-1 ligand-1 and programmed death-1 ligand-2 expression in human esophageal cancer.

PURPOSE: The negative regulatory programmed death-1/programmed death-1 ligand (PD-1/PD-L) pathway in T-cell activation has been suggested to play an important role in tumor evasion from host immunity. In this study, we investigated the expression of PD-L1 and PD-L2 in human esophageal cancer to define their clinical significance in patients' prognosis after surgery. EXPERIMENTAL DESIGN: PD-L1 and PD-L2 gene expression was evaluated in 41 esophagectomy patients by real-time quantitative PCR. The protein expression was also evaluated with newly generated monoclonal antibodies that recognize human PD-L1 (MIH1) and PD-L2 (MIH18). RESULTS: The protein and the mRNA levels of determination by immunohistochemistry and real-time quantitative PCR were closely correlated. PD-L-positive patients had a significantly poorer prognosis than the negative patients. This was more pronounced in the advanced stage of tumor than in the early stage. Furthermore, multivariate analysis indicated that PD-L status was an independent prognostic factor. Although there was no significant correlation between PD-L1 expression and tumor-infiltrating T lymphocytes, PD-L2 expression was inversely correlated with tumor-infiltrating CD8(+) T cells. CONCLUSIONS: These data suggest that PD-L1 and PD-L2 status may be a new predictor of prognosis for patients with esophageal cancer and provide the rationale for developing novel immunotherapy of targeting PD-1/PD-L pathway.

Aged↗
PubMed2005

Role of the programmed death-1 pathway in regulation of alloimmune responses in vivo.

Programmed death-1 (PD-1), an inhibitory receptor up-regulated on activated T cells, has been shown to play a critical immunoregulatory role in peripheral tolerance, but its role in alloimmune responses is poorly understood. Using a novel alloreactive TCR-transgenic model system, we examined the functions of this pathway in the regulation of alloreactive CD4+ T cell responses in vivo. PD-L1, but not PD-1 or PD-L2, blockade accelerated MHC class II-mismatched skin graft (bm12 (I-Abm12) into B6 (I-Ab)) rejection in a similar manner to CTLA-4 blockade. In an adoptive transfer model system using the recently described anti-bm12 (ABM) TCR-transgenic mice directly reactive to I-Abm12, PD-1 and PD-L1 blockade enhanced T cell proliferation early in the immune response. In contrast, at a later time point preceding accelerated allograft rejection, only PD-L1 blockade enhanced T cell proliferation. In addition, PD-L1 blockade enhanced alloreactive Th1 cell differentiation. Apoptosis of alloantigen-specific T cells was inhibited significantly by PD-L1 but not PD-1 blockade, indicating that PD-1 may not be the receptor for the apoptotic effect of the PD-L1-signaling pathway. Interestingly, the effect of PD-L1 blockade was dependent on the presence of CD4+ CD25+ regulatory T cells in vivo. These data demonstrate a critical role for the PD-1 pathway, particularly PD-1/PD-L1 interactions, in the regulation of alloimmune responses in vivo.

Adoptive Transfer↗
PubMed2005

Subcutaneous injections of a TNF-alpha antagonistic peptide inhibit both inflammation and bone resorption in collagen-induced murine arthritis.

The cyclic peptide WP9QY (YCWSQYLCY), which was designed to mimic the most critical tumor necrosis factor (TNF) alpha recognition loop on type1 TNF receptor, antagonizes the effects of TNFalpha. In this study, we investigated the effects of WP9QY peptide on collagen-induced arthritis (CIA) mice to evaluate its effects on inflammatory bone destruction. DBA/1J mice were injected intradermally at the base of the tail with bovine type II collagen, emulsified in complete Freund's adjuvant on day 0 and 21. The three sets of WP9QY peptide injections (24 mg/kg x 8 times per day) were performed before the onset of paw swelling. Mice were sacrificed at day 38 and thereafter, the arthritis scores as well as radiographical and histological outcomes were assessed. WP9QY peptide inhibited CIA-induced increase in the arthritis score. Furthermore, histomorphometric analysis of the tibial epiphysis region revealed that WP9QY peptide inhibited the increase of synovial pannus infiltration and the decrease of bone volume, which were induced by the CIA. The WP9QY treatment prevented the inflammation as well as bone destruction of the joints in the CIA mice, suggesting that the administration of WP9QY peptide might be useful for developing a drug to prevent inflammatory bone destruction.

Animals↗
PubMed2004

Blockade of the interaction between PD-1 and PD-L1 accelerates graft arterial disease in cardiac allografts.

BACKGROUND: Programmed death-1 (PD-1), a member of the CD28 family, has been identified. PD-1 is involved in the negative regulation of some immune responses. We evaluated the role of PD-ligand 1 (PD-L1) in graft arterial disease (GAD) of cardiac allografts and in smooth muscle cells (SMCs). METHODS AND RESULTS: C57BL/6 murine hearts were transplanted into B6.C-H2 KhEg mice for examination of GAD. PD-L1 was expressed in SMCs of the thickened intima in the graft coronary arteries, and administration of anti-PD-L1 monoclonal antibody (mAb) enhanced the progression of GAD (luminal occlusion: 55+/-5.0% versus 9.8+/-4.3%, P<0.05). The expressions of interferon gamma (IFN-gamma) and tumor necrosis factor alpha of cardiac allografts were upregulated in response to anti-PD-L1 mAb treatment. In vitro, PD-L1 expression was induced in SMCs in response to IFN-gamma stimulation. Sensitized splenocytes increased SMC proliferation, and anti-PD-L1 mAb in combination with IFN-gamma stimulation increased this proliferation. CONCLUSIONS: The PD-L1 pathway regulates both the proliferation of SMCs and GAD. Thus, control of this interaction is a promising strategy for suppression of GAD.

Animals↗