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Biomedical subjects

M Koshiba

Publications and source records attributed to M Koshiba.

At least 19 recordsLinked to original sources
PubMed2004

Modification of cytokine milieu by A2A adenosine receptor signaling--possible application for inflammatory diseases.

Pro-inflammatory cytokine TNF-alpha (TNF) production from in vitro lipopolysaccharide (LPS)-stimulated human peripheral blood CD14+ cells (PB-CD14) was inhibited by A2A adenosine receptor (AdoR) (A2AR) or beta2 adrenergic receptor (ADR) (beta2R) signaling in a concentration-dependent manner. These inhibitory effects were presumably mediated by the increase in intracellular cAMP. Furthermore A2AR agonist and beta2R agonist synergistically inhibited the TNF production of LPS-stimulated PB-CD14 cells. These results suggest that the anti-inflammatory effect of extracellular adenosine is, at least in part, due to the modification of the cytokine milieu via A2A signaling, and that the targeting of both A2AR and beta2R may have strong therapeutic potential for the inflammatory diseases.

Anti-Inflammatory Agents↗
PubMed2003

Association of the TAP2*Bky2 allele with presence of SS-A/Ro and other autoantibodies in Japanese patients with systemic lupus erythematosus.

We previously reported that a new allele of transporter associated with antigen processing (TAP) 2 gene, TAP2*Bky2 (Val577), was significantly increased in Japanese patients with Sjögren's syndrome (SS) and had a strong association with SS-A/Ro antibody production. In the present study, it was investigated whether the association of TAP2*Bky2 with SS-A/Ro antibody production was also found in Japanese patients with systemic lupus erythematosus (SLE). Polymorphisms of the TAP1 and TAP2 genes were determined in 114 Japanese SLE patients by the polymerase chain reaction-single-stranded conformation polymorphism (PCR-SSCP) method. The allele frequencies of the TAP1 and TAP2 genes in SLE patients were not significantly different from those in controls, although the allele frequency of TAP2*Bky2 was slightly higher in SLE patients than in healthy control subjects (9.2% vs 5.5%, P = 0.126). The allele frequency of TAP2*Bky2 was significantly higher in SLE patients with oral ulcers than in those without. It was noteworthy that TAP2*Bky2 was significantly associated with the appearance of not only SS-A/Ro antibody but also SS-B/La, nRNP, and Sm antibodies in the patients. The association of TAP2*Bky2 was found with the antibody production to both 60 and 52kDa SS-A/Ro antigens. As TAP2*Bky2 had a strong linkage disequilibrium with DRB1*08032, TAP2*Bky2 or its haplotype with DRB1*08032 may be involved in SS-A/Ro antibody production not only in SS but also SLE patients, indicating that TAP2*Bky2 may be a susceptible gene not only to the disease of SS but also to the SS-A/Ro autoantibody production.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Extraction of all coefficients of coupled-mode equations for natural, single-phase, unidirectional SAW transducers from dispersion characteristics computed by hybrid finite element method.

A numerical method is presented for determining all of the coefficients of coupled-mode equations for natural, single-phase, unidirectional SAW transducers (NSP-UDTs). Substituting numerical results computed by the hybrid finite element method for infinite NSPUDTs with shorted and open electric ports into several relations derived from the coupled-mode theory, we can determine all of the coefficients. Specifically, the edge frequencies of a stop-band and the static capacitances yield the self-coupling coefficients and the amplitudes of mutual-coupling and transduction coefficients, and the electric potential standing wave on the substrate surface, which can be derived from the projection of the standing wave distributions of particle displacements and electric potential in the whole substrate onto the set of those predicted by the coupled-mode theory, yields the phases of mutual-coupling and transduction coefficients. NSPUDTs on ST-25 degrees X quartz, Y-51.25 degrees Z LiTaO3, and 50 degrees Y-25 degrees X La3Ga5SiO14 substrates are investigated. Our results agree well with the earlier experimental ones.

Journal Article↗
PubMed2001

Detection of antinuclear antibodies by use of an enzyme immunoassay with nuclear HEp-2 cell extract and recombinant antigens: comparison with immunofluorescence assay in 307 patients.

BACKGROUND: A new enzyme immunoassay (EIA) for automated detection of antinuclear antibodies (ANAs) uses a mixture of HEp-2 cell extracts and multiple recombinant nuclear antigens immobilized on beads. We compared this EIA and an immunofluorescence (IF) assay in a large group of patients and controls. METHODS: We studied 492 healthy individuals and 307 patients with connective tissue diseases (CTDs). Sera were tested by an automated EIA (COBAS Core HEp2 ANA EIA; Roche Diagnostics) and IF. Samples were also tested for eight disease-specific antibodies, including antibodies against U1RNP, Sm, SSA/Ro, SSB/La, Scl-70, Jo-1, dsDNA, and centromere. RESULTS: Areas under ROC curves for the EIA were greater than (P = 0.008-0.012) or numerically identical to areas for the IF method for each of six CTDs studied. ROC areas for EIA were 0.98 (95% confidence interval, 0.95-0.99), 0.99 (0.96-1.00), and 0.99 (0.98-1.00) in systemic lupus erythematosus (n = 111), systemic sclerosis (n = 39), and mixed connective tissue disease (n = 33), respectively. For all 258 CTD patients with conditions other than rheumatoid arthritis (RA), the sensitivity and specificity of the IF method at a cutoff dilution of 1:40 were 92% and 65%, respectively, vs 93% and 79% for the EIA at a cutoff of 0.6. For the IF method at a cutoff dilution of 1:160, sensitivity and specificity were 81% and 87%, respectively, vs 84% and 94%, respectively, for the EIA at a cutoff of 0.9. For 207 sera containing at least one of eight disease-specific ANAs, positivities for the EIA and the IF method were 97.1% and 97.6%, respectively, at cutoffs of 0.6 and 1:40 (P = 0.76). CONCLUSIONS: An EIA that can be performed by a fully automated instrument distinguishes CTDs (except RA) from healthy individuals with both higher sensitivity and specificity than the IF method when the cutoff index was set at 0.9. Moreover, it can be used to exclude the presence of disease-specific ANAs by setting the cutoff index at 0.6 with almost the same efficacy as the IF method.

Adolescent↗
PubMed2001

Creatine kinase gene mutation in a patient with muscle creatine kinase deficiency.

BACKGROUND: We describe a 56-year-old woman admitted to the hospital with a diagnosis of acute myocardial infarction without an increase of serum creatine kinase (CK) activity during her clinical course. She died on the 11th hospital day, and the diagnosis was confirmed by autopsy. The patient had had no previous muscular symptoms. METHODS: Expression of the CK-muscle (CK-M) protein in cardiac tissue was examined by immunoblotting and immunochemical staining. CK-M mRNA expression was estimated by semiquantitative reverse transcription-PCR. Gene structure of CK-M was determined by Southern blotting and direct sequencing of 2251 bp. Existence of a point mutation in the CK-M gene was examined by restriction fragment length polymorphism analysis of PCR products (PCR-RFLP) in the patient and in 108 controls. RESULTS: CK-M protein in the myocardial tissue of the patient was substantially lower (103 +/- 7 ng/mg protein) than in control myocardial tissue (35 800 +/- 2860 ng/mg protein). Immunoreactive CK-M in the patient tissue sample was 0.3% of the value for the control sample. CK-M mRNA was 53-fold less in the patient sample compared with the control. This very low expression of CK-M mRNA was considered to be the primary reason for CK-M deficiency. Direct sequencing revealed a point mutation at residue 54 in exon 2, which was specific for the patient. No other abnormalities were found in the CK-M gene of the patient. CONCLUSIONS: This report identifies a molecular abnormality in human CK deficiency and discusses the physiologic relevance of CK-M.

Blotting, Southern↗
PubMed1999

Association of the glutathione S-transferase M1 homozygous null genotype with susceptibility to Sjögren's syndrome in Japanese individuals.

OBJECTIVE: To investigate the role of polymorphisms of the glutathione S-transferase M1 (GSTM1) and GSTT1 genes in determining susceptibility to Sjögren's syndrome (SS) and autoantibody production. METHODS: Polymorphisms of the GSTM1 and GSTT1 genes in 106 Japanese patients with primary SS and in 143 healthy controls were analyzed by polymerase chain reaction. RESULTS: Frequency of the GSTM1 homozygous null genotype was significantly increased in SS patients compared with controls (57.5% versus 44.1%; P = 0.035). Moreover, a significantly greater frequency of SSA antibodies was found among SS patients with the GSTM1 null genotype than among those with the GSTM1 non-null genotype (P = 0.0013). Frequency of the GSTT1 polymorphism was not different between SS patients and controls. CONCLUSION: The GSTM1 homozygous null genotype could be a genetic factor that determines susceptibility to SS and may be involved in SSA antibody production.

Adult↗
PubMed1999

Patterns of A2A extracellular adenosine receptor expression in different functional subsets of human peripheral T cells. Flow cytometry studies with anti-A2A receptor monoclonal antibodies.

Signaling through A2A adenosine receptors (A2AR) regulates T lymphocyte expansion and modulates T cell receptor (TCR)-mediated effector functions in vitro. To understand the role of A2ARs in the regulation of immune response, we investigated the expression levels of this receptor in different functional lymphocyte subsets. Monoclonal anti-A2AR antibody was used to develop a flow cytometric assay to quantify the expression A2ARs on lymphocytes. We report that detectable levels of expression of A2ARs are much higher among T cells than B cells. More CD4(+) than CD8(+) T cells express A2ARs, but activation of T cells increases A2AR expression, predominantly in CD8(+) T cells. No significant differences were found in the proportion of A2AR+ cells between CD8(low) and CD8(high) T cells or between TCR/CD3(low) and TCR/CD3(high) T cells. Studies of T helper cell subsets (TH1 and TH2) reveal that lymphokine-producing cells are much more likely to express A2ARs than are cells that do not produce lymphokines. These results suggest that A2ARs are variably expressed on T cell subsets and may regulate cytokine production in activated T lymphocytes.

Antibodies, Monoclonal↗
PubMed1997

Memory of extracellular adenosine A2A purinergic receptor-mediated signaling in murine T cells.

Accumulation of extracellular and intracellular adenosine (Ado) under hypoxic conditions or in the absence of adenosine deaminase results in lymphocyte depletion and in severe combined immunodeficiency, which are currently explained by direct intracellular lymphotoxicity of Ado metabolites. In support of the alternative, "signaling" mechanism, we show that extracellular Ado (extAdo) suppresses all tested T cell receptor (TCR)-triggered effector functions of T lymphocytes including the TCR-triggered FasL mRNA up-regulation in cytotoxic T lymphocytes. Strong evidence against the intracellular lymphotoxicity of Ado (and in support of the signaling model) is provided by abrogation of TCR-triggered growth inhibition in Ado-exposed T cells. The brief exposure to Ado was sufficient to observe inhibition of TCR-triggered effector functions. The "memory" of T cells to exposure to extAdo is best explained by sustained increases in cAMP. Selective agonist (CGS21680) and antagonist (ZM241385) of A2A adenosine receptor were used in functional assays and cDNA probes for different sybtypes of adenosine receptors were used in Northern blot studies. A2A receptors are identified as the predominantly expressed subtype of Gs-coupled Ado receptors in T cells. The demonstration of cross-talk between the A2A receptors and TCR in both directions support the possible role of A2A receptors in mechanisms of extAdo-mediated immunosuppression in vivo under adenosine deaminase deficiency and hypoxic conditions in, e.g., solid tumors.

Adenosine↗
PubMed1997

Role of A2a extracellular adenosine receptor-mediated signaling in adenosine-mediated inhibition of T-cell activation and expansion.

Accumulation of adenosine and of deoxyadenosine in the absence of adenosine deaminase activity (ADA) activity results in lymphocyte depletion and in severe combined immunodeficiency (ADA SCID), which is currently explained by direct cell death-causing effects of intracellular products of adenosine metabolism. We explored the alternative mechanisms of peripheral T-cell depletion as due to inhibition of T-cell expansion by extracellular adenosine-mediated signaling through purinergic receptors. The strong inhibition of the T-cell receptor (TCR)-triggered proliferation and of upregulation of interleukin-2 receptor alpha chain (CD25) molecules, but not the direct lymphotoxicity, were observed at low concentrations of extracellular adenosine. These effects of extracellular adenosine (Ado) are likely to be mediated by A2a receptor-mediated signaling rather than by intracellular toxicity of adenosine catabolites, because (1) poorly metabolized adenosine analogs cause the accumulation of cAMP and strong inhibition of TCR-triggered CD25 upregulation; (2) the A2a, but not the A1 or A3, receptors are the major expressed and functionally coupled adenosine receptors in mouse peripheral T and B lymphocytes, and the adenosine-induced cAMP accumulation in lymphocytes correlates with the expression of A2a receptors; (3) the specific agonist of A2a receptor, CGS21680, induces increases in [cAMP]i in lymphocytes, whereas the specific antagonist of A2a receptor, CSC, inhibits the effects of Ado and CGS21680; and (4) the increases in [cAMP]i mimic the adenosine-induced inhibition of TCR-triggered CD25 upregulation and splenocyte proliferation. These studies suggest the possible role of adenosine receptors in the regulation of lymphocyte expansion and point to the downregulation of A2a purinergic receptors on T cells as a potentially attractive pharmacologic target.

Adenosine↗
PubMed1997

Effects of extracellular ATP and adenosine on different thymocyte subsets: possible role of ATP-gated channels and G protein-coupled purinergic receptor.

To explore the possible role of purinergic receptors in thymocyte development and in pathogenesis of adenosine deaminase SCID, we studied effects of extracellular adenosine triphosphate (ATP(ext)) and adenosine on TCR- and steroid hormone-triggered processes in mouse thymocytes. Reverse transcriptase-PCR analysis confirms the mRNA expression of several types of purinergic receptors, while the functioning of ATP receptors in thymocytes is reflected by ATP(ext)-induced intracellular calcium increases and by thymocyte subset-specific sensitivity to the effects of ATP(ext) and adenosine. Only ATP(ext), but not the ATP catabolites, adenosine, dexamethasone, or TCR cross-linking, was efficient in triggering rapid protein synthesis independent lysis of CD4+8- thymocytes and peripheral CD4+ T cells. In contrast, extracellular adenosine specifically induced the apoptosis of CD4+8+ thymocytes. ATP(ext) also induced a slower process of DNA fragmentation and protein synthesis-dependent apoptosis in all thymocyte subsets. ATP(ext) had an additive effect with TCR cross-linking in the induction of thymocyte death, but, unexpectedly, the effects of ATP(ext) at high concentration were antagonistic to steroid-induced apoptosis. Described here, the properties of ATP(ext) and adenosine are consistent with their involvement in the regulation of T cell development due to differential expression and signaling through purinergic receptors in different thymocyte subsets. The possible role of purinergic receptor signaling in T cell differentiation and adenosine deaminase SCID is discussed.

Adenosine↗
PubMed1997

Transient up-regulation of P2Y2 nucleotide receptor mRNA expression is an immediate early gene response in activated thymocytes.

In studies designed to understand the roles of P2 nucleotide receptors in differentiation of T lymphocytes, we observed a transient and protein synthesis-independent enhancement of mRNA expression for the G protein-coupled P2Y2 receptor in mouse thymocytes after the addition of steroid hormone or T cell receptor (TCR) crosslinking by anti-TCR mAb. Conversely, dexamethasone-induced increases in mRNA expression for the ligand-gated ion channel P2X1 receptor was detected in rat, but not mouse, thymocytes, raising questions about the previously suggested role of P2X1 receptors in thymocyte apoptosis. Flow cytometry analysis of thymocyte subsets excluded the possibility that the observed increases in P2Y2 receptor mRNA expression were due to the enrichment of steroid-treated cells with an P2Y2 mRNA-rich thymocyte subset. Triggering of TCR-mediated intracellular signaling pathways through crosslinking of TCR or by addition of phorbol ester and Ca2+ ionophore also resulted in the up-regulation of P2Y2, but not P2X1, receptor mRNA. It is proposed that the rapid increase of P2Y2 receptor mRNA expression could be a common early event in responses of T cells to different activating stimuli. Taken together with the recently discovered ability of nucleotide receptor-initiated signaling to antagonize or enhance the effects of TCR crosslinking or steroids on thymocytes, the observed rapid up-regulation of P2Y2 receptor mRNA expression may reflect an immediate early gene response where newly expressed cell surface nucleotide receptors provide regulatory feedback signaling from extracellular ATP in the T cell differentiation process.

Animals↗
PubMed1996

Activation of A3 adenosine receptors on human eosinophils elevates intracellular calcium.

Adenosine (ADO) is a potent bronchoconstrictor in allergic patients and has been shown to increase the release of histamine from human lung tissues. Antagonists of ADO A1 and A2A receptors are not effective in attenuating these effects. Therefore, involvement of ADO A3 receptors in the bronchoconstrictor and/or inflammatory effects have to be considered. Eosinophils also play a pivotal role in allergic diseases such as asthma, thus it is natural to consider a link between the A3 receptor and eosinophils. Human peripheral blood eosinophils express the ADO A3 receptor as indicated by detection of the transcript for A3 receptors in polymerase chain reaction-amplified cDNA derived from the cells. A3 receptors on eosinophil membranes were characterized using the A3 receptor agonist radioligand 125I-labeled AB-MECA, which yielded Bmax and Kd values of 1.31 pmol/mg protein and 3.19 nmol/L, respectively. Treatment of eosinophils with the highly potent and selective A3 receptor agonist CI-IB-MECA clearly induced Ca2+ release from intracellular Ca2+ pools followed by Ca2+ influx, suggesting the presence of phospholipase C-coupled A3 receptors. In contrast, the ADO receptor agonists CPA and CGS 21680, selective for A1 and A2A receptors, respectively, at concentrations of < or = 30 mumol/ L did not elevate the intracellular Ca2+ level. These results attest to the existence of ADO A3 receptors on eosinophils and suggest that ADO stimulates these cells to release Ca2+ from intracellular stores via the activation of A3 receptors.

Adenosine↗
PubMed1996

Induction of apoptosis in HL-60 human promyelocytic leukemia cells by adenosine A(3) receptor agonists.

The effects of adenosine (ADO) analogs on cells of the human promyelocytic HL-60 line were examined. ADO A(3) receptor agonists, N(6)-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide (IB-MECA, 30-60 microM) and 2-chloro-N(6)-(3-iodobenzyl)adenosine-5'-N-methyluronamide (CI-IB-MECA, 10-30 microM) induced apoptotic cell death. In contrast, neither an A(1)/A(2) antagonist (XAC) nor other selective ADO receptor agonists (CPA, NECA and CGS21680) induced apoptosis at concentrations of <30 microM. Both IB-MECA and CI-IB-MECA significantly induced Ca(2+) release from intracellular Ca(2+) pools followed by Ca(2+) influx, suggesting the presence of phospholipase C-coupled ADO A(3) receptors on HL-60 cells. This was further supported by the presence of mRNA of ADO A3 receptor in the cells. These results suggest that activation of ADO A(3) receptors is responsible for the ADO-induced apoptosis in HL-60 cells and could be of potential therapeutic value in the treatment of leukemia.

Adenosine↗