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M Nishimura

Publications and source records attributed to M Nishimura.

At least 505 records · Page 28Linked to original sources
PubMed1997

Identification of CCR6, the specific receptor for a novel lymphocyte-directed CC chemokine LARC.

Liver and activation-regulated chemokine (LARC) is a recently identified CC chemokine that is expressed mainly in the liver. LARC functions as a selective chemoattractant for lymphocytes that express a class of receptors specifically binding to LARC with high affinity. To identifiy the receptor for LARC, we examined LARC-induced calcium mobilization in cells stably expressing five CC chemokine receptors (CCR1-CCR5) and five orphan seven-transmembrane receptors. LARC specifically induced calcium flux in K562 cells as well as 293/EBNA-1 cells stably expressing an orphan receptor GPR-CY4. LARC induced migration in 293/EBNA-1 cells stably expressing GPR-CY4 with a bi-modal dose-response curve. LARC fused with secreted alkaline phosphatase (LARC-SEAP) bound specifically to Raji cells stably expressing GPR-CY4 with a Kd of 0.9 nM. Only LARC but not five other CC chemokines (MCP-1, RANTES, MIP-1alpha, MIP-1beta, and TARC) competed with LARC-SEAP for binding to GPR-CY4. By Northern blot analysis, GPR-CY4 mRNA was expressed mainly in spleen, lymph nodes, Appendix, and fetal liver among various human tissues. Among various leukocyte subsets, GPR-CY4 mRNA was detected in lymphocytes (CD4(+) and CD8(+) T cells and B cells) but not in natural killer cells, monocytes, or granulocytes. Expression of GPR-CY4 mRNA in CD4(+) and CD8(+) T cells was strongly up-regulated by IL-2. Taken together, GPR-CY4 is the specific receptor for LARC expressed selectively on lymphocytes, and LARC is a unique functional ligand for GPR-CY4. We propose GPR-CY4 to be designated as CCR6.

B-Lymphocytes↗
PubMed1997

The T cell-directed CC chemokine TARC is a highly specific biological ligand for CC chemokine receptor 4.

Thymus and activation-regulated chemokine (TARC) is a recently identified CC chemokine that is expressed constitutively in thymus and transiently in stimulated peripheral blood mononuclear cells. TARC functions as a selective chemoattractant for T cells that express a class of receptors binding TARC with high affinity and specificity. To identify the receptor for TARC, we produced TARC as a fusion protein with secreted alkaline phosphatase (SEAP) and used it for specific binding. By stably transfecting five orphan receptors and five known CC chemokine receptors (CCR1 to -5) into K562 cells, we found that TARC-SEAP bound selectively to cells expressing CCR4. TARC-SEAP also bound to K562 cells stably expressing CCR4 with a high affinity (Kd = 0.5 nM). Only TARC and not five other CC chemokines (MCP-1 (monocyte chemoattractant protein-1), RANTES (regulated upon activation, normal T cells expressed and secreted), MIP-1alpha (macrophage inflammatory protein-1alpha), MIP-1beta, and LARC (liver and activation-regulated chemokine)) competed with TARC-SEAP for binding to CCR4. TARC but not RANTES or MIP-1alpha induced migration and calcium mobilization in 293/EBNA-1 cells stably expressing CCR4. K562 cells stably expressing CCR4 also responded to TARC in a calcium mobilization assay. Northern blot analysis revealed that CCR4 mRNA was expressed strongly in human T cell lines and peripheral blood T cells but not in B cells, natural killer cells, monocytes, or granulocytes. Taken together, TARC is a specific functional ligand for CCR4, and CCR4 is the specific receptor for TARC selectively expressed on T cells.

Alkaline Phosphatase↗
PubMed1997

Molecular cloning of a novel human CC chemokine EBI1-ligand chemokine that is a specific functional ligand for EBI1, CCR7.

By searching the expressed sequence tag (EST) data base, we identified partial cDNA sequences encoding a novel human CC chemokine. We determined the complete cDNA sequence that encodes a highly basic polypeptide of a total 98 amino acids with 20 to 30% identity to other human CC chemokines. We termed this novel chemokine from EBI1-Ligand Chemokine as ELC (see below). The ELC mRNA was most strongly expressed in the thymus and lymph nodes. Recombinant ELC protein was expressed as a fusion protein with the Flag tag (ELC-Flag). For receptor-binding assays, recombinant ELC protein fused with the secreted form of alkaline phosphatase (SEAP) was used. By stably expressing five CC chemokine receptors (CCR1 to 5) and five orphan receptors, ELC-SEAP was found to bind specifically to an orphan receptor EBI1. Only ELC-Flag, but not MCP-1, MCP-2, MCP-3, eotaxin, MIP-1alpha, MIP-1beta, RANTES (regulated on activation normal T cell expressed and secreted), thymus and activation-regulated chemokine (TARC), or liver and activation-regulated chemokine (LARC), competed with ELC-SEAP for EBI1. ELC-Flag-induced transient calcium mobilization and chemotactic responses in EBI1-transfected cells. ELC-Flag also induced chemotaxis in HUT78 cells expressing endogenous EBI1 at high levels. By somatic hybrid and radiation hybrid analyses, the gene for ELC (SCYA19) was mapped to chromosome 9p13 instead of chromosome 17q11.2 where the genes for CC chemokines are clustered. Taken together, ELC is a highly specific ligand for EBI1, which is known to be expressed in activated B and T lymphocytes and strongly up-regulated in B cells infected with Epstein-Barr virus and T cells infected with herpesvirus 6 or 7. ELC and EBI1 may thus play roles in migration and homing of normal lymphocytes, as well as in pathophysiology of lymphocytes infected with these herpesviruses. We propose EBI1 to be designated as CCR7.

Amino Acid Sequence↗
PubMed1997

An aspartic endopeptidase is involved in the breakdown of propeptides of storage proteins in protein-storage vacuoles of plants.

To understand the mechanism of the maturation of various proteins in protein-storage vacuoles, we purified a 48-kDa aspartic endopeptidase composed of 32-kDa and 16-kDa subunits from castor bean. Immunocytochemical and cell fractionation analyses of the endosperm of maturing castor bean seed showed that the aspartic endopeptidase was localized in the matrix of the protein-storage vacuoles, where a variety of seed storage proteins were also present. The amount of the aspartic endopeptidase increased at the mid-maturation stage of the seeds before accumulation of the storage proteins. To determine how the aspartic endopeptidase is responsible for maturation of seed proteins in concert with the vacuolar processing enzyme, we prepared 35S-labeled proproteins of seed proteins from the endoplasmic reticulum fraction of pulse-labeled maturing endosperm and used the authentic proproteins as substrates for in vitro processing experiments. The purified aspartic endopeptidase was unable to convert any of three endosperm proproteins, pro2S albumin, proglobulin, and proricin, into their mature sizes, while the purified vacuolar processing enzyme could convert all three proproteins. We further examined the activity of aspartic endopeptidase on the cleavage of an internal propeptide of Arabidopsis pro2S albumin, which is known to be removed post-translationally. The aspartic endopeptidase cleaved the propeptide at three sites under acidic conditions. These results suggest that aspartic endopeptidase cannot directly convert pro2S albumin into the mature form, but it may play a role in trimming the C-terminal propeptides from the subunits that are produced by the action of the vacuolar processing enzyme.

2S Albumins, Plant↗
PubMed1997

The prognosis of resected lung carcinoma associated with atypical adenomatous hyperplasia: a comparison of the prognosis of well-differentiated adenocarcinoma associated with atypical adenomatous hyperplasia and intrapulmonary metastasis.

BACKGROUND: Radiologically undetected intrapulmonary solitary nodules are sometimes found in the resected lung. When the main tumor is a well-differentiated (w/d) adenocarcinoma, especially a bronchioloalveolar carcinoma (BAC), it can be difficult to determine morphologically whether the intrapulmonary nodules are atypical adenomatous hyperplasia (AAH) or intrapulmonary metastasis (PM). The authors evaluated the accuracy of the differential diagnosis of these two lesions from the prognostic point of view. METHODS: A retrospective study was conducted of 1360 lung carcinoma patients who had undergone surgical resection. Differential diagnosis was made between AAH and PM based on the conventional histologic specimens. Their clinicopathologic features were also studied. Survival rates were compared between these two groups. RESULTS: AAH was found in 137 patients (10%) with resected lung carcinoma. The 5-year survival rates were 72.9% in Stage I, 60.6% in Stage II, 27.1% in Stage IIIA, 0% in Stage IIIB, and 0% in Stage IV. They were not significantly different from the figures for all patients in the corresponding pathologic stages. Seventy-six cases were diagnosed as w/d adenocarcinoma associated with AAH, whereas PM was found in 46 cases of w/d adenocarcinoma. The 5-year survival rates of AAH and PM differed significantly: 64.6% and 35.5%, respectively (P 0.0004). When a comparison was made between cases of pT1-2, N0 w/d adenocarcinoma, most of which were BAC, with PM (n = 22) and those with AAH (n = 52), the latter had significantly better survival (P = 0.0086). CONCLUSIONS: The prognosis of resected lung carcinoma was not affected by association with AAH. The significant difference in prognosis between AAH and PM in w/d adenocarcinoma, especially in BAC, indicates that their morphologic distinction was correctly made by conventional pathologic examination in most cases.

Adenocarcinoma↗
PubMed1997

Effects of concanavalin A on chondrocyte hypertrophy and matrix calcification.

Resting chondrocytes do not usually undergo differentiation to the hypertrophic stage and calcification. However, incubating these cells with concanavalin A resulted in 10-100-fold increases in alkaline phosphatase activity, binding of 1,25(OH)2-vitamin D3, type X collagen synthesis, 45Ca incorporation into insoluble material, and calcium content. On the other hand, other lectins tested (including wheat germ agglutinin, lentil lectin, pea lectin, phytohemagglutinin-L, and phytohemagglutinin-E) marginally affected alkaline phosphatase activity, although they activate lymphocytes. Methylmannoside reversed the effect of concanavalin A on alkaline phosphatase within 48 h. Concanavalin A did not increase alkaline phosphatase activity in articular chondrocyte cultures. In resting chondrocyte cultures, succinyl concanavalin A was as potent as concanavalin A in increasing alkaline phosphatase activity, the incorporation of [35S]sulfate, D-[3H]glucosamine, and [3H]serine into proteoglycans, and the incorporation of [3H]serine into protein, although concanavalin A, but not succinyl concanavalin A, induced a rapid change in the shape of the cells from flat to spherical. These findings suggest that concanavalin A induces a switch from the resting, to the growth-plate stage, and that this action of concanavalin A is not secondary to changes in the cytoskeleton. Chondrocytes exposed to concanavalin A may be useful as a novel model of endochondral bone formation.

Alkaline Phosphatase↗
PubMed1997

Characterization of T-cell clones derived from peripheral blood lymphocytes of a patient with transfusion-associated graft-versus-host disease: Fas-mediated killing by CD4+ and CD8+ cytotoxic T-cell clones and tumor necrosis factor beta production by CD4+ T-cell clones.

Transfusion-associated graft-versus-host disease (TA-GVHD) is one of the most serious adverse effects of blood transfusion. It is generally thought to be caused by the infused lymphocytes. Donor-derived cytotoxic T lymphocytes (CTLs) directed against the recipient's HLAs, which have escaped the recipient's immune system and are proliferating, are considered to attack recipient organs and tissues. Despite the seriousness of the disease, the precise mechanism of its development remains unclear and no definitive treatment has been developed. With the aim of developing an effective treatment, we established and characterized T-cell clones from peripheral blood lymphocytes (PBLs) of a TA-GVHD patient. Three types of clones were established. Type I clones were CD8+ and specifically lyse cells that express HLA B52. Type II clones were CD4+, specifically lysed cells that express HLA DR15, and proliferated in response to stimulation with cells that express DR15. Type III clones were also CD4+, showed no cytotoxic activity toward any HLA-expressing cells, and proliferated in response to stimulation with cells that express DR15. Furthermore, we found that the Fas/Fas-ligand (Fas-L) system is involved in the cytotoxicity of the type I and II clones and that the type III clones produce and secrete a large amount of tumor necrosis factor beta (TNFbeta) after antigen stimulation. Based on our results, these three types of clones can be classified into two categories: those that have the ability to induce GVHD directly by cytolysis and that show no cytotoxic activity and those that have the ability to cause GVHD indirectly through secretion of cytotoxic lymphokines.

Aged↗
PubMed1997

Detection of Mycoplasma in avian live virus vaccines by polymerase chain reaction.

The polymerase chain reaction (PCR) was evaluated to detect mycoplasma contamination of avian live virus vaccines. The specificity of the primers showed that 34 strains belonging to nine species of avian mycoplasma DNA could be detected. The sensitivity of PCR to detect mycoplasma DNA was 10(0.2) colony forming units (cfu) of Mycoplasma synoviae and 10(0.7) cfu of Mycoplasma gallisepticum. When M. synoviae and M. gallisepticum were spiked into several avian live virus vaccines, PCR gave a positive reaction except for the avian pox and the avian encephalomyelitis vaccines which were prepared from organ homogenates. Short-term incubation of avian encephalomyelitis vaccines improved the sensitivity of PCR to detect both M. synoviae and M. gallisepticum. Therefore, PCR, combined with the short-term incubation, were shown to be most effective in detecting mycoplasma contamination in all of avian live virus vaccines.

Animals↗
PubMed1997

Linkage map of Syrian hamster with restriction landmark genomic scanning.

We have constructed the linkage map with precise genetic analysis of the Syrian hamster, Mesocricetus auratus, according to the restriction landmark genomic scanning (RLGS) spot mapping method. Although only 3.2-6.6% of the total RLGS spots between the two strains, ACN and BIO 14.6, showed genetic variance, 572 loci were found to be polymorphic. Out of 569 RLGS loci and 3 other loci, 531 were mapped with the backcross (ACN x BIO 14.6) F1 x BIO 14.6. The cumulative map was 1111.6 cM, indicating that the spots/loci are located throughout the genome at 1.94 cM intervals on average. Thus, RLGS provides us with a rapid tool to construct the genetic map of any species, even if it has less genetic variation.

Animals↗
PubMed1997

A serine-to-proline mutation in the copper-transporting P-type ATPase gene of the macular mouse.

We have investigated the cDNA sequence of the copper-transporting P-type ATPase (Atp7a) gene of the macular mouse, a model for human Menkes disease. A point mutation (T to C) that results in substitution of proline for serine in a putative eighth transmembrane domain of the ATP7A was identified. This contrasts with abnormalities identified in the Atp7a of other mottled mouse strains: lack of expression of Atp7a mRNA in the dappled mouse, and a splicing mutation in the blotchy mouse.

Adenosine Triphosphatases↗
PubMed1997

Roles of reactive oxygen species and antioxidant enzymes in murine daunomycin-induced nephropathy.

We evaluated the roles of reactive oxygen species and intrinsic antioxidant enzymes in the development of daunomycin (DM)-induced nephropathy in mice. A single dose of DM (20 mg/kg intravenously) induced proteinuria by day 7 and the nephrotic syndrome by day 14 in DM-sensitive strain (A/J) but not in DM-resistant strain (C57BL/6J) (B6). Renal cortical lipid peroxide levels in the A/J mice significantly increased at days 2, 4, and 7 after DM injection, whereas no increase was observed in the B6 mice. The resistance to DM in B6 mice was associated with higher activities in renal cortical superoxide dismutase and glutathione peroxidase. The administration of superoxide dismutase or of dimethylthiourea significantly suppressed the DM-induced proteinuria in the A/J mice. Four days of superoxide dismutase or dimethylthiourea administration suppressed the proteinuria. These findings suggested that murine DM-nephropathy appeared to be mediated by reactive oxygen species and that intrinsic antioxidant enzyme activities may play an important role in the susceptibility to DM-induced nephropathy in mice.

Albuminuria↗
PubMed1997

Regional distribution of GABA transporter 1 (GAT1) mRNA in the rat brain: comparison with glutamic acid decarboxylase67 (GAD67) mRNA localization.

The mRNA expression of GABA transporter 1 (GAT1) was investigated in the rat brain by in situ hybridization histochemistry using an oligonucleotide probe specific to the mRNA. The pattern of GABA transporter 1 mRNA expression was well coincident with that of GABAergic terminals detected by using glutamic acid decarboxylase (GAD) as a marker. In addition, we compared the GAT1 mRNA expression with that of glutamic acid decarboxylase67 (GAD67) mRNA positive neurons. Although the expression patterns of both mRNAs were similar in most nuclei, the number of GABA transporter 1 mRNA positive cells were higher than that of GAD67 mRNA positive cells in some nuclei, such as the inferior colliculus, cerebellar nuclei and cerebral cortex. Furthermore the subthalamic nucleus and inferior olive nucleus, which lacked GAD67 mRNA expression, showed intense GABA transporter 1 mRNA expression. These data suggest that GABA transporter 1 mRNA is expressed not only in GABAergic neurons but also in non-GABAergic neurons and/or glial cells which are involved in GABAergic neurotransmission.

Animals↗
PubMed1997

Differential expression patterns of GABA transporters (GAT1-3) in the rat olfactory bulb.

The localization of GABA transporters 1-3 (GAT1-3) was investigated in the rat olfactory bulb by using in situ hybridization and immunohistochemistry. In the glomerular and the internal granular layers, GAT1 mRNA was expressed in most of periglomerular and granule cells, which are known to be GABAergic. In addition, we compared GAT1 mRNA expression with that of glutamic acid decarboxylase67 (GAD67) mRNA. The expressions were very similar in these two layers, indicating that GAT1 mRNA is mainly expressed in GABAergic neurons. However, in the external plexiform and the olfactory nerve layers, we observed more GAT1 mRNA-positive cells than GAD67 ones, suggesting that GAT1 mRNA is also expressed in non-GABAergic neurons and in glial cells. GAT3 mRNA expression was observed in small glial-like cells which might be involved in GABAergic neurotransmission throughout the olfactory bulb. This was confirmed by double-immunostaining studies which showed the expression of both GAT3 and glial fibrillary acidic protein (GFAP) mRNAs in astrocytes. Moreover, GAT2 mRNA was expressed only in the ependyma and arachnoid. These findings suggest that the expression patterns of GABA transporters differ with the type of cells in the rat olfactory bulb where GAT1 and GAT3 may play an imporatant role in GABA-mediated transmission, such as lateral inhibition.

Animals↗
PubMed1997

Localization of molecules involved in cytokine receptor signaling in the rat trigeminal ganglion.

The localization of some cytokine receptors and their downstream intracellular signaling molecules was examined in the trigeminal ganglia of rats. Among cytokine receptor components, we examined signal transduction subchain, gp130, IL-2Rgamma and IL-5Rbeta, which are common to respective groups of cytokine receptors. Most of the sensory ganglion neurons expressed gp130, but not IL-2Rgamma nor IL-5Rbeta. We further examined the localization of Janus kinase (JAK) family members which were reported to be associated with various kind of cytokine receptors and are thought to be implicated in major cytokine receptor-signaling pathways [6,9,11,13]. While JAK1 and Tyk2 were expressed in all the type of neurons, JAK2 was predominantly expressed in the small neurons. In addition, JAK3 immunoreactivity was only found in satellite cells. The present results indicate that most of neurons express gp130, and that the localization of JAK family members differs with the cell type. This also suggests that the cytokine receptor-signaling pathway may be different in neuronal and glial cells.

Animals↗
PubMed1997

Role of tyrosine specific phosphorylation of cellular proteins, especially EGF receptor and p125FAK in human lung cancer cells.

To clarify the role of tyrosine phosphorylation of cellular proteins in human lung cancer cells, phosphotyrosine (PTYR)-containing proteins in lung cancer cell lines and in paired tissues resected from cancerous and normal lungs were studied by immunoblotting with an anti-PTYR antibody. We found that the profiles of protein phosphorylation were very similar among those cell lines which had different histological features. The major PTYR-containing proteins (180-190 KDa, 120-130 KD, and 95-100 KDa) were detected in lung cancer cell lines. The expression of EGF receptor (EGF-r) (p185) and o-erb B2 protein, and tyrosine phosphorylation of p125FAK were examined in cancerous lung tissues and normal lung tissues. In surgical specimens, approximately half of the samples of lung cancer tissues showed clear elevation of tyrosine phosphorylation. In these cancerous tissues, no clear amplification of EGF-r and c-erb B2 protein expression was observed. However, elevation of tyrosine phosphorylation of p125FAK was observed in cancerous lung tissues but not in normal lung tissues, and its phosphorylation was closely correlated with the nodal involvement of cancer and disease-free survival time. These results suggested that the intracellular signaling pathway via tyrosine phosphorylation plays a role in the generation and immortalization of lung cancer, and assessment of tyrosine phosphorylation of cellular proteins. especially p125FAK, may be available clinically as a prognostic factor.

Adult↗
PubMed1997

Pharmacologic inhibition of twin-pulse facilitation of release of transmitter quanta at the mouse neuromuscular junction.

1. The frequency (F,s-1) of miniature endplate potentials and the quantal content (m) of endplate potentials were simultaneously measured intracellularly at mouse diaphragm endplates in a bath solution that contained 0.6 mM Ca2+ ions and 5 mM Mg2+ ions. 2. Twin pulses at 4-ms intervals gave the quantal contents of the first (m1) and second (m2) responses. The ratio of m2/m1 was taken as an indicator of the temporal facilitation of the release of transmitter. 3. Lead ions (Pb2+; 10 microM), bis (o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA; loaded for 60 min at 200 microM), and chlortetracycline (CTC; loaded for 30 min at 80 microM) reduced the values of F and m2/m1. Pb2+ ions and CTC reduced the value of m, whereas BAPTA did not.omega-Agatoxin (omega AGT; 10 ng/ml) reduced the value of m without affecting F or m2/m1. 4. These results suggest that synaptic facilitation is modifiable by agents that can affect systems which buffer intracellular levels of Ca2+ ions.

Animals↗
PubMed1997

Twitch potentiation induced by caffeine in the mouse diaphragm depends on external calcium ions in the absence of potassium ions.

1. Removal of external K+ ions increases the amplitude of directly elicited twitch contractions of the mouse diaphragm (Nishimura et al., 1996). This increase depends on external Ca2+ ions. 2. We examined the effect of caffeine (2 mM) on this increase in twitch amplitude. The mouse diaphragm muscle was directly stimulated in the presence of d-tubocurarine (10 microM). 3. Caffeine increased the amplitude of twitches in a standard bathing solution. This effect was maintained in a solution without either K+ or Ca2+ ions but was abolished in a solution from which both ions were absent. Readdition of Ca2+ ions restored the potentiating effect of caffeine. 4. In the presence of caffeine, removal of both K+ and Ca2+ ions decreased the resting membrane potentials of muscle fibers to about -53 mV. The readdition of 2 mM Ca2+ ions restored the membrane potentials. 5. Twitch potentiation in the absence of external K+ ions was attenuated by 10 microM bepridil but not by 3 microM verapamil or 10 microM Cd2+ ions. 6. These results support the hypothesis that Na(+)-Ca2+ exchange can support twitch contraction during the inhibition of Na(+)-K(+)-ATPase activity. The influx of Ca2+ ions into the cells might be stored in the sarcoplasmic reticulum.

Animals↗