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Biomedical subjects

M Nishimura

Publications and source records attributed to M Nishimura.

At least 595 records · Page 33Linked to original sources
PubMed1996

HLA-A null allele with a stop codon, HLA-A*0215N, identified in a homozygous state in a healthy adult.

A healthy adult having no serologically detectable HLA class I A locus antigens was identified. The parents of the individual are consanguineous. Results of a family study indicated that the individual is homozygous for the B46-Cw1-DR8.1 haplotype, which was shown to be positively associated with A*0207 in our previous study. The HLA-A null individual is healthy and exhibits no apparent immunological abnormality. Total RNAs extracted from peripheral blood were converted to cDNAs. The reverse transcriptase-polymerase chain reaction (PCR) product, which is of the same size as the normally expressed gene, was easily obtained from the cDNAs with HLA-A locus-specific primers. The nucleotide sequence of this null allele (A*0215N) was the same as that of A*0207 except for a single nucleotide substitution which resulted in a stop codon in exon 4. From its nucleotide sequence, a truncated molecule was expected to be produced; however, the immunoprecipitation study failed to detect the predicted product. Genomic DNAs from 29 unrelated individuals who expressed only one HLA-A antigen with HLA-B46, were analyzed by a PCR-sequence-specific oligonucleotide method. None of the samples possessed this stop codon. Therefore, A*0215N is likely to be a rare allele generated by a single point mutation from A*0207.

Adult↗
PubMed1996

The role of Ca2+ release from sarcoplasmic reticulum in the regulation of sinoatrial node automaticity.

The role of Ca2+ release channels in the sarcoplasmic reticulum in modulating physiological automaticity of the sinoatrial (SA) node was studied by recording transmembrane action potentials and membrane ionic currents in small preparations of the rabbit SA node. Ryanodine, which modifies the conductance and gating behavior of the Ca2+ release channels, was used to block Ca2+ release from the sarcoplasmic reticulum. Superfusion of 1-mM ryanodine decreased the spontaneous firing frequency as well as the maximal rate of depolarization of the SA, and these reductions reached a steady state within approximately 5 min. The action potential recordings revealed that the latter part of diastolic depolarization was depressed and that the take-off potential became less negative. This suggested that the negative chronotropic effect of ryanodine resulted from the blockade of physiological Ca2+ release from the sarcoplasmic reticulum. In voltage clamp experiments, using double-microelectrode techniques, ryanodine did not markedly reduce the Ca2+ current (ICa) but decreased the delayed rectifying K+ current (IK), the steady-state inward current (Iss), and the hyperpolarization-activated inward current (Ih). These observations suggest that, even when the function of C2+ channels in the cell membrane is normally maintained, depression of Ca2+ release channels in the sarcoplasmic reticulum would prevent sufficient elevation of the Ca2+ concentration in SA node cells for the activation of various ionic currents, and, thus adversely affect the physiological automaticity of this primary cardiac pacemaker.

Action Potentials↗
PubMed1996

Modification of fatty acid composition by over- and antisense-expression of a microsomal omega-3 fatty acid desaturase gene in transgenic tobacco.

omega-3 fatty acid desaturases, which catalyse the conversion of linoleic acid (18:2) to linolenic acid (18:3) in lipids, are located in the microsomes and plastid membranes. Transgenic tobacco plants were produced that express the transcripts of a tobacco microsomal omega-3 fatty acid desaturase gene (NtFad3) in antisense and sense orientations under the control of the cauliflower mosaic virus 35S promoter. The antisense construct has the 0.5-kb fragment of the NtFad3 cDNA containing a 3'-flanking region and a part of the coding region in antisense orientation. The antisense-transformant lines showed decreases of the steady-state NtFad3 mRNA level to 30% of the control plants. In these lines, the 18:3 content decreased to about 80% in root tissues and to about 70-80% in leaf tissues when compared with the control plants. The sense construct has the 1.4-kb full-length cDNA of NtFad3. In one of the sense-transformant lines, the NtFad3 mRNA level increased 8 times when compared with that of the control plants. In this line, the 18:3 content increased by about 1.5-fold in root tissues and by about 1.1-fold in leaf tissues. These results indicate that the up- and down-regulation of the transcript level in the microsomal omega-3 fatty acid desaturase gene is useful to modify the 18:3 content in the vegetative tissues of higher plants.

Antisense Elements (Genetics)↗
PubMed1996

Electron microscopic study of retinas of macular mice.

BACKGROUND: Menkes' disease may be due to a lack or deficiency of copper in various organs. The macular mouse is known as a model for Menkes' disease. We examined melanin granules in the retinal pigment epithelium and the activity of cytochrome oxidase, a copper-containing enzyme, in the retinas of macular mice by electron microscopy. METHODS: In the retinas of hemizygote macular mice we demonstrated cytochemically (oxidative polymerization of diaminobenzidine to an osmophilic reaction product) the activity of cytochrome oxidase. The distribution of melanin granules in the retinal pigment epithelium related to the activity of another copper-containing enzyme, tyrosinase was also studied. Stereological methods were applied to obtain quantitative data. RESULTS: In the retinal photoreceptor inner segments of the macular mouse, the mitochondria were more numerous than in normal litermates and they appeared swollen. There were fewer melanin granules in the retinal pigment epithelium of macular mice than in that of normal littermates. The cytochrome oxidase activity was significantly lower in the macular mice than in the controls. CONCLUSION: Macular mice have lower activity of cytochrome oxidase and fewer melanin granules than do normal mice. Both changes may be related to copper deficiency. These results correspond to the retinal changes seen in patients with Menkes' disease.

Animals↗
PubMed1996

99mTc-MIBI accumulation in the parathyroid autograft in a patient with recurrent hyperparathyroidism.

A case with recurrent hyperparathyroidism secondary to chronic renal insufficiency is reported. The patient had undergone total parathyroidectomy and autotransplantation of parathyroid tissue five years ago. Bone scintigraphy clearly demonstrated skeletal involvement of secondary hyperparathyroidism and 99mTc-methoxyisobutylisonitrile scintigraphy clearly demonstrated a hyperfunctioning parathyroid autograft.

Bone and Bones↗
PubMed1996

Localization of Menkes gene expression in the mouse brain; its association with neurological manifestations in Menkes model mice.

Menkes gene (Mc1 or MNK, encoding putative copper-transporting ATPase) expression was investigated and compared in normal and macular mutant mouse brain. Northern blot analysis showed a distinct 8.3-kb transcript and no obvious difference in size or extent in normal mice and macular mutants on postnatal days 0, 4, 7, 10 or 13. In situ hybridization revealed that certain specific populations of cells in the brain express Menkes mRNA, and that their localization in normal and mutant mice did not differ and was conserved on days 4, 10 and 13. The most intense hybridization signals were observed in the hippocampal CA1 region and dentate gyrus, the olfactory bulb nuclei, the cerebellar granular cell layer, the choroid plexus and the ependyma, with less intense signals in the hippocampal CA3 region and cerebellar Purkinje cells. In addition, necrotic neuronal cell death was predominantly observed in the CA3 region and the Purkinje cells of macular mice after postnatal day 10. The finding that the regions that had lower expression level of Menkes mRNA corresponded to those showing neuronal necrosis suggests that the Menkes gene may be responsible for the neuronal degeneration in some specific portions of the brain and clinical manifestations in this mutant.

Adenosine Triphosphatases↗
PubMed1996

Effects of M1-selective antimuscarinics on respiratory chemosensitivity in humans.

We examined effects of selective M1 antagonists on hypercapnic and hypoxic ventilatory responses in 17 healthy human volunteers. Subjects were intravenously treated with placebo, pirenzepine (10 mg) and biperiden lactate (4 mg) on three separate days in a randomized double-blind design. Ventilatory responses to hyperoxic progressive hypercapnia and isocapnic progressive hypoxia were studied after the drug administration. There were no statistically significant differences in the mean delta VE/delta PET CO2 or delta VE/delta SaO2 among the three treatments. However, the delta VE/delta PET CO2 with placebo negatively correlated with the difference in delta VE/delta PET CO2 between the biperiden and placebo studies (r=-0.65, P < 0.01), but not with that between the pirenzepine and placebo studies. On the other hand, the delta VE/delta SaO2 with placebo negatively correlated with the difference in delta VE/delta SaO2 between the pirenzepine and placebo studies (r = -0.79, P < 0.001), but not with that between the biperiden and placebo studies. These data suggest the possible involvement of M1 cholinergic receptors in the central CO2 and peripheral O2 sensing mechanisms in humans, although the degree of its involvement is not consistent among subjects. These findings may explain the interindividual variation in the control of breathing in humans.

Adolescent↗
PubMed1996

Islet amyloid polypeptide/amylin contents in pancreas change with increasing age in genetically obese and diabetic mice.

To search for a possible relationship between islet amyloid polypeptide (IAPP)/amylin and the pathophysiology of non-insulin-dependent (type 2) diabetes mellitus (NIDDM), we examined the changes in IAPP contents in the pancreata of genetically obese and diabetic mice (C57BL/6J ob/ob and C57BL/KsJ db/db mice). In the male ob/ob mice, IAPP and insulin contents began to increase at 16 weeks and continued to increase. In the male db/db mice, IAPP content began to increase at 8 weeks of age and insulin content at 4 weeks. Both contents continued to increase until 16 weeks, but drastically decreased at 24 weeks. Immunohistochemical studies using anti-IAPP8-17 antibody showed the increase of islet cell mass and the heterogeneous immunoreactivity for IAPP in islet cells in the ob/ob mice at 24 weeks of age. In the db/db mice at the same age, the immunoreactivity was heterogeneous and weak in many islet cells. These results suggest that genetic factors that are important in the manifestation of NIDDM influence the capacity of beta-cells to synthesize and secrete IAPP, and that IAPP synthesis and secretion change in the course of the disease.

Aging↗
PubMed1996

Twitch responses dependent on external calcium ions in the mouse diaphragm in a potassium-free bathing solution.

We examined the effect of external Ca2+ on twitches of mouse diaphragm under the inhibition of Na(+)-K(+)-ATPase in vitro. The muscle was directly stimulated in presence of d-tubocurarine (dTc). K+ removal potentiated the amplitude of twitches with a transient prior reduction. This potentiation depended on external Ca2+. Membrane potentials decreased after removal of both external K+ and Ca2+ and were restored after an addition of Ca2+. The removal of both ions increased the content of Na+ in tissues and decreased K+. These changes were restored to the levels in the K(+)-free bathing solution by the addition of Ca2+. These results imply that Na+/Ca2+ exchange can support twitch contraction under the inhibition of Na(+)-K(+)-ATPase activity.

Animals↗
PubMed1996

Effect of pulverization on dehydration behavior of crystals of GK-128, a serotonin3 receptor antagonist.

The effect of pulverization on the dehydration behavior of 2-[(2-methylimidazol-1-yl)methyl]benzo[i]thiochromen-1-one monohydrochloride hemihydrate (GK-128), a newly developed serotonin3 receptor antagonist, was studied using powder X-ray diffraction analysis, thermogravimetry (TG), and differential scanning calorimetry (DSC). The crystalline forms of GK-128 obtained by pulverizing with a jet mill and an agale mortar were each confirmed to be the same as that of intact GK-128, since the powder X-ray diffraction patterns of the two prepared samples were identical with that of intact GK-128. However, after pulverization by jet mill, the dehydration temperature of GK-128 was markedly lowered and the DSC endothermic peak due to dehydration disappeared. The activation energy for dehydration, calculated by the Ozawa method using TG data, decreased with decreasing particle size and/or crystallinity of GK-128 crystals, e.g., the activation energies for dehydration of intact and jet-milled GK-128 were 128.1 and 75.9 kj/mol, respectively. The results of comparison of powder X-ray diffraction patterns of pulverized GK-128 and intact GK-128 and of determination of the crystal structure of GK-128 suggested that water molecules could be removed easily along the c-axis of GK-128 crystals.

Calorimetry, Differential Scanning↗
PubMed1996

Tyrosine phosphorylation of 100-130 kDa proteins in lung cancer correlates with poor prognosis.

To search for the signalling pathways in lung cancer relevant to its aggressive behaviour, we studied tyrosine phosphorylated proteins in lung cancer cell lines and surgical specimens. We found that the profiles of protein phosphorylation were closely matched among these cell lines and cancer tissues of different histological origins, and 100-130 kDa proteins were the major components of phosphorylated proteins. In surgical specimens, approximately half of the cases showed tyrosine phosphorylation of these proteins in a tumour-specific manner, and phosphorylation of these proteins showed good correlation with the survival length of patients after operation. By immunoprecipitation with specific antibodies, we found that p125FAK, p120 and beta-catenin were the major components of tyrosine-phosphorylated proteins in the surgical specimens. These results suggest that tyrosine phosphorylation of these proteins may play a role in tumour relapse and is available as a clinical marker.

Aged↗
PubMed1996

A genetic linkage map of the Syrian hamster and localization of cardiomyopathy locus on chromosome 9qa2.1-b1 using RLGS spot-mapping.

The Syrian cardiomyopathic hamster (BIO14.6) has an inherited form of progressive myocardial necrosis and congestive heart failure. Although widely studied as an animal model for human hypertrophic cardiomyopathy, further genetic analysis has been limited by a scarcity of DNA markers. Until now, only six autosomal linkage groups have been described and the number of polymorphic loci was extremely limited. In this study, we applied the restriction landmark genome scanning (RLGS) spot-mapping method to construct a genetic map of the Syrian hamster (Mesocricetus auratus) using 72 back-cross progeny. Although the polymorphic rate is very low (3-7%) between the strains, 531 polymorphic spots/loci were mapped, showing the power of this approach and reasonable applicability to other organisms lacking a well-defined genetic map. Further, the spot markers which flank the cardiomyopathy (cm) locus were cloned to determine the chromosomal location of cm by fluorescent in situ hybridization (FISH) analysis, resulting in the assignment of the locus to the centromeric region of hamster chromosome 9qa2.1-b1. Several candidate genes responsible for hypertrophic cardiomyopathy in humans have been excluded.

Animals↗
PubMed1996

Evidence of involvement of cytotoxic antibodies directed against patients's HLA class II produced by transfused donor-derived B cells in post-transfusion graft-versus-host disease.

Post-transfusion graft-versus-host disease (PTGVHD) is one of the most severe side-effects of blood transfusion. To characterize the effector cells causing this disease, we cloned lymphocytes from a PTGVHD patient's peripheral blood. T-cell and B-cell clones were established, the origins of which were proven to be transfused donor lymphocytes. It was found that the B cells produced IgG that mediated complement-dependent cytotoxicity to the cells bearing the patient's HLA class II genotype. Our results suggest, for the first time, the involvement of B-cell-produced cytotoxic antibodies directed against patient's HLA class II in the pathogenesis of PTGVHD.

Antibodies↗
PubMed1996

Transport of chimeric proteins that contain a carboxy-terminal targeting signal into plant microbodies.

Malate synthase is a glyoxysome-specific enzyme. The carboxy-terminal tripeptide of the enzyme is Ser-Arg-Leu (SRL), which is known to function as a peroxisomal targeting signal in mammalian cells. To analyze the function of the carboxy-terminal amino acids of pumpkin malate synthase in plant cells, a chimeric gene was constructed that encoded a fusion protein which consisted of beta-glucuronidase and the carboxyl terminus of the enzyme. The fusion protein was expressed and accumulated in transgenic Arabidopsis that had been transformed with the chimeric gene. Immunocytochemical analysis of the transgenic plants revealed that the carboxy-terminal five amino acids of pumpkin malate synthase were sufficient for transport of the fusion protein into glyoxysomes in etiolated cotyledons, into leaf peroxisomes in green cotyledons and in mature leaves, and into unspecialized microbodies in roots, although the fusion protein was no longer transported into microbodies when SRL at the carboxyl terminus was deleted. Transport of proteins into glyoxysomes and leaf peroxisomes was also observed when the carboxy-terminal amino acids of the fusion protein were changed from SRL to SKL, SRM, ARL or PRL. The results suggest that tripeptides with S, A or P at the -3 position, K or R at the -2 position, and L or M at the carboxyl terminal position can function as a targeting signal for three kinds of plant microbody.

Amino Acid Sequence↗
PubMed1996

Isolation and characterization of a cDNA encoding mitochondrial chaperonin 10 from Arabidopsis thaliana by functional complementation of an Escherichia coli groES mutant.

Chaperonin (Cpn) is one of the molecular chaperones. Cpn10 is a co-factor of Cpn60, which regulates Cpn60-mediated protein folding. It is known that Cpn10 is located in mitochondria and chloroplasts in plant cells. The Escherichia coli homologue of Cpn10 is called GroES. A cDNA for the Cpn10 homologue was isolated from Arabidopsis thaliana by functional complementation of the E. coli groES mutant. The cDNA was 647 bp long and encoded a polypeptide of 98 amino acids. The deduced amino acid sequence showed approximately 50% identity to mammalian mitochondrial Cpn10s and 30% identity to GroES. A Northern blot analysis revealed that the mRNA for the Cpn10 homologue was expressed uniformly in various organs and was markedly induced by heat-shock treatment. The Cpn10 homologue was constitutively expressed in transgenic tobaccos. Immunogold and immunoblot analyses following the subcellular fractionation of leaves from transgenic tobaccos revealed that the Cpn 10 homologue was localized in mitochondria and accumulated at a high level in transgenic tobaccos.

Amino Acid Sequence↗
PubMed1996

Distribution of and age-related changes in ciliary neurotrophic factor protein in rat tissues.

We developed a sensitive enzyme-linked immunosolvent assay for measuring ciliary neurotrophic factor (CNTF) and examined age-related changes in the CNTF contents of a variety of rat tissues during postnatal development. CNTF contents were substantially higher in the sciatic nerve and spinal cord than in the other tissues tested, the kidney coming third. In all the tissues except the sciatic nerve (90 ng/g), the CNTF content was less than 1 ng/g at 1 week of age, then gradually increased. It was highest at 5 weeks of age in the sciatic nerve (3171 ng/g), spinal cord (118 ng/g), and kidney (36.8 ng/g), after which it slowly decreased. In contrast, the maximum in the brain stem (9 ng/g) and cerebellum (3.6 ng/g) was at 8 weeks of age, whereas in skeletal muscle it was at 2 weeks of age (14.6 ng/g). These findings indicate that CNTF functions in the postnatal development of the rat.

Aging↗
PubMed1996

Cloning and expression of the inorganic pyrophosphatase gene from thermophilic bacterium PS-3.

The thermophilic bacterium PS-3 ppa gene encoding inorganic pyrophosphatase (PPiase) has been cloned and sequenced. The deduced amino acid sequence was exactly the same as that determined on Edman degradation (Ichiba, T., Takenaka, O., Samejima, T. and Hachimori, A., J. Biochem. 108: 572-578, 1990) except that amino acid-70 is threonine instead of proline and two more amino acids, Asn-Lys, are present at the carboxyl terminus. The structural gene containing BamH I and Hind III restriction cleaving sites at the 5' and 3' ends, respectively, was amplified by the PCR method. Escherichia coli JM109 was transformed with a synthetic DNA, and we found that thermophilic ppa was expressed in E. coli, as judged on enzyme assaying, SDS-PAGE and immune assaying.

Amino Acid Sequence↗