[Prospects in telemedicine].
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Clinical Laboratory Techniques↗
Biomedical subjects
M Okuda
Publications and source records attributed to M Okuda.
Involvement of NADH/NADPH oxidase in human platelet ROS production.
Platelets play an important role in atherosclerotic and thromboembolic vascular diseases. It has been reported that reactive oxygen species (ROS) could modify platelet function, and platelets themselves have the ability to produce ROS. However, the enzymatic sources of ROS in platelets have not been fully determined. The NADH/NADPH oxidase system was originally identified as the major source of ROS in phagocytes. Recently, it has become evident that this oxidase is functionally expressed not only in phagocytes but also in various cell types. The present study was undertaken to test the hypothesis that NADH/NADPH oxidase might be expressed in human platelets. Lucigenin-enhanced chemiluminescence (L-CL) and electron spin resonance (ESR) method demonstrated that human platelets obtained from healthy volunteers released ROS, and the released ROS were increased by stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA) or calcium ionophore. Homogenates of human platelets, as well as MEG01 cells, megakaryocytic cell line, had the enzymatic activity to produce superoxide in NADH/NADPH-dependent manners. This enzymatic activity was suppressed by diphenylene iodonium (DPI), an inhibitor of NADH/NADPH oxidase. Western blot analysis demonstrated that platelets and MEG01 cells expressed p22(phox) and p67(phox) proteins, components of NADH/NADPH oxidase. Thus, human platelets have the enzymatic activity of p22(phox)-based NADH/NADPH oxidase, and this oxidase is likely one of the important sources of ROS in platelets.
Blood Platelets↗
Inhibitory effects of 6-O-acylated L-ascorbic acids possessing a straight- or branched-acyl chain on Epstein-Barr virus activation.
6-O-Acylated L-ascorbic acids possessing a straight- or branched-acyl chain of varying length from C(4) to C(18) have been synthesized and evaluated their anti-tumor promoting effects on the activation of the Epstein-Barr virus early antigen. The derivatives having a straight- or branched-acyl chain of C(6) to C(11) carbon atoms exhibited marked effects.
Anticarcinogenic Agents↗
Molecular cloning and sequencing of canine T-cell costimulatory molecule (CD28).
T-cells express CD28 and CTLA-4, and through binding to their shared ligands (CD80/CD86) on antigen presenting cells, provide a potent co-stimulatory signal for T-cell activation and proliferation. To investigate the role of CD28 in canine immune system, we hereby report the molecular cloning and sequencing of the full-length complementary DNA (cDNA) coding for canine CD28, from pokeweed mitogen stimulated canine peripheral blood lymphocytes. The cloned cDNA contains an open reading frame of 663 nucleotides, encoding for a polypeptide of 221 amino acids. The amino acid sequence of the canine CD28 showed 91.9, 80, and 79.6% similarities with those of the cat, cattle, and human counterparts, respectively. Five sequence motifs of TATT or ATTTA involved in the regulation of gene expression by influencing mRNA stability are found in the 3' untranslated region. The hexapeptide motif (MYPPPY), five cysteine residues, a potential N-glycosylation site and a cytoplasmic phosphatidylinositol 3-kinase binding site in canine CD28 molecule are completely conserved in canine CTLA-4. The availability of full length canine CD28 will provide a useful molecule for studying its role in dog immune system.
Abatacept↗
Effect of 1,1,1-trichloroethane on calcium current of rat dorsal root ganglion neurons.
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Animals↗
Comparison of the S RNA segments among Japanese isolates and Taiwanese isolates of Watermelon silver mottle virus.
The complete nucleotide sequences of the S RNA of two Japanese isolates of Watermelon silver mottle virus were determined. One was isolated from naturally infected watermelon and causes malformation on upper leaves of Tetragonia expansa. The other was isolated from melon and causes characteristic yellow necrotic lesions on upper leaves of T. expansa. The total nucleotide sequences of the S RNA of WS-O and WS-Y were 3553 nt and 3558 nt long, respectively. Both the nucleotide sequences and the deduced amino acid sequences of WS-O and WS-Y were quite similar even though the symptoms on T. expansa are quite different. They were also significantly similar to those of the Taiwanese isolates, Topso-W and Tospo-To. These results suggested that the Japanese isolates and the Taiwanese isolates of WSMoV were classified as one group not only serologically but also genetically. Within the S RNA sequences, the most variable region was the intergenic region between the N gene and the NSs gene. This was due to a 20 nt insertion between the Japanese isolates and the Taiwanese isolates.
Amino Acid Sequence↗
Grading the severity of allergic rhinitis for treatment strategy and drug study purposes.
Grading the severity of rhinitis symptoms is essential for the evaluation of drug efficacy. However, a uniform grading system has not been standardized. A review of 44 journal articles and symposia from Japan, Copenhagen, and the World Health Organization indicates major differences among systems. All except the World Health Organization used a sum of rhinitis scores. The lack of standardization limits comparisons of subjective symptom changes after comparable treatments. Corrective measures include the use of evidence-based definitions of independent nasal symptoms, avoidance of biases in evaluation, and establishment of international consensus for a standardized scale in this era of globalization.
Asthma↗
Molecular cloning and expression analysis of feline melanoma antigen (MAGE) obtained from a lymphoma cell line.
Melanoma antigens (MAGE) are regarded as inducing tumor-specific immune response and thought to be potential therapeutical agents for cancer immunotherapy. We hereby report the cloning of feline MAGE cDNA obtained from a lymphoma cell line derived from cat malignant lymphoma, and its expression pattern in tumor and normal tissues. The cDNA encoding the MAGE is 1668 base pairs (bp) in length, and contains an open reading frame (ORF) of 936 bp encoding a protein of 311 amino acids. The predicted amino acid sequence has 29-46% of homology with other MAGE proteins from human and mouse. mRNA transcripts for the feline MAGE were detected in certain tumors, but not in adult cat normal tissues except in testis, by reverse transcription polymerase chain reaction (RT-PCR) analysis. This indicates that the expression pattern of feline MAGE mRNA is similar to those of other MAGE family genes in tumors and normal tissues.
Amino Acid Sequence↗
RT-PCR for detecting five distinct Tospovirus species using degenerate primers and dsRNA template.
RT-PCR procedures for detection of multiple species of tospovirus from plant tissues were investigated. Downstream primers were designated from the 3' untranslated sequences of the S RNA. An upstream primer was designated from the degenerated sequences of the nucleocapsid protein. Approximately 450 bp DNA fragments were detected when Tomato spotted wilt virus (TSWV)- or Impatiens necrotic spot virus (INSV)- infected tissues were examined. Approximately 350 bp DNA fragments were detected when Watermelon silver mottle virus (WSMoV)- or Melon yellow spot virus (MYSV)-infected tissues were examined. Template RNA was extracted using CF 11 cellulose powder, and nonspecific amplification became unnoticeable when double-stranded RNA was used. The amplified fragments of WSMoV were differentiated from those of MYSV by AluI or TaqI digestion. The amplified fragments of TSWV were differentiated from those of INSV by DraI or HindIII digestion. An alstroemeria plant that was infected with an unidentified tospovirus was also examined, and a 350 bp fragment that was 97% identical to Iris yellow spot virus was detected. This method, therefore, detected at least five distinct Tospovirus species.
Amino Acid Sequence↗
Effect of high population density environment on skin barrier function in mice.
Previous reports have demonstrated that stressful stimuli markedly influenced physiological conditions and homeostasis. In this study, we examined the influence of high population density environment on barrier function and water retention property of skin in mice. Overcrowding stress induced dramatic body weight reduction and significant increase in adrenal gland weight. Macroscopic skin appearance showed moderate exfoliation and slight wrinkle formation in the stress group. Moreover epidermis hyperplasia not concomitant with inflammatory reactions such as infiltration of immunocytes and vasodilation was observed in the skin from the stress group. Skin surface conductance in the stress group was significantly lower than in the control group. On the other hand, transepidermal water loss in the stress group increased significantly, compared to the control group. Moreover, transdermal penetration of indomethacin and nicotinic acid amide was enhanced significantly. These results suggest that overcrowding stress induced impairment of barrier function and water retention property. To elucidate the mechanism of overcrowding stress-induce dry skin, contents of ceramide and pyrrolidone carboxylic acid, which are the important compounds contributing to those functions were evaluated. Both of them in stratum corneum declined significantly in the stress group. Taken together, these results demonstrate that overcrowding stress induced dry skin, and that the impairment of barrier function and water retention property correlated with decrease in ceramide and pyrrolidone carboxylic acid.
Adrenal Glands↗
Distinct characteristics of organic cation transporters, OCT1 and OCT2, in the basolateral membrane of renal tubules.
PURPOSE: This study was performed to determine the detailed mRNA distribution of organic cation transporters, rOCT1 and rOCT2, along the rat nephron and to distinguish the substrate affinities of these transporters. METHODS: The distributions of rOCT1 and rOCT2 mRNA were determined by reverse transcriptase polymerase chain reaction analysis of microdissected nephron segments. Using MDCK cells transfected with rOCT1 or rOCT2 cDNA, the inhibitory effects of various compounds on the uptake of [14C]tetraethylammonium were assessed. RESULTS: rOCT1 mRNA was detected primarily in the superficial and juxtamedullary proximal convoluted tubules, whereas rOCT2 mRNA was detected widely in the superficial and juxtamedullary proximal straight and convoluted tubules, medullary thick ascending limbs, distal convoluted tubule, and cortical collecting duct. The IC50 values for cationic drugs and endogenous cations on [14C]tetraethylammonium uptake across the basolateral membranes in the transfectants indicated that rOCT1 and rOCT2 had similar inhibitor specificity for many compounds but showed moderate differences in the specificity for several compounds, such as 1-methyl-4-phenylpyridinium, dopamine, disopyramide, and chlorpheniramine. CONCLUSIONS: rOCT1 and rOCT2 possess similar but not identical multispecificities for various compounds with distinct distributions along the nephron, indicating that the two transporters share physiologic and pharmacologic roles in the renal handling of cationic compounds.
Animals↗
Exon 3 of tyrosine hydroxylase gene: lack of association with Japanese schizophrenic patients.
Tyrosine hydroxylase (TH) is the rate-limiting enzyme in dopamine (DA) biosynthesis. Exon 3 of the human TH gene encodes the sequence from Ser31 to Glu104 of type 1 enzyme, which contains the critical parts for regulation of the catalytic activity. The amino acid residues Gly36-Arg37-Arg38 were identified as a key sequence for DA to exert its inhibitory effect on catalytic activity. Therefore, we screened the nucleotide sequences of exon 3 from 201 Japanese patients with schizophrenia to explain the elevation in the synaptic or presynaptic DA concentrations in the schizophrenic brain, based on the hypothesis that any mutation changing the amino acid sequence Gly36-Arg37-Arg38 would result in the elevation of DA synthesis, due to a reduced inhibitory effect of DA on the catalytic activity. However, no mutated sequences of exon 3 and both exon-intron boundaries were detected in any of the patients examined. Polymorphisms generating Val81 and Met81 were compared of the distributions of genotype and allele between the patients and 175 Japanese healthy controls, which did not suggest an association between the polymorphism and schizophrenia. These results indicate that exon 3 of the human TH gene lacks association with schizophrenia in Japanese patients.
Adult↗
A possible role of hypervariable region 1 quasispecies in escape of hepatitis C virus particles from neutralization.
We examined serial changes in the hypervariable region 1(HVR1) quasispecies both in immune and nonimmune complexed hepatitis C virus (HCV) particles from 12 patients with chronic hepatitis C to elucidate the mechanism by which genetic diversification of HCV during the course of infection allows escape of virus from the humoural immune response. Immune and nonimmune complexes were separated by differential flotation centrifugation and immunoprecipitation, and their HVR1 quasispecies were determined by subcloning and sequencing. The presence of a specific antibody against a specific viral clone in serum was examined in two patients by Western blotting of the corresponding recombinant HVR1 protein. The distribution of HVR1 quasispecies in both immune and nonimmune complexes conspicuously changed over time in most of the patients studied. In seven patients, various HCV clones serially shifted from nonimmune complexes to immune complexes. In four of them, a group of clones with similar HVR1 sequences to each other remained predominant in nonimmune complexes, whereas minor clones with sequences considerably divergent from the predominant clones shifted from nonimmune complexes to immune complexes. These results suggest a mechanism for persistent infection of HCV, in which major HCV clones escape from neutralization by anti-HVR1 antibodies by generating considerably divergent minor 'decoy' clones which may be preferentially neutralized.
Adult↗
Breast-feeding prevents Helicobacter pylori infection in early childhood.
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Age Distribution↗
Schinzel-Giedion syndrome.
A 2-month-old girl was brought to the Department of Pediatrics at Wakayama Rosai Hospital because of poor feeding since 1 month of age. She was the third child of young healthy non-consanguineous parents whose first son was healthy but whose second son had died of 18 trisomy. Physical examination showed midfacial hypoplasia with coarse dysmorphic features, choanal stenosis, remarkable abdominal distention and bilateral talipes equivarus. Abdominal ultrasonography, computed tomography and drip infusion pyelogram showed left severe hydronephrosis and right moderate hydronephrosis. Having diagnosed Schinzel-Giedion syndrome, a left ureteroneocystostomy with tailoring was performed to preserve renal functions and to eliminate the urinary tract infection at the age of 3 months.
Abnormalities, Multiple↗
Characterization of monoclonal antibodies specific for feline serum amyloid (SAA) protein.
Serum amyloid A (SAA) has been characterized as an inflammatory marker in many species. In this study, we have developed and characterized monoclonal antibodies (MAbs) against feline SAA (fSAA) derived from culture hybridomas. These hybridomas were produced from the fusion of Balb/c-derived myeloma s/p20-Ag14 and splenocytes from mice immunized with purified recombinant feline SAA (rfSAA). Six hybridomas secreting MAbs, M2, M5, M7, M8, M13, and M15, were selected and subcloned on the basis of their specificity to rfSAA by enzyme-linked immunoabsorbent assay (ELISA), and confirmed based on their specificity to rf-SAA by immunoblot analysis. Out of six clones, two clones (M5 and M7) showed higher reactivity with rf-SAA, and were selected for further analysis of ELISA additivity and Western blot cross-reactivity tests. As a result, M5 and M7 clones recognized the same or excessively near epitopes on rfSAA and reacted with rfSAA, fSAA and equine recombinant SAA, but showed no reaction with human recombinant SAA. Because of their specificity, these MAbs may be usefully applied in studying the measurement of SAA concentration in cat serum.
Amino Acid Sequence↗
Mutation of an arginine biosynthesis gene causes reduced pathogenicity in Fusarium oxysporum f. sp. melonis.
Restriction enzyme-mediated integration (REMI) mutagenesis was used to tag genes required for pathogenicity of Fusarium oxysporum f. sp. melonis. Of the 1,129 REMI transformants tested, 13 showed reduced pathogenicity on susceptible melon cultivars. One of the mutants, FMMP95-1, was an arginine auxotroph. Structural analysis of the tagged site in FMMP95-1 identified a gene, designated ARG1, which possibly encodes argininosuccinate lyase, catalyzing the last step for arginine biosynthesis. Complementation of FMMP95-1 with the ARG1 gene caused a recovery in pathogenicity, indicating that arginine auxotrophic mutation causes reduced pathogenicity in this pathogen.
Arginine↗
Expression of glutaredoxin in human coronary arteries: its potential role in antioxidant protection against atherosclerosis.
Oxidative stress is considered an important factor in atherogenesis. Mammalian cells have a complex network of antioxidants such as catalase, superoxide dismutase, and glutathione peroxidase. However, the mechanisms that regulate the cellular redox state in the vessel wall remain unclear. Recent study has shown that thioredoxin, a thiol-disulfide oxidoreductase, is expressed in atherosclerotic plaques of human carotid arteries. In this study, we investigated the localization and expressional change of glutaredoxin and thioredoxin, two important members of the thiol-disulfide oxidoreductases, in autopsy samples of human coronary arteries. In nonatherosclerotic coronary arteries, glutaredoxin was expressed in endothelial cells, in fibroblasts of the adventitia, and most intensely in medial smooth muscle cells. Interestingly, in atherosclerotic lesions such as hypercellular lesions, the infiltrating macrophages highly expressed glutaredoxin. The expressional pattern of thioredoxin was quite similar to that of glutaredoxin. Western blot analysis demonstrated that hydrogen peroxide stimulated the expression of glutaredoxin in a time- and dose-dependent manner in cultured human coronary artery smooth muscle cells. Fluorescence microtopography with dihydroethidium demonstrated that the generation of reactive oxygen species was associated with the expression of glutaredoxin. These results suggest the possible involvement of thiol-disulfide oxidoreductases in antioxidant protection in human coronary arteries.
Aged↗