Biomedical subjects

S Harada

Publications and source records attributed to S Harada.

At least 271 records · Page 15Linked to original sources

PubMed1995

Biological responses in Caenorhabditis elegans to high magnetic fields.

Here we describe a device for testing possible influences of high magnetic fields on biological processes, by which alternating-current magnetic stimuli as high as 1.7 T can be administered. Experiments with a simple multicellular organism, the nematode Caenorhabditis elegans, revealed that intermittent exposure to the magnetic fields modestly inhibited the animal's reproduction as well as its post-embryonic development, and caused a marked but transient derangement in its locomotory behavior. Available evidence indicates that alternating high magnetic fields can elicit both chronic and acute biological effects, but that the effects may be well tolerated or compensated for by the living organism.

Animals↗

PubMed1995

Demonstration of specific insulin binding to cytosolic proteins in H35 hepatoma cells, rat liver and skeletal muscle.

We previously demonstrated that internalized insulin enters the cytoplasm before accumulating in nuclei of H35 rat hepatoma cells. This finding raises the possibility that insulin may interact with cytosolic proteins in addition to insulin-degrading enzyme (IDE). In the present study, cytosol from H35 hepatoma cells, rat liver or muscle was incubated with A14- or B26-125I-insulin at 4 degrees C for 5-120 min in the absence or presence of 25 micrograms/ml unlabelled insulin. 125I-insulin was cross-linked to cytosolic proteins by disuccinimidyl suberate and analysed by reducing or non-reducing SDS/PAGE and autoradiography. Our results demonstrate the presence of both tissue-specific and common cytosolic proteins which specifically bind insulin. In muscle cytosol, only two proteins of 27 and 110 kDa were specifically labelled with B26-125I-insulin. Seven major bands, of 27, 45, 55, 60, 76, 82 and 110 kDa, were labelled in rat liver cytosol. Detection of cytosolic insulin-binding proteins in H35-cell cytosol was dependent on cell-culture conditions. Labelling in cytosol from serum-deprived cells was decreased or absent compared with cytosol prepared from serum-fed or serum-deprived cells treated with 100 ng/ml insulin for 1 h before preparation of the cytosol, in which six bands, of 32, 41, 45, 55, 82 and 110 kDa, were specifically labelled with B26-125I-insulin. This result suggests that the concentration or binding activity of some cytosolic insulin-binding proteins is rapidly regulated. Labelling of both rat liver and H35 cytosolic insulin-binding proteins was time-dependent, and decreased or disappeared at 120 min in parallel with the degradation of labelled insulin. Fewer bands were specifically labelled with A14-125I-insulin than with B26-125I-insulin. The number of labelled bands observed under reducing and non-reducing conditions was not different in any of the cytosols. The 110 kDa band in all cytosols was identified as IDE by Western-blot analysis; the other proteins did not react with anti-IDE antibody and remain unidentified. 1,10-Phenanthroline (2 mM) increased IDE labelling, but decreased the labelling of 82 and 27 kDa bands. The marked difference in the number of cytosolic insulin-binding proteins in muscle and either H35 cells or liver suggests both that the labelling is specific and that these proteins serve a function and may be involved in some heretofore unknown mechanism of the signalling pathway by which insulin regulates cell growth or differentiation.

Animals↗

PubMed1995

Reversal of drug sensitivity in MDR subline of P388 leukemia by gene-targeted antisense oligonucleotide.

We attempted to reverse multidrug resistance (MDR) by treatment with 25-mer antisense phosphorothioate oligonucleotide. The phosphorothioate analogs, the sequences of which are sense or antisense to the initiation codon of mouse mdr1 mRNA, were tested against murine leukemic P388/S and adriamycin-resistant P388/ADR cell lines. A weak inhibitory effect on the growth of P388/S and P388/ADR cells was observed at a sense and antisense oligonucleotide concentration of 30 microM. Using the monoclonal antibody to P-glycoprotein and a flow cytometry technique, we showed that the level of expression of P-glycoprotein in P388/ADR cells treated with antisense oligonucleotide was lower than when treated with sense oligonucleotide. The antisense oligonucleotide potentiated the growth-inhibitory effect of vinblastine on P388/ADR cells, whereas sense oligonucleotide did not. This was accompanied by an increase in vinblastine retention in the cells. The reversal of the resistance by antisense oligonucleotide was increased by the combination with 1 microM verapamil. These results suggest that the antisense oligonucleotide and low dose verapamil may be useful in circumventing the resistance to anticancer drugs of MDR tumors.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1995

Trends in nonmelanoma skin cancer in Japan.

We compared the prevalence of skin cancer and solar keratosis (SK) in patients who attended 26 Japanese university hospitals between 1976-1980 with those who attended between 1986-1990 to investigate whether the incidence of skin cancer has increased or not. Age-adjusted incidence rates of basal cell carcinomas (BCC) and SK, but not squamous cell carcinomas (SCC), from 1986-1990 were higher than those from 1976-1980. In addition, a population-based incidence study was conducted in Kasai City, Hyogo prefecture, to establish the frequency of skin cancer and SK. A total of 4736 people over 20 years of age were examined. Two BCC and 36 SK patients were identified clinically and histopathologically. SCC was not found. Age-adjusted incidence rates of BCC and SK per 100,000 were 16.5 and 486.1, respectively. The BCC incidence rate in Kasai City was significantly higher than the incidence of Japanese nonmelanoma skin cancer (NMSC) reported by Gordon in 1976. Further, subjects classified as skin type I showed statistically higher SK prevalence rates compared to skin types II and III. The present study indicates that the prevalences of NMSC and SK in Japanese have increased during the last three decades and that skin type I may be a risk factor for NMSC in Japanese.

Adolescent↗

PubMed1995

Alcohol and aldehyde dehydrogenase genotypes and drinking behavior of Chinese living in Shanghai.

Alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH), the principal enzymes responsible for oxidative metabolism of ethanol, exist in multiple, genetically determined molecular forms. Widely different kinetic properties in some of these isozymes account for the individual differences in alcohol sensitivity. In this study we used the polymerase chain reaction/restriction fragment length polymorphism method to determine the genotypes of the ADH2 and ALDH2 loci of alcoholic and nonalcoholic Chinese living in Shanghai. We also investigated the subjects' drinking patterns by means of semistructured interviews. The alcoholics had significantly lower frequencies of the ADH2(2) and ALDH2(2) alleles than did the nonalcoholics, suggesting the inhibitory effects of these alleles for the development of alcoholism. In the nonalcoholic subjects, ADH2(2) had little, if any, effect, despite the significant effect of the ALDH2(2) allele in decreasing the alcohol consumption of the individual. Taken together, these results fit the proposed hypothesis for the development of alcoholism, i.e., drinking behavior is greatly influenced by the individual's genotypes of alcohol-metabolizing enzymes, and the risk of becoming alcoholic is proportionate with the ethanol consumption of the individual.

Adult↗

PubMed1995

Chlordiazepoxide attenuates stress-induced activation of neurons, corticotropin-releasing factor (CRF) gene transcription and CRF biosynthesis in the paraventricular nucleus (PVN).

Corticotropin-releasing factor (CRF) plays a role in coordinating endocrine, autonomic, and behavioral responses to stressful stimuli. Benzodiazepines exert many effects which oppose those of CRF, including anxiolysis and suppression of the pituitary-adrenal axis. In the present study, we employed in situ analysis of CRF heteronucleous RNA (hnRNA) and c-fos mRNA to assess stimulus-induced CRF gene transcription rate following stress and its modulation by chlordiazepoxide (CDP). Male albino rats were exposed to restraint stress for 30 min and sacrificed 30 and 120 min after the onset of stress. Either CDP or vehicle was given intraperitoneally 60 min before stress. To determine plasma ACTH levels by immunoradiometric assay, another group of rats was decapitated 10 min after the onset of restraint stress. Restraint stress induced rapidly and significantly c-fos mRNA and CRF hnRNA expression in the PVN at the 30 min time point. Increases in both RNA copies were significantly inhibited by administration of CDP at doses of 5 and 10 mg/kg. CRF mRNA concentrations were increased significantly in the PVN 120 min after stress and again, CDP attenuated significantly these increases in the PVN. The plasma ACTH increase in response to stress was inhibited significantly by CDP administration at every dose tested. CDP did not change CRF mRNA levels in the non-stressed animal.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1995

Frequency of glutathione-S-transferase 1 gene deletion and its possible correlation with cataract formation.

This experiment was conducted to investigate the possible association between an increased frequency of glutathione-S-transferase (GST)1 gene deletion and the presence of cataracts in elderly patients. Genomic DNA was isolated from blood samples obtained from 138 elderly patients who had undergone cataract surgery, and from 62 random blood donors. All subjects lived in the same geographic area (Ibaraki Prefecture, Japan). The DNA sequences among three different exon ranges (exons 3-5, exons 4-5 and exons 5-6) of the GST1 gene were amplified by the polymerase chain reaction (PCR) technique to determine if GST1 gene deletion occurred. Cataract patients had a significantly higher frequency of GST1 gene deletion than random controls did (P < 0.001, odds ratio = 2.91, 1.56-5.44; 95% of confidence interval). Mean age of cataract patients lacking GST1 gene was significantly younger (n = 101, mean age = 70.4, s.d. = 10.2) than that of patients possessing the GST1 gene (n = 37, mean age = 75.0, s.d. = 8.7) (P < 0.02). These results show that the deletion of the GST1 gene may be one of determinants of genetic susceptibility to cataractgenic agents.

Aged↗

PubMed1995

Nuclear localization of the truncated hepatitis C virus core protein with its hydrophobic C terminus deleted.

The core protein of hepatitis C virus (HCV) is considered to be cleaved from the N terminus of the large precursor polyprotein by cellular signalase. The HCV cDNA encoding the core protein was expressed (i) in monkey COS cells by a plasmid expression vector driven by the SR alpha promoter, and (ii) in insect cells by a recombinant baculovirus. The expressed product had an M(r) of 22,000 and was located in the cytoplasm. When the C-terminal hydrophobic domains were deleted, however, the truncated core proteins were translocated into the nucleus. The truncated core proteins were located in the nucleus even when they were expressed as a fusion protein with E. coli beta-galactosidase, which is essentially localized in the cytoplasm. Plasmids containing HCV cDNAs with a deletion in one of the regions encoding clusters of basic amino acids were expressed in COS cells and the localization of the core protein was examined. The residues PRRGPR were suggested to play an important role in nuclear localization. HCV is an RNA virus and its life cycle was originally considered to be confined to the cytoplasm; the present study, however, suggests that the HCV core protein can translocate into the nucleus under certain circumstances.

Amino Acid Sequence↗

PubMed1995

Establishment of a cell line constitutively expressing E2 glycoprotein of hepatitis C virus and humoral response of hepatitis C patients to the expressed protein.

A Chinese hamster ovary cell line was established which abundantly expresses the second envelope protein (E2) of hepatitis C virus under the control of an exogenous promoter. The expressed E2 protein was found to be a glycoprotein of 58 kDa by immunoprecipitation with sera from patients that had chronic hepatitis C. Using this cell line as antigen in immunofluorescence tests, as high as 93% of patients with non-A non-B hepatitis had antibodies against E2 protein. In Western blots using SDS-denatured E2 protein, however, the detectability of the antibody was drastically reduced to 30%. Immunoprecipitation assays and ELISA, using both native and denatured E2 protein, revealed that antibodies to E2 protein were present in most of the chronic hepatitis C patients and that they reacted only to the native forms.

Animals↗

PubMed1995

Changing pattern of mental health utilization in Hachijojima.

We have reviewed retrospectively 229 patients interviewed by the mental health team in the Hachijojima since the establishment of our services in 1987. The 6 year observation period was classified into three chronological stages for the purposes of identifying any change in the utilization pattern of the mental health services of the island. The rate of schizophrenia was observed to have decreased from 48% in the first 2 years of the study period to 28% in the last 2 years; while that of other mental disorders increased yearly. Interestingly, an increasing number of patients were referred to our services by professionals in areas other than mental health (from 42 to 77%), and patients without any history of psychiatric treatment were more frequently seen (from 36 to 75%) in the later part of the study period. These changes suggest the importance of access to mental health services.

Adolescent↗

PubMed1995

Stress-induced activation of neuronal activity and corticotropin-releasing factor gene expression in the paraventricular nucleus is modulated by glucocorticoids in rats.

Intronic in situ hybridization methodology provides a means of determining the rate of gene transcription under basal and stimulated conditions. In the present study, we have used intronic in situ hybridization to the corticotropin-releasing factor (CRF) gene to measure hypothalamic CRF gene transcription after stress as well as its modulation by glucocorticoids. Using this and conventional exonic in situ hybridization we examined the time course of changes in c-fos mRNA, and CRF heteronuclear RNA (hnRNA) and mRNA concentrations in the paraventricular nucleus (PVN) of male Wistar rats after restraint stress. In addition, we determined the effects of adrenalectomy and dexamethasone administration on c-fos and CRF gene expression in the PVN. Restraint stress induced a rapid induction (within 5 min) of c-fos mRNA and CRF hnRNA expression in the PVN. Both RNA concentrations peaked at 30 min then decreased and were undetectable 2 h after stress onset. In contrast, the concentration of CRF mRNA increased gradually and a significant elevation was first detected 60 min after the beginning of stress. Adrenalectomy augmented and dexamethasone pretreatment inhibited c-fos mRNA, CRF hnRNA, and mRNA induction after stress. The data suggest that stress-induced activation of neurons, CRF gene transcription, and CRF synthesis in the PVN are modulated by glucocorticoids.

Adrenalectomy↗

PubMed1995

Electron microscopic visualization of insulin translocation into the cytoplasm and nuclei of intact H35 hepatoma cells using covalently linked Nanogold-insulin.

Insulin affects numerous metabolic processes as well as nuclear events such as gene transcription. Our previous ultrastructural and biochemical studies demonstrated insulin accumulation in nuclei of cultured and rapidly proliferating cells, and biochemical evidence suggested that insulin entered the cell cytoplasm before accumulating in the nucleus. The present study was undertaken to develop a covalently linked electron-dense insulin complex that could be used to visualize the intracellular translocation of insulin and confirm that insulin enters the cytoplasm of cells. Insulin was cross-linked to 1.4-nm diameter Nanogold particles. The complex binds to the plasma membrane insulin receptor, is biologically active, and is degraded by cellular insulin-degradative enzymes. Ultrastructural analysis after silver intensification of the gold particles confirmed that insulin internalization culminates in the translocation of some internalized insulin to the cytoplasm and nuclei. When cytoplasmic insulin-degrading enzyme (IDE) activity was inhibited with 1,10-phenanthroline, an increase in the number of cytoplasmic and nuclear Nanogold-insulin particles was observed. The results of this and previous studies suggest that 1) the translocation of insulin to the cytoplasm, 2) the regulation of insulin degradation in the cytoplasm by IDE, 3) the possible interaction of insulin with cytoplasmic proteins other than IDE, and 4) the subsequent accumulation of intact insulin or insulin complexed with cytoplasmic proteins in nuclei may play a role in insulin's regulation of gene transcription and cell proliferation.

Adipose Tissue↗

PubMed1995

Synthesis of saframycins. X. Transformation of (-)-saframycin A to (-)-saframycin Mx type compound with the structure proposed for saframycin E.

Treatment of (-)-saframycin A (1a) with selenium oxide in acetic acid afforded (-)-saframycin G (1g), and a catalytic reduction and regioselective oxidation sequence afforded the saframycin Mx type compound (3). We applied this methodology to the transformation of (+/-)-5-hydroxysaframycin B (11) to the hydroquinone (1e). Acetylation of 1e with acetic anhydride in pyridine gave the triacetate (13), which is identical with the triacetyl derivative of natural saframycin E.

Anti-Bacterial Agents↗

PubMed1995

Male reproductive toxicity study of nefiracetam in rats.

Sprague-Dawley male rats were administered nefiracetam orally at daily doses of 500 and 1500 mg/kg/day for 4 or 9 weeks. Although the copulation index was not affected by nefiracetam treatment, the fertility index was extremely low in the 1500 mg/kg/day group for both treatment periods. This high dose group consistently exhibited decreased testicular weights. Epididymal and prostate weights were also reduced in the 1500 mg/kg/day group after both 4- and 9-week treatments and in the 500 mg/kg/day group after the 9-week treatment. Severe degenerative changes such as degeneration of germ cells, loss of germ cells and atrophy of seminiferous tubules were observed in all rats of the 1500 mg/kg/day groups after both 4 and 9 weeks of treatment. Retention of spermatids in stage IX, X and XI seminiferous tubules was also noted after the 4- and 9-week treatments at 500 mg/kg/day. The testicular sperm head counts were markedly decreased following the 4- and 9-week treatments at 1500 mg/kg/day, and mildly reduced after the 4-weeks treatment at 500 mg/kg/day. From these results it is concluded that histopathological examination and the testicular sperm head count method are highly useful for detecting testicular toxicity and that testicular lesions caused by nefiracetam can be detected after 4 weeks of exposure.

Animals↗

PubMed1995

[Reproductive toxicity of iodixanol, a new non-ionic, iso-tonic contrast medium in rats and rabbits].

The reproductive toxicity of iodixanol, a non-ionic isotonic contrast medium, was investigated in rats and rabbits. Iodixanol was administered intravenously to male rats at doses of 0.3, 1.0 or 2.0 gI/kg/day for 80 days or more until successful copulation. Female rats were treated with iodixanol at the same dose levels for at least 14 days prior to and in the early stage of pregnancy. No remarkable adverse effects on fertility were noted at any dose level. Increased water consumption and swollen paws/face were observed among male rats receiving > 1.0 gI/kg/day. In addition, decreased food consumption was observed in males of the 2.0 gI/kg/day group. In female rats, no remarkable adverse effects were noted at any dose level. Iodixanol elicited no evidence of teratogenicity when administered during the fetal organogenesis period to pregnant rats and rabbits at doses of 0.3, 1.0 or 2.0 gI/kg/day. No remarkable changes were observed in rat or rabbit fetuses, nor in dams of any dose group. In a perinatal and postnatal dosing study in rats using doses of 0.3, 1.0 or 2.0 gI/kg/day, iodixanol did not adversely affect the reproductive performance of the treated parent animals and pre- and postnatal development of their offspring.

Abnormalities, Drug-Induced↗

PubMed1995

Belactins A and B, new serine carboxypeptidase inhibitors produced by Actinomycete. I. Taxonomy, production, isolation and biological activities.

Belactins A and B, new inhibitors of serine carboxypeptidase were discovered in the fermentation broth of Saccharopolyspora sp. MK19-42F6. They were purified by ethyl acetate extraction, silica gel chromatography, Sephadex LH20 chromatography, Capcellpak C18 SG120 reversed phase HPLC and centrifugal partition chromatography (CPC) following their inhibitory activity against carboxypeptidase Y (CP-Y). The inhibition constants (Ki) of belactins A and B against CP-Y are 0.14 and 0.27 microM respectively. Belactins A and B have highly specific inhibitory activities for CP-Y among various peptidases, have no antimicrobial activities at 100 micrograms/ml and have low toxicities.

Benzamides↗

PubMed1995

TAN-1511 A, B and C, microbial lipopeptides with G-CSF and GM-CSF inducing activity.

The microbial lipopeptides, TAN-1511 A, B and C, were isolated from the culture broth of Streptosporangium amethystogenes subsp. fukuiense AL-23456. Their structures were elucidated on the basis of their reactions and spectroscopic analyses. These lipopeptides were mixtures of molecules having different lengths of fatty acids. The metabolites stimulated the proliferation of bone marrow cells from BALB/c female mice at very low concentrations (concentration giving 30% increase: A and B, 0.313 ng/ml; C, 1.25 ng/ml). We confirmed that chemically synthesized TAN-1511 A analogue [(2R,6R)-2-tetradecanoylamino-6,7- bis(hexadecanoyloxy)-4-thiaheptanoyl-Gly-Gly-Gly-Glu-Thr-Thr -OH] stimulated the proliferation of bone marrow cells in a manner similar to that of natural TAN-1511 A. This analogue induced the secretion of both granulocyte colony stimulating factor (G-CSF) and granulocyte-macrophage colony stimulating factor (GM-CSF), and potentiated the generation of Gr-1 positive cells in the bone marrow cell culture. Moreover, it effected the G-CSF mediated restoration of granulocytopoiesis in a murine leukopenia model.

Amino Acid Sequence↗

PubMed1995

Synthesis and biological activities of TAN-1511 analogues.

TAN-1511 analogues were synthesized and their effects on the proliferation of bone marrow cells were examined. To exert potent activity the following conditions are necessary: the configuration of the 2-amino-6,7-dihydroxy-4-thiaheptanoic acid moiety must be (2R,6R), long chain acyl groups (C14 to C18) must be bound to both hydroxyl groups, the amino group must be free or acylated with the long chain fatty acid (ca. C14) and the peptide moiety must have glutamic acid as a component. Among the synthesized compounds, trisodium (2R,6R)-2-amino-6,7-bis (hexadecanoyloxy)-4-thiaheptanoyl glycyl glutamyl glutamate, which has improved solubility, was effective in experimental leukocytopenia in mice.

Amino Acid Sequence↗