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M Tokunaga

Publications and source records attributed to M Tokunaga.

At least 73 records · Page 4Linked to original sources
PubMed1999

Ruthenium-Catalyzed Intermolecular Hydroamination of Terminal Alkynes with Anilines: A Practical Synthesis of Aromatic Ketimines.

Without solvent and in the open air, the hydroamination of phenylacetylene with aniline in the presence of [Ru(3)(CO)(12)]/NH(4)PF(6) proceeds with high regioselectivity [Eq. (1)]. Simple distillation gives a product with greater than 99 % purity in an excellent yield. The application of this method to a new two-component reaction gives the important class of substances: the quinolines.

Journal Article↗
PubMed1999

Encephalopathy caused by visceral larva migrans due to Ascaris suum.

We described a patient with encephalopathy associated with visceral larva migrans (VLM) caused by Ascaris suum. He suffered from drowsiness, quadriparesis, eosinophilia and elevated serum IgE levels. Brain magnetic resonance (MR) imaging revealed multiple cerebral cortical and white matter lesions. Serological tests indicated recent infection with A. suum. Pulse steroid therapy relieved the patient's central nervous system symptoms and marked improvement of lesions on brain MR images. We concluded that the encephalopathy in this patient was probably caused by VLM due to Ascaris suum.

Adult↗
PubMed1999

A single myosin head moves along an actin filament with regular steps of 5.3 nanometres.

Actomyosin, a complex of actin filaments and myosin motor proteins, is responsible for force generation during muscle contraction. To resolve the individual mechanical events of force generation by actomyosin, we have developed a new instrument with which we can capture and directly manipulate individual myosin subfragment-1 molecules using a scanning probe. Single subfragment-1 molecules can be visualized by using a fluorescent label. The data that we obtain using this technique are consistent with myosin moving along an actin filament with single mechanical steps of approximately 5.3 nanometres; groups of two to five rapid steps in succession often produce displacements of 11 to 30 nanometres. This multiple stepping is produced by a single myosin head during just one biochemical cycle of ATP hydrolysis.

Actins↗
PubMed1999

Deletions and single-base mutations within the carboxy-terminal region of the latent membrane protein 1 oncogene in Epstein-Barr virus-related gastric cancers of southern Japan.

The 30-base pair (bp) deletion of the cytoplasmic carboxy-terminal domain of the latent membrane protein 1 (LMP-1) gene was analyzed in 37 frozen tissues from patients with Epstein-Barr virus (EBV)-related gastric cancer and 18 throat washings from healthy adults in southern Japan. The 30-bp deletion was identified in 33 (91.7%) of 36 specimens of EBV-related gastric cancers and in 15 (83.3%) of 18 throat washings from healthy adults. In one case of gastric cancer, an additional 9-bp deletion was identified downstream of the 30-bp deletion. From the last transmembrane domain to the end of the carboxy terminal of LMP-1, mutations were examined in 37 cases of gastric cancers and in three cases of throat washings. Twenty-eight nonsilent mutations were identified in this region of EBV-related gastric cancer and throat washings. Five nonsilent mutations at positions 168,755, 168,746, 168,687, 168,357, and 168,355 were identified in all 30-bp-deleted cases of EBV-related gastric cancers and throat washings. However, these nonsilent mutations were not identified in three patients without the 30-bp deletion. Although the deletion and single-base mutations of the LMP-1 gene in gastric cancers and throat washings were similar to those of nasopharyngeal carcinoma in Taiwan and China, more single-base mutations were found in southern Japan. These data indicate that high prevalence of the 30-bp deletion of the LMP-1 gene in gastric cancers may reflect the prevalence of the deletion variant in the normal population in southern Japan.

DNA Mutational Analysis↗
PubMed1999

Identification and partial purification of DnaK homologue from extremely halophilic archaebacteria, Halobacterium cutirubrum.

The levels of synthesis of six proteins were increased at elevated growth temperature of the extremely halophilic archaebacterium Halobacterium cutirubrum. One of these proteins, with an apparent molecular mass of 97 kDa on sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE), bound to an ATP-agarose column in the presence of 4 M NaCl, but not in the absence of salt, indicating that this protein retained its ATP-binding activity only at high salt concentration. The NH2-terminal sequence of this protein and the internal sequences of the tryptic peptides covering 1/3 of the total number of residues coincided with that deduced from the nucleotide sequence of the dnaK gene isolated from H. cutirubrum. The results strongly suggest that this apparent 97-kDa protein is the gene product of dnaK, although the molecular mass calculated from the nucleotide sequence is only 68,495, much smaller than the value of this protein determined by SDS-PAGE. Ferguson plot analysis indicated that this protein showed anomalous mobility on SDS-PAGE. We have purified DnaK homologue to greater than 90% homogeneity with stepwise elution from an ATP-agarose column.

Amino Acid Sequence↗
PubMed1999

[Detection and characterization of gastric carcinoma associated with epstein-barr herpes virus].

The present investigation was carried out to estimate the incidence of Epstein-Barr virus (EBV)-associated cases among gastric carcinoma (GC) patients in Russia. Carcinoma specimens from 184 patients with GC treated at the Cancer Research Center were investigated by EBV encoded RNA-1 (EBER-1) in situ hybridization. Seventeen (9.24%) cases showed uniform EBER-1 expression restricted to the carcinoma cells. Hybridized signals were not detected in the non-neoplastic gastric epithelium. EBV involvement was significantly more frequent among males, especially in the tumors belonging to less differentiated types (moderately differentiated tubular adenocarcinomas and poorly differentiated solid adenocarcinomas) and located in the upper stomach (cardia and middle part). Most EBV-positive GCs were characterized by great lymphoid compartment involvement. The findings of the distribution of EBV-positive. GCs by sex, site, and histology are similar to those in Japan; however, the detection rate of EBV-positive cases in Russia is higher than that in Japan (6.7%) and lower than that in the USA (16%).

Adenocarcinoma↗
PubMed1999

Epstein-Barr virus-associated gastric carcinoma in Mexico: analysis of 135 consecutive gastrectomies in two hospitals.

Epstein-Barr virus (EBV) has been implicated in the genesis of gastric carcinoma. The presence of clonal episomal viral forms in the nuclei of neoplastic gastric epithelial cells suggests that viral infection occurs before the development of gastric carcinoma. Mexico is a country at high risk for gastric cancer-it is the second cause of death among patients who die of cancer in that country. A series of 135 consecutive non-selected gastrectomies from two hospitals in Mexico City were analyzed to search for EBV in gastric carcinomas. EBV-encoded small non-polyadenylated RNA (EBER) in situ hybridization was performed on 5-microm paraffin-embedded tissue sections. Age, gender, anatomical site, histological type, and invasiveness of gastric carcinomas were obtained from the records in the corresponding Departments of Pathology. Eleven (8.15%) of the 135 cases were EBER-1-positive gastric carcinomas. Six occurred in males and five in females. In three women, the neoplasia was localized in the antrum. Five of the 11 cases were lymphoepithelioma-like carcinomas and, in two of them, an unusual foreign body-type inflammation was observed. Environmental factors could influence the distinctive pathologic features of EBV-associated gastric carcinoma in the Mexican population.

Adult↗
PubMed1998

Molecular cloning of the dnaK locus, and purification and characterization of a DnaK protein from Bacillus brevis HPD31.

Using part of the dnaK gene from Bacillus subtilis as a probe, a 4. 4-kbp SacI-BglII fragment of chromosomal DNA of Bacillus brevis, a protein-hypersecreting bacterium, was cloned. Nucleotide sequencing revealed 3 open reading frames in the order of grpE-dnaK-dnaJ homologues. We purified DnaK protein to homogeneity from B. brevis HPD31 harboring a multi-copy dnaK expression plasmid. Purified DnaK showed ATPase activity which was synergistically stimulated 14-fold by the addition of glutathione S-transferase-DnaJ and glutathione S-transferase-GrpE fusion proteins. DnaK hydrolyzed not only ATP but also CTP, UTP, and GTP at about 40% of the efficiency of ATP. The specific activity of DnaK-ATPase was 7.25x10-3 unit/mg protein (the turnover number against ATP was 0.47 min-1) under our assay conditions. The DnaK dimers dissociated into monomers on addition of ATP, GTP, CTP, UTP and ATPgammaS, but not ADP or AMP. DnaK formed a stable complex with permanently unfolded carboxymethylated alpha-lactalbumin but not with native alpha-lactalbumin, and this complex was dissociated by addition of ATP/Mg. Formation of this complex was inhibited in the presence of inorganic phosphate.

Adenosine Triphosphatases↗
PubMed1998

Simultaneous observation of individual ATPase and mechanical events by a single myosin molecule during interaction with actin.

We have developed a technique that allows mechanical and ligand-binding events in a single myosin molecule to be monitored simultaneously. We describe how steps in the ATPase reaction are temporally related to mechanical events at the single molecule level. The results show that the force generation does not always coincide with the release of bound nucleotide, presumably ADP. Instead the myosin head produces force several hundreds of milliseconds after bound nucleotide is released. This finding does not support the widely accepted view that force generation is directly coupled to the release of bound ligands. It suggests that myosin has a hysteresis or memory state, which stores chemical energy from ATP hydrolysis.

Actin Cytoskeleton↗
PubMed1998

Characterization of deletion mutations in the carboxy-terminal peptide-binding domain of the Kar2 protein in Saccharomyces cerevisiae.

Hsp70 is structurally composed of three domains, an amino-terminal ATPase domain, a proximal 18 kDa peptide-binding domain and a distal 10 kDa carboxy-terminal (C-terminal) domain. To dissect the functional significance of the distal 10 kDa domain, and the boundary region between the proximal and distal C-terminal domains of Kar2p in vivo in Saccharomyces cerevisiae, we constructed a series of plasmids which were truncated or had internal deletion mutations in this region. We found that all these mutations are recessive, and that the distal 10 kDa C-terminal domain, including the HDEL ER-retention sequence, is not essential for cell growth, although the major role of this 10 kDa C-terminal domain is due to the function of the HDEL ER-retention signal. We also found that the Kar2p region (Thr492-Thr512), corresponding to the beta 8-sheet in the peptide-binding domain, which constitutes the bottom plate of the binding pocket in E. coli DnaK, is essential for cell viability, and that the following Kar2p region (Glu513-Lys542), corresponding to alpha-helices A and B of E. coli DnaK, which was proposed to compose the lid of the binding pocket, is critical but not essential for yeast cell growth. This was further supported by the fact that the latter deletion showed a fully reversible ts phenotype in its growth and only a slight inhibitory effect on the secretion of alpha-amylase at non-permissive temperature.

Binding Sites↗
PubMed1998

Single muscle fiber analysis of myoclonus epilepsy with ragged-red fibers.

We examined two muscle biopsy specimens from a proband and her mother with myoclonus epilepsy with ragged-red fibers (MERRF), both obtained at an interval of about 10 years, using histochemistry, in situ hybridization, and single-fiber polymerase chain reaction. Total (wild-type and mutant) mitochondrial DNAs (mtDNAs) were greatly increased in ragged-red fibers (RRF) over non-RRF in all muscle specimens analyzed. The proportion of mutant mtDNA was also significantly higher in RRF than in non-RRF. By comparing the first and second muscle biopsied specimens in each patient, we found that while the proportion of RRF, cytochrome coxidase deficient fibers, and mutant DNA in muscle changed over a 10-year period, the proportion of wild-type and mutant mtDNAs in RRF and in non-RRF was similar between the two specimens. These results suggest that the ratio of wild-type to mutant mtDNAs in RRF and non-RRF in MERRF is at a steady state level in each muscle fiber, without replicative advantage of mutant mtDNA.

Adult↗
PubMed1998

Upregulation of hypothalamic nitric oxide synthase gene expression in streptozotocin-induced diabetic rats.

Plasma arginine vasopressin (AVP) is known to be elevated in patients with uncontrolled insulin-dependent diabetes mellitus who have plasma hyperosmolality with hyperglycaemia. Although osmotic stimuli cause an increase in nitric oxide synthase (NOS) activity as well as synthesis of AVP and oxytocin in the paraventricular (PVN) and supraoptic nuclei (SON), it is not known whether NOS activity in the hypothalamus changes in the diabetic patients who have plasma hyperosmolality with hyperglycaemia caused by insulin deficiency. Expression of the neuronal (n) NOS gene in the PVN and SON in streptozotocin (STZ)-induced diabetic rats was investigated by using in situ hybridization histochemistry and NADPH-diaphorase histochemical staining. Four weeks after intraperitoneal (i. p.) administration of STZ, male Wistar rats developed hyperglycaemia and plasma hyperosmolality. The expression of nNOS gene and NADPH-diaphorase staining in the PVN and SON remarkably increased in STZ-induced diabetic rats compared to control rats. Three weeks after administration of STZ, the diabetic rats were subcutaneously treated with insulin for 1 week, which resulted in significant suppression of the induction of nNOS, AVP and oxytocin gene expression in the PVN and SON. Furthermore, the induction of nNOS gene expression in the PVN and SON was suppressed in STZ-induced diabetic rats treated with phlorizin and diet to normalize hyperglycaemia without insulin treatment. These results suggest that upregulation of nNOS gene expression as well as AVP and oxytocin gene expression in the PVN and SON in STZ-induced diabetic rats may be associated with hyperglycaemia and plamsa hyperosmolality.

Animals↗
PubMed1998

Skin tumor risk among atomic-bomb survivors in Japan.

OBJECTIVES: Elevated risks of skin cancer following high doses of ionizing radiation have long been known. Recent reports on atomic-bomb survivors indicate that nonmelanoma skin cancer can be induced at low to medium doses. We studied atomic-bomb survivors to determine the effects of radiation on specific histologic types of skin cancer and to describe the dose-response relationship. METHODS: Cases of melanoma, nonmelanoma skin cancers, and Bowen's disease were ascertained between 1958 and 1987 for the 80,000 cohort members through the population-based Hiroshima and Nagasaki (Japan) tumor registries augmented by searches of other records. RESULTS: An excess of basal cell carcinoma (n = 80), with some suggestion of a non-linear dose-response, was observed. The excess risk decreased markedly as age at exposure increased, and there was no evidence for an interaction between ionizing and ultraviolet radiation. No dose-response was found for squamous cell carcinoma (n = 69). The excess relative risk point-estimates were large, but statistically nonsignificant for both melanoma (n = 10) and Bowen's disease (n = 26). CONCLUSIONS: The basal layer of the epidermis appears to be quite sensitive to radiation carcinogenesis, particularly at a young age. The suprabasal layer seems to be more resistant, as shown by the lack of an association for squamous cell carcinomas.

Adolescent↗
PubMed1998

RNA from decades-old archival tissue blocks for retrospective studies.

The validity of molecular studies using DNA and RNA extracted from decades-old formalin-fixed and paraffin-embedded tissue blocks has been demonstrated. The quality and usability of DNA and RNA from archival tissues are modified by various factors, such as the fixative, the fixation time, and the postmortem time. However, in contrast to DNA, there are no comprehensive studies quantitatively addressing the feasibility of RNA from old (more than 10 years) archival samples. This study examined the integrity of RNA extracted from 738 autopsy liver and 63 autopsy thyroid cancer tissue blocks procured during a span of nearly four decades, beginning in 1952 and ending in 1989, from the atomic bomb survivors. The integrity of RNA was assessed by amplification of c-BCR messenger RNA (mRNA) between two sequential exons with an intervening intron by reverse-transcription polymerase chain reaction (RT-PCR). The integrity of RNA was influenced by the age of the samples and the postmortem time, but not by the formalin-fixation period. It was possible to amplify more than 60% of the samples. Using these RNAs, the HCV genome in liver cancers and the H4-RET gene in thyroid cancers were detectable. This study illustrates the possibility of molecular studies using RNA from routinely prepared paraffin blocks stored for long periods and provides the statistics and critical factors to consider in assessing the feasibility of such contemplated studies.

Drosophila Proteins↗
PubMed1998

Expression and purification of cytokine receptor homology domain of human granulocyte-colony stimulating factor receptor in Escherichia coli.

In an attempt to generate a stable non-glycosylated cytokine receptor homology (CRH) domain (Tyr97-Ala309) of human granulocyte-colony stimulating factor (G-CSF) receptor, two free cysteines in the CRH domain were converted to serine by site-directed mutagenesis. Taking advantage of the tight regulation for the expression of T7 RNA polymerase, the mutated CRH domain was successfully expressed in Escherichia coli (E. coli) with a pelB signal sequence at its NH2-terminus and with a His tag at its COOH-terminus. The processed and secreted CRH domain after solubilization and in vitro refolding retained G-CSF binding activity, and its yield (approximately 40 micrograms/30 ml culture) was more than 100-fold higher than that of the mouse CRH domain expressed by the MalE fusion system in E. coli.

Amino Acid Sequence↗
PubMed1998

[The influence of sleep breathing disorder on growth hormone secretion in children with tonsil hypertrophy].

INTRODUCTION: Recently it has been revealed that Obstructive Sleep Apnea Syndrome has an influence on the endocrine system, especially to the secretion of growth hormone (GH). It was previously reported that secretion of GH has a circadian rhythm along with a correlation with slow wave sleep. Normally in a pediatric patient, after 90 min of sleep the brain wave shows patterns of deep sleep, and a matching peak in GH secretion is observed. We have studied the effect of GH secretion in sleep breathing disorder of pediatric patients by observing the pre- and post-operative difference in GH secretion in children with tonsillar hypertrophy who had tonsillectomies. MATERIAL AND METHODS: Ten pediatric patients who complained of sleep apnea or hypopnea, and were highly suspected of having sleep breathing disorders due to tonsil hypertrophy and underwent surgery were chosen. Pre and post-operative blood somatomedin C and urinary growth hormone levels were measured. In 6 of the 10 patients blood GH levels were measured after overnight collection of blood every 30 min through a catheter placed in the vein. METHOD OF ANALYSIS: Integral blood GH calculated from GH levels of the blood samples taken every 30 min was used to express the efficacy of GH secretion during sleep, and was compared with blood somatomedin C and urinary GH levels. Pre- and post-operative somatomedin C and urinary GH measurements were compared. RESULT: 1) In 6 of the 10 patients, there was a positive correlation with integral GH and urinary GH, and no correlation with integral GH or somatomedin C. 2) Post-operatively, 7 of the 10 patients had increased urinary GH levels, 1 showed no change and 2 had decreased levels. There was a significant increase in the mean urinary GH levels from 18.6 +/- 16.1 pg/ml pre-operatively to 35.8 +/- 19.6 pg/ml post-operatively (P < 0.05). 3) Eight of the 10 patients had a post-operative increase in somatomedin C levels, 1 showed no change and the other patient had a decreased level. There was a significant increase in mean somatomedin C levels from 175.9 +/- 124.8 ng/ml pre-operatively to 226.6 +/- 134.3 ng/ml post-operatively (P < 0.01). CASE: An 8-year-old-boy was diagnosed as having habitual tonsillitis and suspected sleep breathing disorder, and was admitted to our hospital for a tonsillectomy. Apnea and hypopnea were observed during sleep in the hospital. After receiving his parents' consent, GH secretion was evaluated before and after the tonsillectomy. Integrated GH measurements, somatomedin C and urinary GH increased significantly after the operation. CONCLUSION: In this study, the improvement of sleep breathing disorder by surgery was associated with a positive effect to the GH secretion. GH secretion is important for growth of a child. Wen the cause of sleep breathing disorder is evident in the upper airway system, active treatment and cure by surgery is advised. Measurement of urinary GH in the morning is useful and simple for evaluating the ability to secrete GH.

Biomarkers↗