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Biomedical subjects

M Hayashi

Publications and source records attributed to M Hayashi.

At least 505 records · Page 28Linked to original sources
PubMed1999

[Susceptibilities of Enterococcus faecium, PRSP and MRSA to RP59500 and their correlations with those to other drugs].

Investigations on emergence of vancomycin-resistant Enterococcus faecium (VREF) which has recently been attracting attention, especially in the Western countries, have been conducted in Japan. A total of 1,239 isolates of E. faecium were collected from 19 institutions during the period of April 1995 and June 1996, in the purpose of evaluating susceptibilities to variety of antimicrobial agents, including RP59500 and vancomycin (VCM), and detecting vancomycin-resistant genes (van genes). Susceptibilities of penicillin-resistant Streptococcus pneumoniae (PRSP) and methicillin-resistant Staphylococcus aureus (MRSA) were also studied. As a result, 2 isolates of E. faecium were found to be moderately resistant to VCM showing MIC of 8 micrograms/ml though the final identification in species level and the detection of van genes by PCR method have not been completed. On the other hand there detected no MRSA nor PRSP showing moderately resistant or resistant to VCM. It was concluded that RP59500 and VCM possessed favorable activity against clinically isolated E. faecium, PRSP and MRSA. Among other species of enterococci, moderately resistant strains to VCM showing MIC of 8 micrograms/ml were detected; 10 isolates of E. gallinarum, 4 of E. casseliflavus and 2 of E. flavescens. In those isolates, vanC1 and vanC2 were detected by PCR, and vanB was also detected in a isolates of E. gallinarum simultaneously.

Anti-Bacterial Agents↗
PubMed1999

[Coagulation and fibrinolytic activation in lacunar infarct patients].

Lacunar infarcts are related to occlusion of penetrating arteries. Lipohyalinosis affects the smaller arteries 40-200 microns in diameter, and atherosclerosis involves larger arteries 200-850 microns in diameter. We hypothesized that the processes of thrombus formation might be different among these two kinds of lacuner infarcts, including those caused by lipohyalinosis and atherosclerosis. We studied acute coagulation and fibrinolytic activation in lacunar infarct patients which were divided into two groups according to their size: smaller lacunar group and larger lacunar group. Then we divided lacunar infarct patients into two groups in terms of the progression of motor deficits: those who showed the progression and those did not. And coagulation and fibrinolytic activation were compared each other. One hundred and twenty four patients were enrolled in this study, including 34 control subjects, 39 patients with smaller lacune (3 mm-10 mm in diameter), 28 patients with large lacune (10 mm-20 mm), and 23 patients with atherothrombotic infarcts confirmed by angiography. The levels of TAT activity in large lacune and atherothrombotic infarcts were significantly higher than those in control subjects (p = 0.009, p < 0.0001, respectively), whereas those in small lacune were not. Also, the levels of D-dimer activity in large lacune and atherothrombotic infarcts were significantly higher than those in control subjects (p = 0.0003, p < 0.0001, respectively), whereas those in small lacune were not. The progression of motor deficits were more frequently recognized in large lacune than in small lacune: three patients out of 39 small lacune patients and 22 patients out of 28 large lacune patients (difference was significant, p = 0.001). The level of TAT activity in patients who showed progression of motor deficits was significantly higher than that in those who did not (p = 0.0002), whereas the difference of the levels of D-dimer activity in two groups did not reach significant differencial levels. The process of thrombin and fibrin formation in large lacunar infarcts which are related to microatheroma and atheroscrelosis appears to be different from that in small lacunar infarcts. Antiplatelet and anticoagulation therapy should be tailored to large lacunar infarct patients.

Aged↗
PubMed1999

Abnormal FHIT transcripts found in both lung cancer and normal lung tissue.

Occurrence of abnormal transcripts of the FHIT (fragile histidine triad) gene has been reported in various types of cancer. On the other hand, aberrant transcripts are sometimes found in non-neoplastic tissues, so the relationship between the presence of abnormal transcripts of the FHIT gene and cancer pathogenesis is controversial. We investigated alterations in the FHIT locus, detected by nested reverse transcription-polymerase chain reaction and/or allelic status, in 88 primary lung cancers and normal lung tissues, and 22 normal lung tissues with metastatic lung cancer as a control. The frequencies of abnormal transcripts were 59% in lung cancer, 35% in paired normal lung, and 64% in normal control lung; the difference in frequencies between lung cancer and paired normal lung was significant, while that between lung cancer and normal control lung was not. Sequence analysis revealed that there were no cancer-specific abnormal transcripts entirely missing two or more exons, nor were the abnormal transcripts of lung cancer identical with those of paired normal lung in the same individual. Furthermore, we found no correlation between loss of heterozygosity in the FHIT locus and occurrence of abnormal FHIT transcripts. These results suggest that the presence of abnormal FHIT transcripts, in terms of their frequency and variety, is not cancer-specific in lung carcinogenesis, and the abnormality may be mainly due to abnormal splicing and processing of the transcripts. To estimate the precise function of the FHIT gene, further study of the FHIT protein in lung carcinogenesis is needed.

Acid Anhydride Hydrolases↗
PubMed1998

The expression of late infantile neuronal ceroid lipofuscinosis (CLN2) gene product in human brains.

We raised polyclonal antibodies against a gene product responsible for late infantile neuronal ceroid lipofuscinosis (CLN2). By Western blotting, all three antisera recognized the CLN2 protein at approximately 49 kDa in human brain homogenates. Immunohistochemistry using the antisera demonstrated the granular labelling in the cytoplasm of cerebral neurons and glial cells. The immunoreactivity on Western blots was absent from the brain of a patient with CLN2. Our results suggest the usefulness of these antibodies for the diagnosis of CLN2, which currently requires demonstration of characteristic ultrastructure by electron microscopy.

Adolescent↗
PubMed1998

Effects of ACTH on adrenal angiotensin II receptor subtype expression in vivo.

Control of adrenal aldosterone secretion is an important endocrine mechanism mediated by angiotensin II (Ang. II). Recently three subtypes of angiotensin receptors have been demonstrated in the rat adrenal. Our aim was to examine if these receptors are affected by hormone treatment in vivo. Treatment of rats with ACTH resulted in a decrease in AT2 receptor binding from 766 +/- 95 to 310 +/- 51 fmol/mg protein (P < 0.05), without significant changes in AT1 receptors. AT2 receptor mRNA was also decreased (18 +/- 2%, of control, P < 0.05; 25+/-7% of control, P < 0.05) after both 2 and 7 day treatment with ACTH. Changes in AT1a receptor mRNA or total AT1 mRNA did not reach statistical significance. Moreover, treatment with dexamethasone or aldosterone did not affect AT1a, AT1b, or AT2 mRNA. These results demonstrate that ACTH treatment results in subtype-specific changes in adrenal AT receptor gene expression in vivo.

Adrenal Glands↗
PubMed1998

Spindle disturbances induced by benzo[a]pyrene and 7, 12-dimethylbenz[a]anthracene in a Chinese hamster cell line (V79-MZ) and the stability of the numerical chromosome aberrations that follow.

We previously reported that benzo[a]pyrene (BP) and 7, 12-dimethylbenz[a]anthracene (DMBA) induce aneuploidy and polyploidy, respectively, in the Chinese hamster cell line V79-MZ in the absence of S9 mix. In the present study we investigated the effect of BP and DMBA on the mitotic spindle. BP caused incomplete spindle formation and DMBA inhibited spindle formation completely. The combined results indicate that incomplete spindles caused by BP resulted in aneuploidy, and the absence of spindle formation caused by DMBA resulted in polyploidy. The induced polyploidy was stable for several serial passages in fresh medium. BP and DMBA induced different chromosome number distributions. After BP treatment, the normal distribution of chromosome number was restored in 4 days. After DMBA treatment, on the other hand, a tetraploid peak was maintained for 2 months following an initial transient broad distribution of chromosome number after 1 day. The results suggest that different mechanisms were involved in the induction of numerical aberrations by BP and DMBA. Furthermore the induction of numerical aberrations by BP and DMBA was reproducible over 5 months of passages. In four clones tested, the frequency of cells with the modal chromosome number in control cultures gradually decreased from 82% on the average just after cloning to 62% 5 months later. BP and DMBA induced characteristic ploidy changes following succeeding cell passages for up to 5 months, indicating that the ability to respond to BP and DMBA was stable for that period of time. Because these findings were specific to V79-MZ cells, this cell line might be a good tool for studying chemicals that induce numerical chromosome aberrations.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1998

Conformation analysis of eel calcitonin--comparison with the conformation of elcatonin.

The solution structure of eel calcitonin in a mixture of 60% water and 40% trifluoroethanol has been determined in this study by the combined use of 1H-NMR spectroscopy and distance geometry calculations. 1H-NMR spectroscopy provided 181 distance constraints, 5 dihedral angle constraints and 14 hydrogen bond constraints. The observed NOEs demonstrated the presence of an amphiphilic a-helix in position Leu4-Gln20. The seven best converged structures exhibit backbone atomic rmsd of 0.027 nm for the backbone atoms from the averaged coordinate position in the region of Cysl-Leu19. In the previous study [Ogawa, K., Nishimura, S., Doi, M., Kyogoku, Y., Hayashi, M. & Kobayashi, Y. (1994) Eur: J. Biochem. 222, 659-666], the conformation of elcatonin, an analogue of eel calcitonin, was characterized by an amphiphilic a-helix between Thr6 and Thr21 and a turn structure in the first five residues of the N-terminus. The major difference of structure between eel calcitonin and elcatonin exists within the cyclic moiety at the helical N-terminus. Some of the turn structure detected at the N-terminus in elcatonin is not found in eel calcitonin. This is attributed to the difference in the ring formation caused by the disulfide bridge and ethylene bridge. A medium-range NOE, dalphaN(i,i+2), is detected between the CalphaH of Arg24 and the NH of Asp26, so a turn structure occurs in this segment. This NOE connectivity profile in the C-terminal region is also the same in elcatonin, suggesting that this is important for receptor binding and immunological properties.

Amino Acid Sequence↗
PubMed1998

Evaluation of a tissue homogenization technique that isolates nuclei for the in vivo single cell gel electrophoresis (comet) assay: a collaborative study by five laboratories.

We evaluated a tissue homogenization technique that isolates nuclei for use in the in vivo comet assay. Five laboratories independently tested the technique using the liver, kidney, lung, spleen, and bone marrow of untreated and mutagen-treated male CD-1 mice. The direct mutagen methylmethanesulfonate (MMS) or the promutagen diethylnitrosamine (DEN) were injected intraperitoneally at maximum tolerated doses. Three and twenty-four hours later, the organs were removed and, except for bone marrow, were minced and homogenized and a nuclear suspension was prepared. The nuclear suspensions and bone marrow cells were used in the comet assay. None of the nuclear suspensions from the non-treated mice induced a positive response. All nuclear suspensions derived from the MMS-treated mice and those of the liver, kidney, and lung from DEN-treated mice induced positive responses in all the laboratories similarly. Reproducibility was demonstrated by five replicate studies in one laboratory. Furthermore, the organ-specific responses to MMS and DEN reflected the characteristic genotoxicity of the chemicals. We concluded from these results that the homogenization technique is a valid one to be used for mouse organs in the in vivo comet assay.

Animals↗
PubMed1998

Reversal of multidrug resistance (MDR) by aspidofractinine-type indole alkaloids.

A series of indole alkaloids of the aspidofractinine-type was assessed for their potential in reversing MDR in vincristine-resistant KB cells. Of the compounds tested, kopsiflorine, kopsamine, pleiocarpine, 11-methoxykopsilongine, lahadinine A and N-methoxycarbonyl-11,12-methylenedioxy-delta 16,17-kopsinine were found to show appreciable activity.

Alkaloids↗
PubMed1998

Molecular cloning and characterization of human PDE8B, a novel thyroid-specific isozyme of 3',5'-cyclic nucleotide phosphodiesterase.

We have identified a novel human isozyme of 3',5'-cyclic nucleotide phosphodiesterase (PDE), which we designated PDE8B. cDNA of 2844 bp encoding the C-terminal 659 amino acids of PDE8B was cloned following the identification of an expressed sequence tag (EST) obtained through a search of the EST database. The predicted protein sequences of PDE8B showed highest homology (65% identity, 83% similarity) to that of PDE8A. Northern blot analysis indicated that the mRNA encoding PDE8B is expressed specifically and abundantly in thyroid gland as a approximately 4.2 kb mRNA, in contrast to the wide expression of PDE8A mRNA in various tissues. The carboxyl-terminal 584 amino acids of PDE8B were expressed in E.coli as a fusion protein. The recombinant PDE8B exhibited cAMP PDE activity which was not inhibited by various PDE inhibitors including vinpocetine, milrinone, rolipram, and IBMX with the exception of dipyridamole which caused 50% inhibition at a concentration of 40 microM. cAMP hydrolytic activity was unaffected by cGMP and no cGMP PDE hydrolysis were detectable at concentrations up to 100 microM. These findings suggest that PDE8B is a new member of the PDE8 family.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1998

Adrenal angiotensin II type 1 and type 2 receptors in Cushing's and Conn's syndromes.

Cushing's and Conn's syndromes are well recognised endocrine diseases, but the pathobiology of the tumors causing these disorders is unclear. In this study we examined AT1 and AT2 gene expression in adrenal adenomas of Cushing's and Conn's syndromes. AT1 and AT2 receptor mRNA, as well as alternatively spliced AT1 transcripts, were detected by RT-PCR using adjacent adrenal cortex tissue as controls. Whereas no consistent differences in AT1 mRNA were seen compared to control adrenal cortex, AT2 mRNA levels were significantly decreased in the adenomas of Cushing's and Conn's syndromes. No changes in alternative splicing of AT1 mRNA were observed in the adrenal tumors. The fact that no consistent changes were seen in AT1 mRNA or its splicing, whereas AT2 mRNA were reduced in both forms of hormone producing adrenal tumor suggests that the AT2 receptor, rather than the AT1 subtype, may be correlated with adrenal tumorigenesis.

Adrenal Cortex↗
PubMed1998

Recommendations for statistical designs of in vivo mutagenicity tests with regard to subsequent statistical analysis.

A workshop was held on September 13 and 14, 1993, at the GSF, Neuherberg, Germany, to start a discussion of experimental design and statistical analysis issues for three in vivo mutagenicity test systems, the micronucleus test in mouse bone marrow/peripheral blood, the chromosomal aberration tests in mouse bone marrow/differentiating spermatogonia, and the mouse dominant lethal test. The discussion has now come to conclusions which we would like to make generally known. Rather than dwell upon specific statistical tests which could be used for data analysis, serious consideration was given to test design. However, the test design, its power of detecting a given increase of adverse effects and the test statistics are interrelated. Detailed analyses of historical negative control data led to important recommendations for each test system. Concerning the statistical sensitivity parameters, a type I error of 0.05 (one tailed), a type II error of 0.20 and a dose related increase of twice the background (negative control) frequencies were generally adopted. It was recommended that sufficient observations (cells, implants) be planned for each analysis unit (animal) so that at least one adverse outcome (micronucleus, aberrant cell, dead implant) would likely be observed. The treated animal was the smallest unit of analysis allowed. On the basis of these general consideration the sample size was determined for each of the three assays. A minimum of 2000 immature erythrocytes/animal should be scored for micronuclei from each of at least 4 animals in each comparison group in the micronucleus assays. A minimum of 200 cells should be scored for chromosomal aberrations from each of at least 5 animals in each comparison group in the aberration assays. In the dominant lethal test, a minimum of 400 implants (40-50 pregnant females) are required per dose group for each mating period. The analysis unit for the dominant lethal test would be the treated male unless the background frequency of dead implants (DI) is so low that multiple males would need to be integrated to meet the minimum observation of one adverse outcome (DI) per analysis unit. A three-step strategy of data analysis was proposed for the cytogenetic assays. Use of negative historical controls was allowed in certain circumstances for interpretation of results from micronucleus tests and chromosomal aberration tests.

Animals↗
PubMed1998

An automated new technique for scoring the rodent micronucleus assay: computerized image analysis of acridine orange supravitally stained peripheral blood cells.

We developed an automated image analysis system to obtain objective data for the rodent peripheral blood micronucleus assay with acridine orange (AO) supravital staining. The system was able to identify micronucleated reticulocytes (MNRETs) and to evaluate inhibition of bone marrow cell proliferation by measuring the reticular area of reticulocytes (RETs). We also developed automated equipment to produce homogeneous acridine orange-coated glass slides. This study was designed to compare automated scoring with manual scoring using 4 model clastogens and 2 mouse strains. The MNRET incidence induced by each clastogen was similar for automated and manual scoring, and there was good correlation (r=0.92) between the methods. In addition, an index of bone marrow toxicity based on the reticular area of RETs was compared to the conventional index (% of polychromatic erythrocytes (PCE) to total erythrocytes; PCE ratio) and was similar. The results indicated that our technique for computer-assisted image analysis for the micronucleus assay with AO supravitally stained peripheral blood RETs was comparable to conventional microscopic scoring, and it was superior in objectivity and statistical power.

Acridine Orange↗
PubMed1998

Development of GAP-43 mRNA in the macaque cerebral cortex.

To estimate the extent of axonal growth in various areas of the cerebral cortex, we measured the amount of GAP-43 mRNA in the cerebral cortex of developing macaque monkeys. In four areas, i.e., the prefrontal area (FD delta), the temporal association area (TE), the primary somatosensory area (PC), and the primary visual area (OC), the amount of GAP-43 mRNA was measured from the intermediate fetal period [embryonic day 120 (E120)] to the adult stage. In two other areas, i.e., the parietal association area (PG) and the secondary visual area (OB), the amount of GAP-43 mRNA was measured during the postnatal period. The amount of GAP-43 mRNA was highest at E120, decreased roughly exponentially, and approached the asymptote by postnatal day 70 (P70). The amount of GAP-43 mRNA was higher in the association areas (FD delta, TE, and PG) than in the primary sensory areas (PC and OC) during development and at the adult stage. These findings suggest that axonal growth in the cerebral cortex is most exuberant before or during the intermediate fetal period and approximately ends by P70. Furthermore, axonal growth is evidently more intensive in the association areas than in the primary sensory areas during the stage following the intermediate fetal period.

Aging↗
PubMed1998

D-aspartate modulates melatonin synthesis in rat pinealocytes.

It has been known that pinealocytes contain the highest level of D-aspartate among various neuroendocrine cells in the rat. Here, we report that exogenous D-aspartate strongly inhibited norepinephrine-dependent melatonin synthesis in the rat pineal gland, the concentration required for 50% inhibition being 75 microM. This inhibition was due at least partly to decreased norepinephrine-dependent serotonin N-acetyltransferase activity. Upon incubation, D-aspartate was gradually released from pinealocytes and accumulated in the incubation medium as determined by high-performance liquid chromatography on a Pirkle-type chiral column. These results suggest that D-aspartate acts as a negative regulator for melatonin synthesis in the pineal gland.

Animals↗
PubMed1998

Identification of D-aspartate in rat pheochromocytoma PC12 cells.

D-Aspartate is now known to be present in mammalian neuronal and endocrine cells in vivo, and may play some role(s) in neurocrine and endocrine functions. However, origin of D-aspartate is unknown. Here, we report that free D-aspartate (108 pmoles/3 x 10(7) cells) is present in the cultured PC12 cells, a rat pheochromocytoma cell line, as determined with immunohistochemical techniques as well as high performance liquid chromatography (HPLC) on a Pirkle-type chiral column. The amount of D-aspartate does not change with the passage. The culture medium does not contain D-aspartate. These results strongly suggest the presence of a de novo biosynthetic pathway for D-aspartate in the endocrine cells.

Animals↗