Search PubMed⌕ Search

Biomedical subjects

S Ohnishi

Publications and source records attributed to S Ohnishi.

At least 73 records · Page 4Linked to original sources
PubMed1998

Hybridization-AT-tailing (HybrAT) method for sensitive and strand-specific detection of DNA and RNA.

delta Tth DNA polymerase catalyzed polymerization of dATP and dTTP into a high-molecular-weight d(A-T) copolymer using oligo-d(A-T) as the template/primer (Hanaki et al., Biochem. Biophys. Res. Commun. 244, 210-219). Taking advantage of this reaction, we developed a highly sensitive method for strand-specific detection of DNA or RNA. The probe consisted of a 40- to 50-base-long complementary sequence on the 5' side and 10 repeats of AT on the 3' side. After hybridization using the 5' side, the 3' side AT repeat region was elongated by delta Tth DNA polymerase in the presence of the dATP, dTTP, and digoxigenin (dig)-11-dUTP. The elongation condition was 52-62 degrees C for 3 h. The method named HybrAT (hybridization-AT-tailing) was at least 100-fold more sensitive than the conventional hybridization with 5' end dig-11-dUTP labeled probe.

Adenine↗
PubMed1998

Cloning, functional expression, and chromosomal localization of the human and mouse gp180-carboxypeptidase D-like enzyme.

We previously reported that a host cell glycoprotein, gp180, binds duck hepatitis B virus particles, and is encoded by a member of the carboxypeptidase gene family (Kuroki, K., Eng, F., Ishikawa, T., Turck, C., Harada, F., Ganem, D., 1995. gp180, a host cell glycoprotein that binds duck hepatitis B virus particles, is encoded by a member of the carboxypeptidase gene family. J. Biol. Chem. 270, 15022-15028). After that report, carboxypeptidase D (CPD) was subsequently purified from bovine pituitary and characterized as a novel carboxypeptidase E (CPE)-like enzyme, with many characteristics in common with duck gp180 (Song, L., Fricker, L.D., 1995. Purification and characterization of carboxypeptidase D, a novel carboxypeptidase E-like enzyme, from bovine pituitary. J. Biol. Chem. 270, 25007-25013). CPD is now supposed to play an important role in a secretory pathway. To clarify the function of gp180 further, we have isolated and analyzed human and mouse homologues of duck gp180. cDNA clones derived from human HepG2 cells and mouse livers have been isolated on the basis of homology to the duck gp180. The suggested open reading frames of the human and mouse cDNA encode 1380 and 1377 amino acid proteins, respectively and have three carboxypeptidase homologous domains (A, B, and C). Domains A and B have completely conserved the residues known to have the enzymatic activity of carboxypeptidase, but domain C in each cDNA does not. Northern blotting revealed a ubiquitous tissue distribution of human gp180 mRNA with several transcript species. Expression of human gp180 cDNA in transfected 293T cells exhibited carboxypeptidase activity upon radiometric assay. The human and mouse homologues of duck gp180 have many characteristics in common with bovine CPD. Fluorescence in-situ hybridization reveals that the gene encoding human gp180 is located in region 17q11.2.

Amino Acid Sequence↗
PubMed1998

Effects of protein kinase and phosphatase inhibitors on slow shortening of guinea pig cochlear outer hair cells.

The intracellular mechanisms of slow shortening in isolated guinea pig cochlear outer hair cells were investigated using inhibitors and/or an activator of protein kinases and protein phosphatases. The slow shortening was induced by tetanic electrical field stimulation, and changes in the cell length, volume and intracellular Cl- concentration were microscopically monitored using a chloride-sensitive fluorescent dye. The slow shortening was inhibited by a calmodulin inhibitor, W-7, and a calcium calmodulin-dependent protein kinase II (CaMKII) inhibitor, KN-62. The inhibition by W-7 or KN-62, was abolished by the supplemented conductance of K+ with valinomycin. Among the protein phosphatase inhibitors tested, a type 1 and 2A protein phosphatase inhibitor, calyculin A, inhibited the slow shortening. The inhibition by calyculin A was abolished by the increased Cl- permeability, but neither by the increased K+ conductance with valinomycin nor by the increased Ca2+ conductance with A23187. A protein serine/threonine phosphatase activator, N-acetylsphingosine, inhibited the shortening, which was abolished by either valinomycin or a type 2A protein phosphatase inhibitor, okadaic acid, but not by calyculin A. These findings suggest the following signaling mechanisms in the slow shortening of outer hair cells; the K+ channel opening is facilitated through protein phosphorylation by CaMKII and suppressed via okadaic acid-sensitive dephosphorylation, and the Cl- channel opening depends on calyculin A-sensitive protein phosphatase activity.

Animals↗
PubMed1998

Correlation between surface morphology and surface forces of protein A adsorbed on mica.

We have investigated the morphology and surface forces of protein A adsorbed on mica surface in the protein solutions of various concentrations. The force-distance curves, measured with a surface force apparatus (SFA), were interpreted in terms of two different regimens: a "large-distance" regimen in which an electrostatic double-layer force dominates, and an "adsorbed layer" regimen in which a force of steric origin dominates. To further clarify the forces of steric origin, the surface morphology of the adsorbed protein layer was investigated with an atomic force microscope (AFM) because the steric repulsive forces are strongly affected by the adsorption mode of protein A molecules on mica. At lower protein concentrations (2 ppm, 10 ppm), protein A molecules were adsorbed "side-on" parallel to the mica surfaces, forming a monolayer of approximately 2.5 nm. AFM images at higher concentrations (30 ppm, 100 ppm) showed protruding structures over the monolayer, which revealed that the adsorbed protein A molecules had one end oriented into the solution, with the remainder of each molecule adsorbed side-on to the mica surface. These extending ends of protein A overlapped each other and formed a "quasi-double layer" over the mica surface. These AFM images proved the existence of a monolayer of protein A molecules at low concentrations and a "quasi-double layer" with occasional protrusions at high concentrations, which were consistent with the adsorption mode observed in the force-distance curves.

Adsorption↗
PubMed1998

Possible mechanism of ECG features in patients with idiopathic ventricular fibrillation studied by heart model and computer simulation.

The possible contribution of localized conduction delay and abnormal action potentials to ventricular fibrillation (VF) was studied by applying an anisotropic cardiac computer model to clinical cases of the Brugada-type electrocardiogram (ECG), which shows right bundle branch block (RBBB), a normal QT interval, ST-segment elevation, and late r' in leads V1 and V2. The anisotropic heart model was composed of 50,000 discrete units with a spatial resolution of 1.5 mm and was mounted in a human torso model. The longitudinal/transverse conduction velocity ratio was 3:1. For the normal ECG, a conduction velocity of 0.75 m/s was required. In the abnormal area of the right anterior epicardial wall, the conduction velocity was set at 0.2 m/s, with decreasing action potential amplitude and 10% prolonged action potential duration. The ECG features of ST-segment elevation and Brugada-type right bundle branch block pattern were simulated. The action potential duration was able to change dynamically with coupling interval of stimulation, with a ratio of 9% for normal ventricular muscle and 50% for Purkinje fibers. Five successive stimuli were applied to the left lateral epicardium 300 ms after the first sinus excitation, and sustained VF was induced with the transmural conduction delay at the right anterior ventricle as a block increasing the vulnerability. At the initiation of VF, reentry circuits were shown around the border zone of the right epicardium and were very heterogeneous around the conduction delayed area and septal area. In an area with the characteristics of nontransmural conduction delay, sustained VF was prevented, and the pattern of transient right bundle branch block appeared on the simulated ECG and body surface potential maps. The late r' wave was calculated in the precordial leads and right anterior site on the body surface potential maps. These results suggest that increased multipolarity in the border zone between the Purkinje fibers and delayed conduction area in the right ventricle might play an important role as a functional block for the persistence of VF.

Action Potentials↗
PubMed1998

Amiodarone-induced thyrotoxicosis associated with thyrotropin receptor antibody.

Since amiodarone was first marketed in 1992 in Japan, the incidence of amiodarone-induced thyrotoxicosis (AIT) has been increasing. Among 12 thyrotoxic patients, a patient with arrhythmogenic right ventricular dysplasia, who had been taking amiodarone for 4 years, developed thyrotoxicosis with subacute onset, accompanied by transiently positive thyrotropin (TSH) receptor antibody (TRAb), or thyrotropin-binding inhibiting immunoglobulin (TBII). The immunoglobulin G (IgG) obtained from the TRAb-positive serum of the patient elicited no thyroid hormone-releasing activity in cultured human thyroid follicles, whereas all IgGs obtained from untreated Graves' disease elicited positive results. These in vitro findings and clinical course suggest that TRAb/TBII without thyroid-stimulating activity may develop in patients with amiodarone-induced destructive thyroiditis, as reported in patients with destructive thyroiditis, such as subacute and silent thyroiditis.

Adult↗
PubMed1998

[The effect of MS-551 on two VT/Vf patients in CABG: a case report].

Uncontrollable arrhythmia is one of the causes of operative death in CABG. We report two cases of successful treatment of recurrent VT/Vf in CABG operation by MS-551. One case was a 72-year-old male, who had unstable angina and OMI with left ventricular dysfunction (LVEF 24%). After weaning from CPB, the rhythm turned into VT/Vf suddenly. Lidocaine and verapamil were not effective. VT/Vf recurred over the over. The another case was a 52-year-old male, who had OMI and effort angina. He became myocardial infarction during operation. VT/Vf occurred after the weaning from CPB. Lidocaine and amiodarone were of no effect on his VT/Vf. In both cases, intravenous injection of MS-551 promptly stopped VT/Vf. Use of MS-551 was limited mainly on DCM patients. But other antiarrhythmic agents were ineffective on these cases. We have had to continue support circulation for a long time without MS-551. As MS-551 is being used as a trial on medically followed patients, it was unknown how effective clinically on the acute myocardial infarction. In this regard, our cases may suggest a positive answer to that.

Aged↗
PubMed1998

[Study on abuse and neglect of the disabled elderly living at home].

To obtain basic information on elderly abuse and neglect in order to provide for its early discovery and prevention, a questionnaire survey about the present situation regarding abuse and neglect of the disabled elderly living at home was performed on health/medical service/welfare professions. Forty-two cases were analyzed. The major results were as follows; 1. Victims were 13 men and 29 women, approximately 1/3 of whom were in their 70's and another 1/3 in their 80's. Approximately 1/4 of the abusers were either sons or daughters of the victims. 2. Verbal abuse was the most frequent type of abuse with a rate of 69.0%. The percentage of psychological abuse was 61.9, passive neglect was 57.1%, active neglect was 50.0%, physical abuse was 47.6%, passive self-neglect was 28.6%, financial/material exploitation was 23.8%, active self-neglect was 16.7% and the others was 11.9%. The average number of abuse and neglect that an elderly received was 3.5. 3. The main causes of abuse and neglect did not appear to be simple but was complicated by related causes, many of both victims and abusers had elements in their personality, developmental history and interpersonal relationships that over a period of years formed the basis for the problematic behavior. Many abuse or neglect cases arose from caregiving burden of family caregivers or insufficient social systems for support to meet the needs of caring for the elderly. 4. The results suggest that for prevention and countermeasures for elderly abuse, there is an urgent need to arrange for expansion of health and welfare services to reduce burden of caregivers, provide for education of professionals of health/medical service/welfare for early discovery and proper handling of abuse problems, development of a checklist for early discovery, expansion of opportunities of improving care skill for family caregivers and establishment of consultation system for the elderly and caregivers, and organizing emergency shelter for victims of elderly abuse.

Aged↗
PubMed1998

[Treatment of aplastic anemia with anti-thymocyte globulin and cyclosporine].

Antithymocyte globulin (ATG) is an effective immunosuppressive therapy for aplastic anemia (AA). We administered ATG combined with cyclosporine (CyA), to 9 patients (4 men and 5 women; median age, 55 years). AA was severe in 8 patients and moderate in 1. The ATG and CyA regimen was the initial treatment for 3 patients, but sequential treatment for the other 6, who were refractory to other agents. Peripheral T lymphocytes, including CD4-positive and CD8-positive cells, decreased rapidly after treatment. Although 1 patient died of pulmonary hemorrhage during the 6-month period following treatment with this combined regimen, 3 showed a favorable response, 2 moderate response, and 3 no response at all. Adverse drug reactions, including transient fever and rash, were not severe. These findings suggested that ATG and CyA in combination are a safe and effective immunosuppressive regimen for initial and refractory patients with AA.

Adult↗
PubMed1997

Structural features of membrane fusion between influenza virus and liposome as revealed by quick-freezing electron microscopy.

The structure of membrane fusion intermediates between the A/PR/8(H1N1) strain of influenza virus and a liposome composed of egg phosphatidylcholine, cholesterol, and glycophorin was studied using quick-freezing electron microscopy. Fusion by viral hemagglutinin protein was induced at pH 5.0 and 23 degrees C. After a 19-s incubation under these conditions, small protrusions with a diameter of 10-20 nm were found on the fractured convex faces of the liposomal membranes, and small pits complementary to the protrusions were found on the concave faces. The protrusions and pits corresponded to fractured parts of outward bendings of the lipid bilayer or "microprotrusions of the lipid bilayer." At the loci of the protrusions and pits, liposomal membranes had local contacts with viral membranes. In many cases both the protrusions and the pits were aligned in regular polygonal arrangements, which were thought to reflect the array of hemagglutinin spikes on the viral surface. These structures were induced only when the medium was acidic with the virus present. Based on these observations, it was concluded that the microprotrusions of the lipid bilayer are induced by hemagglutinin protein. Furthermore, morphological evidence for the formation of the "initial fusion pore" at the microprotrusion was obtained. The protrusion on the convex face sometimes had a tiny hole with a diameter of <4 nm in the center. The pits transformed into narrow membrane connections <10 nm in width, bridging viruses and liposomes. The structures of the fusion pore and fusion neck with larger sizes were also observed, indicating growth of the protrusions and pits to distinct fusion sites. We propose that the microprotrusion of the lipid bilayer is a fusion intermediate induced by hemagglutinin protein, and suggest that the extraordinarily high curvature of this membrane structure is a clue to the onset of fusion. The possible architecture of the fusion intermediate is discussed with regard to the localization of intramembrane particles at the microprotrusion.

Acids↗
PubMed1997

Idiopathic ventricular fibrillation induced with vagal activity in patients without obvious heart disease.

BACKGROUND: Recently, idiopathic ventricular fibrillation (VF) has gained much attention. Although several subgroups have been described, its pathogenesis, mechanism, treatment, and prognosis remain unknown. METHODS AND RESULTS: We studied six cases of idiopathic VF with transient late r' waves and ST elevation (late r'/ST elevation) in leads V1 through V3. Late r'/ST elevation was augmented before and after VF episodes. Signal-averaged ECGs showed late potentials even when no late r'/ST elevation occurred. During late r', a conduction delay was observed by use of body-surface maps at the anterior wall and outflow tract of the right ventricle without inhomogeneity of the repolarization phase. There was a decrease or total disappearance of late r'/ST elevation with isoproterenol, atropine, and exercise stress testing and induction or exacerbation with propranolol, edrophonium, and hyperventilation. VF was induced by programmed electrical stimulation in all cases but two, in which it was induced only after edrophonium injection. In two cases, VF was exacerbated by propranolol, and in all cases, it was uninducible with isoproterenol. Heart rate spectral analysis just before VF episodes showed a sudden rise in vagal activity in two cases. As the VF mechanism, a conduction delay exists at the anterior wall and outflow tract of the right ventricle that is possibly exacerbated by an abrupt rise in vagal activity, inducing random reentry that results in VF. Class I antiarrhythmic agents and beta-blockers were ineffective for this VF. All subjects required implantable cardioverter-defibrillators. CONCLUSIONS: We propose this VF associated with late r'/ST elevation in the precordial leads and influenced by vagal activity as a new possible mechanism of idiopathic VF.

Adult↗
PubMed1997

Cytotoxic T lymphocyte response and viral load in hepatitis C virus infection.

A cytotoxic T lymphocyte (CTL) response to the hepatitis C virus (HCV) nucleoprotein residues 88-96 that are the minimal and optimal epitope for human leukocyte antigen (HLA) B44-restricted CTLs was assessed in 27 HLA B44-positive patients with chronic HCV infection. Serum HCV RNA concentration and the amino acid sequence of the residues 81-100 were also determined. Three patients were infected with HCV with uncommon amino acid substitutions within the epitope. One was infected with HCV with an amino acid substitution in the flanking residues of the epitope. To stimulate CTLs in the peripheral blood, 9-mer peptides that corresponded to the residues 88-96 of the individual patients were synthesized and used. Seven of the 27 patients demonstrated a CTL response to the residues 88-96 with specific cytotoxic activities higher than 20%. The CTL activities were significantly higher in patients with a low titer of serum HCV RNA than in those with a high titer of serum HCV RNA (P = .0006). Some of the patients that demonstrated a CTL response to the residues 88-96 also demonstrated a CTL response to a newly identified HLA B44-restricted CTL epitope or a known HLA A11-restricted CTL epitope or both. No apparent association was observed between the CTL response and the stage of disease, or between the CTL response and the grade of necroinflammatory activity. The results suggest that the HLA B44-restricted CTLs together with other HCV-specific CTLs may inhibit the outgrowth of HCV and that high-titer infection with HCV may suppress the CTL responses.

Adult↗
PubMed1997

Clinical significance of thallium-201 and gallium-67 scintigraphy in pulmonary tuberculosis.

One hundred and thirty-nine patients with pulmonary tuberculosis were evaluated by means of gallium-67 and thallium-201 scintigraphy. The disease was clinically active in 83 and inactive in 56. The uptake ratio between the lesion and the contralateral normal lung field was calculated. The ratio determined by 67Ga scintigraphy was expressed as GR, and that determined by 201T1 scintigraphy (early or delayed) as ER or DR. The 201T1 retention index (RI) was calculated using the following equation: RI = DR - ER/ER x 100. The sensitivity, specificity and accuracy of 201T1 scintigraphy with respect to the activity of pulmonary tuberculosis were better than those of 67Ga scintigraphy (the figures for 201T1 scintigraphy were 88.0%, 82.1% and 85.6%, respectively, and those for 67Ga scintigraphy, 83.1%, 60.7% and 74.1%). We found a significant correlation between GR and ER, but there was no significant correlation between RI and ER. When the relationships between ER, C-reactive protein and 1-h erythrocyte sedimentation rate were examined among patients with abnormal uptake, no significant relationships were noted. In the 27 patients who could be followed up, GR and ER decreased with duration of the therapy, indicating a decrease in disease activity which was consistent with clinical findings. The RI was significantly higher in the early stages of therapy than in the later stages (P < 0.01), suggesting a temporary delay in 201T1 washout in the early stages. The washout seemed to be promoted by the effects of the therapy. Overall, it is concluded that 201T1 scintigraphy is more useful and more suitable than 67Ga scintigraphy for the evaluation of disease activity and therapeutic effects in patients with pulmonary tuberculosis.

Blood Sedimentation↗
PubMed1997

Mitochondrial DNA and nuclear DNA indicate that the Japanese Fasciola species is F. gigantica.

For elucidation of the taxonomic status of the Japanese Fasciola species, whole mitochondrial DNA of Fasciola hepatica from Australia, F. gigantica from Malaysia, and Fasciola sp. from Japan was digested with three four-base-cutting endonucleases: HinfI, MspI, and RsaI. The resulting digestion patterns showed that for each enzyme there were some bands specific for each geographical isolate and that the Japanese Fasciola sp. shared more bands with F. gigantica than with F. hepatica. Nucleotide sequences of two regions, the second internal transcribed spacer (ITS2) of the nuclear ribosomal RNA cluster and mitochondrial cytochrome c oxidase subunit I (COI), were also compared among them. The ITS2 sequence was highly conserved among the three isolates. F. gigantica and the Japanese Fasciola sp. were identical, but they differed from the Australian F. hepatica at six sites, one of which was a deletion. The COI sequence was less conserved but implied a similar relationship between the isolates. There seems no reason to regard the Japanese Fasciola sp. as anything other than a strain of F. gigantica.

Animals↗
PubMed1997

Cardiac memory in patients with intermittent Wolff-Parkinson-White syndrome.

This study used body surface mapping to evaluate the ventricular repolarization process in the absence of delta waves in 13 patients with the intermittent Wolff-Parkinson-White (WPW) syndrome. The findings were compared with data from 30 normal individuals and 50 patients with the overt WPW syndrome. The QRST isointegral maps of patients with the overt WPW syndrome exhibited abnormal areas and the QRST departure maps showed a peculiar distribution to each accessory pathway site. The QRST isointegral map exhibited abnormal areas in 11 of the 13 cases (85%) of the intermittent WPW syndrome in the absence of delta waves. In 8 of these 11 cases (73%), the distribution of the departure map resembled that in the overt WPW syndrome. These findings suggest that abnormal ventricular repolarization due to cardiac memory is present in patients with the intermittent WPW syndrome even in the absence of delta waves.

Adolescent↗
PubMed1997

Increased expression of vascular endothelial growth factor in human hepatocellular carcinoma.

BACKGROUND/AIMS: Angiogenesis is critical for the development and progression of solid tumors. The purpose of this study was to evaluate the possible role of vascular endothelial growth factor (which is considered to be one of the most important factors involved in tumor-associated angiogenesis), in human hepatocellular carcinoma. METHODS: Vascular endothelial growth factor gene and protein expression were analyzed by means of Northern hybridization and immunohistochemical methods in 5 hepatocellular carcinoma cell lines. Secretion of vascular endothelial growth factor was evaluated by immunoblotting of conditioned medium of these hepatocellular carcinoma cells. Further, we compared the level of vascular endothelial growth factor expression in hepatocellular carcinoma tissues along with that in surrounding tumor-free tissues obtained from 20 patients. We also analyzed mRNA expression of Flt-1, one of the vascular endothelial growth factor specific high-affinity receptors, in hepatocellular carcinoma cell lines. RESULTS: Northern hybridization analysis and immunohistochemistry revealed that all cultured hepatocellular carcinoma cells exhibited a high level of vascular endothelial growth factor mRNA. In addition, vascular endothelial growth factor secretion by Hep G2, one of the hepatocellular carcinoma cell lines, was shown by Western blot. In vivo, we observed vascular endothelial growth factor expression in both hepatocellular carcinoma and non-hepatocellular carcinoma tissues. However, in 12 of 20 cases, vascular endothelial growth factor mRNA levels were significantly up-regulated in hepatocellular carcinoma tissues. In the majority of cases (10 out of 12 cases) with abundant tumor vascularity, vascular endothelial growth factor mRNA up-regulation in hepatocellular carcinoma tissues was observed. We failed to detect Flt-1 mRNA expression in hepatocellular carcinoma cells. CONCLUSIONS: This study suggests that the possibility that hepatocellular carcinoma cells overexpress the vascular endothelial growth factor gene and protein. These findings support the hypothesis that vascular endothelial growth factor is one of the important factors involved in the angiogenesis of hepatocellular carcinoma, and may even be involved in the development and/or progression of hepatocellular carcinoma itself.

Adolescent↗
PubMed1997

Influence of early coronary reperfusion on QT interval dispersion after acute myocardial infarction.

We studied the influence of early coronary reperfusion on QT interval dispersion in patients with acute myocardial infarction (MI). There were 54 males and 18 females with a mean age of 60 +/- 10 years. Of the 51 patients with recanalization of the infarct related vessel in the recovery phase, 28 (group A) had early coronary reperfusion (5.5 +/- 2.7 hours), 23 other patients (group B) were not confirmed with early coronary reperfusion. Twenty-one patients (group C) did not undergo recanalization of the infarct related vessel in the recovery phase. Corrected QT (QTc) maximum, QTc minimum, and QTc dispersion calculated as the difference between the maximum and minimum QTc intervals, were compared among these three groups at both acute and recovery phase. At the acute phase after MI, there were no significant differences in the QTc maximum, QTc minimum, QT dispersion, and QTc dispersion among these three groups. At the recovery phase after MI, there were also no significant differences in the QTc maximum and QTc minimum. However, there were significant differences in the QT dispersion (0.035 +/- 0.010 in group A, 0.049 +/- 0.015 in group B, and 0.061 +/- 0.031 s in group C, respectively; P = 0.0001), and QTc dispersion (0.038 +/- 0.012 in group A, 0.050 +/- 0.015 in group B, and 0.063 +/- 0.032 s in group C, respectively; P = 0.0003) among the three groups. Comparison of QTc dispersion between acute and recovery phase revealed significant reduction from acute to recovery phase in group A. The number of premature ventricular contraction was lower in group A and B than group C. In summary, early coronary reperfusion may reduce electrophysiological instability by reducing QT dispersion in the recovery phase after acute MI.

Cardiac Catheterization↗