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Biomedical subjects

S Yoshitake

Publications and source records attributed to S Yoshitake.

At least 19 recordsLinked to original sources
PubMed1992

Thioredoxin regenerates proteins inactivated by oxidative stress in endothelial cells.

The thioredoxin/thioredoxin reductase system has been studied as regenerative machinery for proteins inactivated by oxidative stress in vitro and in cultured endothelial cells. Mammalian glyceraldehyde-3-phosphate dehydrogenase was used as the main model enzyme for monitoring the oxidative damage and the regeneration. Thioredoxin and its reductase purified from bovine liver were used as the regenerating system. The physiological concentrations (2-14 microM) of reduced thioredoxin, with 0.125 microM thioredoxin reductase and 0.25 mM NADPH, regenerated H2O2-inactivated glyceraldehyde-3-phosphate dehydrogenase and other mammalian enzymes almost completely within 20 min at 37 degrees C. Although the treatment of endothelial cells with 0.2-12 mM H2O2 for 5 min resulted in a marked decrease in the activity of glyceraldehyde-3-phosphate dehydrogenase, it had no effect on the activities of thioredoxin and thioredoxin reductase. Essentially all of the thioredoxin in endothelial cells at control state was in the reduced form and 70-85% remained in the reduced form even after the H2O2 treatment. The inactivated glyceraldehyde-3-phosphate dehydrogenase in a cell lysate prepared from the H2O2-treated endothelial cells was regenerated by incubating the lysate with 3 mM NADPH at 37 degrees C and the antiserum raised against bovine liver thioredoxin inhibited the regeneration. The inhibition of thioredoxin reductase activity by 13-cis-retinoic acid resulted in a decrease in the regeneration of glyceraldehyde-3-phosphate dehydrogenase in the H2O2-treated endothelial cells. The present findings provide evidence that thioredoxin is involved in the regeneration of proteins inactivated by oxidative stress in endothelial cells.

Animals↗
PubMed1992

Immunogenic properties of structurally modified human tissue plasminogen activators in chimpanzees and mice.

Immunogenic properties of second generation human tissue plasminogen activator (tPA) derivatives were examined in chimpanzee and mouse systems. Five species of modified tPAs (mtPAs) (designated 2660, 2663, 2810, 8000, and 9200), recombinant native tPA or bovine serum albumin (BSA) as a positive control were subcutaneously injected nine times at suitable intervals into chimpanzees, genetically the closest species to man. These animals were tested for antigen(Ag)-specific antibodies to the corresponding proteins by means of enzyme-linked immunosorbent assay and Western blot analysis. Neither 9200, one of the five mtPAs tested, nor tPA was immunogenic, although BSA and the other four mtPAs were immunogenic under these conditions. Thus, an antigenic determinant was not exposed by the modification on 9200 and this modified tPA is expected not to be immunogenic in humans. In the mouse studies, mice were immunized with mtPAs. Serum samples from these animals were tested for antibodies to the mtPAs which did not concomitantly recognize native tPA by immune adsorption of the antibodies to tPA. The amount of such antibodies after the elimination of native tPA-reactive antibodies was little or none when the serum samples from 9200 and from the other mtPAs, except 8000, were tested. Taking into consideration the results of the chimpanzee studies, it can be concluded that Ag-specific antibodies are dominantly produced to unchanged epitopes present in modified proteins in the mouse system, in which the native protein is immunogenic.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1992

A highly sensitive enzyme-linked immunosorbent assay for Clostridium perfringens enterotoxin.

A highly sensitive enzyme-linked immunosorbent assay (ELISA) for quantitation of Clostridium perfringens enterotoxin (CPE) by a sandwich method with polystyrene beads was elaborated. The ELISA was very sensitive with a detection limit of 1 pg/ml of CPE. Clostridium perfringens culture fluid did not interfere with the assay. This ELISA may be useful for the mass screening for Cl. perfringens producing small amounts of CPE.

Biotechnology↗
PubMed1992

Gene analyses of abnormal fibrinogens with a mutation in the gamma chain.

In four abnormal fibrinogens with a point mutation in the gamma chain, all characterized by impaired fibrin polymerization, we identified single base exchanges in the respective mutant gamma chain genes by polymerase chain reaction followed by DNA sequence analysis. These base exchanges accounted for the amino acid substitutions previously reported from our laboratory. They were exchanges of C to T (CGC for gamma Arg-275 to TGC for Cys) in fibrinogen Osaka II, T to G (AAT for gamma Asn-308 to AAG for Lys) in fibrinogen Kyoto I, T to C (ATG for gamma Met-310 to ACG for Thr) in fibrinogen Asahi, and G to T (GAT for gamma Asp-330 to TAT for Tyr) in fibrinogen Kyoto III. These base exchanges were found to reside in exon VIII of the gamma chain gene. Since many abnormal molecules are associated with polymerization defects, unless associated with the impaired release of fibrinopeptides A and/or B, exon VIII of the gamma chain gene may deserve careful study to define the structural alterations.

DNA Mutational Analysis↗
PubMed1992

[Coagulation changes during liver resection].

Thrombin-antithrombin III complex (TAT) and plasminogen activator inhibitor (PAI) were measured during liver resection surgery in 8 patients. TAT and PAI activities of patients under liver resection were compared with those of 11 patients under resection of esophageal carcinoma. TAT activity increased during liver resection (P < 0.001) and reached 14 times (P < 0.001) of its control value in the recovery room. PAI activity was very stable during operation, but increased to twice (P < 0.01) of its control value in the recovery room. TAT activity of patients after liver surgery in the recovery room was (P < 0.05) more than twice of that of patients after esophageal surgery. We conclude that hypercoagulable state occurred during liver resection to a greater degree compared with that observed with esophageal surgery, and that its cause might be liver resection itself.

Antithrombin III↗
PubMed1992

[Effects of sevoflurane and isoflurane on intraocular pressure in adult patients].

We investigated effects of two anesthetic agents, sevoflurane and isoflurane, on intraocular pressure (IOP). Forty adult patients were allocated randomly to two groups; Group S (sevoflurane) or Group I (isoflurane). All patients were given a bolus of thiamylal (4-5 mg.kg-1) and vecuronium (0.1-0.2 mg.kg-1) and maintained with 1-3% sevoflurane or isoflurane supplemented with nitrous oxide. Ventilation was controlled and ETCO2 was monitored. IOP, blood pressure and heart rate were measured before induction of anesthesia (control) and at the 7 points of 5, 10, 15, 30, 45, 60 and 120 minutes after intubation. In Group S, IOP was reduced significantly until 30 min compared with control IOP and the maximum decrease was 40% at 10 min. In Group I, IOP was consistently lower than control and the maximum decrease was 40% at the same time as in Group S. But there were no significant differences between the two groups at each point. It seems that the remarkable reductions of IOP after inductions are mainly caused by induction agents. In both groups, hemodynamic parameters showed no remarkable changes during maintenance. These results suggest that both sevoflurane and isoflurane are useful anesthetics for elderly patients receiving ophthalmic surgeries.

Adult↗
PubMed1992

[Influence of calcium channel blockers and beta-blockers on pain relief with iontophoresis].

We studied usefulness of iontophoresis on pain relief using several Ca channel blockers, propranolol and guanethidine. Subjects were 18 healthy adult volunteers. We used 4% lidocaine with/without several drugs (2 mg of nicardipine, verapamil, diltiazem, propranolol and 10mg of guanethidine), and evaluated the pain relief effect with Nakahama's algesimeter. In all groups except for propranolol group, the pain recognition time was elongated significantly in comparison with control. In making comparison between each Ca channel blocker, we did not observe any significant differences, but, in propranolol group, elongation of pain threshold time was observed in some subjects. During these processes, systemic blood pressure and heart rate showed no remarkable changes. Our results suggest that it is possible to achieve more prolonged analgesic effect by the Ca channel blockers.

Adult↗
PubMed1991

Thrombolytic properties of a novel modified human tissue-type plasminogen activator (E6010): a bolus injection of E6010 has equivalent potency of lysing young and aged canine coronary thrombi.

The thrombolytic properties of a novel modified human tissue plasminogen activator (E6010), in which cystein 84 in the epidermal growth factor domain is replaced by serine and that has a prolonged biological half-life, were examined. The thrombolytic efficacies of E6010 and recombinant human tissue plasminogen activator (rt-PA) on the duration of coronary artery thrombus were evaluated in a canine model (123 anesthetized dogs) with copper coil-induced left anterior descending coronary artery thrombus. Thrombi established for periods of 1, 3, or 6 h, as documented by coronary arteriography, were employed. A single bolus i.v. injection of E6010 or rt-PA and an i.v. infusion of rt-PA over 60 min were compared (n = 6). Thrombolytic efficacy was evaluated by three criteria: time to reperfusion (TR), reperfusion rate at 60 min (RR), and reocclusion rate at 60 min after reperfusion (OR). With a bolus i.v. injection of E6010 at a dose of 0.2 mg/kg or an i.v. infusion of rt-PA at a dose of 0.6 mg/kg/h, these parameters were as follows: TR, 30.0 +/- 15.3 and 27.5 +/- 4.8 min; RR, 100 and 100%; OR, 17 and 33% for 1-h aged thrombi; TR, 30.0 +/- 9.5 and 35.0 +/- 8.2 min; RR, 83 and 50%; OR, 20 and 67% for 6-h aged thrombi. These data indicate that a bolus injection of E6010 is almost equally efficacious in lysing thrombi aged both 1 and 6 h. On the other hand, in the case of rt-PA, the thrombi aged 6 h were lysed significantly less than the thrombi aged 1 h. Plasma half-lives of E6010 were t1/2 alpha, 4.8 +/- 0.95 (estimated by antigen level) and 3.0 +/- 0.78 min (estimated by activity), and t1/2 beta, 51 +/- 5.4 (antigen level) and 22 +/- 7.0 min (activity). The half-lives of rt-PA were t1/2 alpha, 3.6 +/- 0.23 (antigen level) and 2.1 +/- 0.61 min (activity), and t1/2 beta, 36 +/- 2.3 (antigen level) and 7.0 +/- 3.5 min (activity). We conclude that a bolus injection of E6010 may have a more potent and longer-lasting effect than i.v.-infused rt-PA in clot lysis therapy.

Aging↗
PubMed1991

[A comparison of three drugs (pethidine, magnesium sulfate and droperidol) in patients with post-anesthesia shivering].

Shivering is one of the common complications after general anesthesia. It increases metabolic rate markedly and may cause myocardial ischemia in patients with decreased cardiopulmonary function. We investigated, therefore, the effects of three drugs (pethidine, magnesium sulfate and droperidol) on shivering in 30 patients under enflurane anesthesia. The patients were of ASA class I or II physical status and had no cardiovascular or respiratory disease. For post-anesthesia shivering, every patient was administered with pethidine 0.5 mg.kg-1, MgSO4 30 mg.kg-1 or droperidol 0.15 mg.kg-1. Their VO2 and VCO2 were measured for 5 minutes during shivering and 15 minutes after drug administration. We also evaluated the therapeutic efficacy of each drug for the treatment of shivering by examining VO2 and VCO2 before and after drug administration. With pethidine and MgSO4, both VO2 and VCO2 decreased significantly after each drug administration. With pethidine, VO2 and VCO2 decreased significantly more than that in MgSO4 (P less than 0.01 vs P less than 0.05). But with droperidol, there was no significant change. These results suggest that pethidine is most effective followed by MgSO4.

Adult↗
PubMed1990

[Intraoperative cardiac output monitoring by the transtracheal Doppler method].

We investigated the cardiac output measured with a transtracheal doppler (TTD) tube in surgical patients and evaluated its usefulness during operations. This tube was located at the point where maximum velocity of blood flow was obtained in the ascending aorta. At this point the cross-sectional area of the ascending aorta was calculated and the cardiac output was obtained by multiplying this area with average blood flow velocity which had been ultrasonically determined. The cardiac output by the TTD method was compared with that of the thermodilution method in 10 patients. The results with both groups showed a direct linear correlation. We obtained the following linear equation: TTD C.O. = 0.61 Thermo C.O. + 1.3 C.O.: cardiac output. The direct correlation coefficient proved to be 0.85 at a risk factor of 0.01. The result suggests that this non-invasive TTD method is easily available and useful in monitoring the intraoperative cardiac output.

Aged↗
PubMed1990

[Clinical characteristics of the lung diseases due to Mycobacterium avium and Mycobacterium intracellulare classified by DNA probe test].

Clinical characteristics of the lung diseases due to M. avium and M. intracellulare classified by DNA probe test were investigated. Between M. avium and M. intracellulare, there was no significant differences in the samples' backgrounds and the clinical characteristics except for their prognoses. The prognosis of the lung diseases due to M. intracellulare was better than those due to M. avium, and M. avium was revealed to be highly susceptible to Cyclocerine than M. intracellulare. No remarkable difference was found in the susceptibility to other antituberculous drugs.

DNA Probes↗
PubMed1987

Hemophilia B (factor IXSeattle 2) due to a single nucleotide deletion in the gene for factor IX.

To understand the molecular basis for hemophilia B in patients with little or no circulating Factor IX antigen, a patient who had less than 0.2% circulating Factor IX antigen (Factor IXSeattle 2) was selected for analysis of his Factor IX gene. Genomic DNA fragments from the abnormal gene were cloned into bacteriophage lambda vectors and recombinant phage were identified using radiolabeled genomic probes obtained from the normal Factor IX gene. The exons and flanking regions of the abnormal gene were sequenced by the dideoxy chain-termination method and this sequence was compared with that of the normal gene. Only one significant difference was observed, the deletion of a single adenine nucleotide in exon V. This resulted in a frameshift that converted an aspartic acid at position 85 in the protein to a valine and the formation of a stop signal at position 86. These data indicate that the gene for Factor IXSeattle 2 codes for an 85 residue polypeptide that terminates after the first epidermal growth factor domain. Thus, the putative Factor IXSeattle 2 polypeptide lacks the second epidermal growth factor domain, the activation peptide, and the catalytic domain present in the normal protein. This provides an explanation for the coagulation disorder in this patient and represents the first report of a single nucleotide deletion and frameshift resulting in hemophilia B.

Adenine↗
PubMed1979

An enzyme immunoassay for the measurement of thyroglobulin in human serum.

An enzyme-linked sandwich immunoassay using silicone rods coated with rabbit (anti-human thyroglobulin) immunoglobulin G and rabbit (anti-human thyroglobulin) monovalent fragment of immunoglobulin F (Fab') conjugated with beta-D-galactosidase was developed for the measurement of thyroglobulin in human serum. The volume of serum needed for the assay was as little as 2 microliters. The sensitivity of the assay was 3.5 ng/ml, which is equal to or rather higher than that of radioimmunoassay. The specificity of the assay was demonstrated by the following observations: (1) The absence of crossreaction of thyroxine and triiodothyronine, (2) non-detectability of thyroglobulin in the sera of patients who underwent total thyroidectomy, (3) parallelism of the standard curve with dilutions of reference serum. The precision of the assay was proven by the demonstration of the sufficient recovery of human thyroglobulin added to sera (92--99%) and coefficients of variance in within and between assays were 6.2--9.3 and 2.5--5.3%, respectively. Furthermore, a highly significant correlation was observed between thyroglobulin concentrations measured by our enzyme immunoassay and those by radioimmunoassay (r = 0.99, p less than 0.001, n = 63). Human thyroglobulin in serum was detectable in 90% of 146 normal subjects, the concentration (mean +/- S.D.) being 13.3 +/- 10.3 ng/ml.

Female↗
PubMed1979

Efficient conjugation of rabbit Fab' with beta-D-galactosidase from Escherichia coli.

An efficient procedure for the conjugation of rabbit Fab' with beta-D-galactosidase from Escherichia coli using N,N-o-phenylenedimaleimide is described. Thiol groups of Fab' were stabilized by the presence of ethylenediaminetetraacetate, and malemide groups were shown to be stable at pH 5 at 4 degrees C. The stability of thiol and maleimide groups enabled an efficient introduction of maleimide groups into Fab' and the average number of maleimide groups introduced into Fab' was 0.76 (range 0.73-0.79; n = 10) per molecule. As a result, 43.4% (range 41.3-46.9%; n = 6) of Fab' used could be conjugated with most of beta-D-galactosidase used. The average number of Fab' molecules conjugated per enzyme molecule was calculated to be 4.2 (range 4.0-4.6; n = 6). Both the enzyme and antibody activities were well preserved in the conjugate. There was no self-coupling of Fab', although the enzyme was polymerized to some extent during the conjugation reaction. The enzyme activity and cross-link in the conjugate was stable at pH 6.0-7.0 at 4 degrees C for at least 3 months.

Adsorption↗