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Biomedical subjects

S Ohtaki

Publications and source records attributed to S Ohtaki.

At least 127 records · Page 7Linked to original sources
PubMed1983

Steady state kinetics and regulation of thyroid peroxidase-catalyzed iodination.

The kinetics of iodination of tyrosine and monoiodotyrosine was studied with purified hog thyroid peroxidase. The Lineweaver-Burk plots and stopped flow kinetic data both yielded a value of about 2 X 10(7) M-1 S-1 as the rate constant for reaction of Compound I with iodide. From Lineweaver-Burk plots, the rate constants for transfer of an assumed enzyme-bound iodinium cation to tyrosine, monoiodotyrosine, and GSH were estimated at 2.1 X 10(6), 3.3 X 10(6), and 3.9 X 10(7) M-1 S-1, respectively. This iodine-transferring reaction was stimulated by thyroxine, triiodothyronine, and diiodothyronine, and the effect increased with the decrease in the number of iodines in the molecules. Diiodotyrosine, during its coupling reaction, inhibited the iodinating reaction by changing the steady state intermediates of thyroid peroxidase from Compound I to Compound II. The quantitative relationship between iodinating and coupling activities of thyroid peroxidase is shown in a scheme, which suggests "preferential formation" of thyroxine from tyrosine by this enzyme system.

Animals↗
PubMed1983

A Japanese family with high density lipoprotein deficiency.

Two siblings with marked reduction of plasma high density lipoprotein (HDL) were found in a Japanese family. Their plasma cholesterol levels were very low (30-60 mg/dl), especially in the HDL fraction (0-1 mg/dl). The concentration of apolipoprotein (Apo) A-I in their plasma was 2-3 mg/dl and that of Apo A-II was 1.5-2.0 mg/dl, determined by means of a single radial immunodiffusion technique. An ultracentrifugally separated HDL fraction contained two different populations of lipoprotein particles, as shown by electron microscopy; a small particle with a diameter of 50-70 A and a relatively large particle at 200 A. Plasma lecithin: cholesterol acyltransferase activity was substantially retained in both cases. Hepatosplenomegaly was present and liver biopsy revealed lipid deposition in reticuloendothelial cells, although the tonsils were apparently normal. No severe atherosclerotic lesions were noticed. The results from these two cases were consistent with the characteristic features of homozygotes of familial HDL deficiency (Tangier disease). HDL cholesterol levels were relatively low in the parents and two children from one patient, which is consistent with the heterozygote state. Two other cases in the kindred were also found to have relatively low HDL cholesterol levels, besides these 4 cases of obligate heterozygotes. Apo A-I and Apo A-II levels in the plasma of the obligate heterozygotes, however, were within the normal range. Plasma low density lipoprotein in the patients moved faster in polyacrylamide gel electrophoresis than those of normal subjects, as did those in the heterozygotes.

Adult↗
PubMed1983

Orientation of thyroid peroxidase in hog thyroid microsomes.

The orientation of thyroid peroxidase in hog thyroid microsomes was studied by trypsin treatment, gel filtration, binding to Concanavalin A Sepharose and iodination of thyroglobulin. Trypsin treatment of microsomes did not solubilize the thyroid peroxidase activity completely but solubilized the NADPH-cytochrome c reductase activity almost completely. The apparent molecular size of thyroid peroxidase was not altered by trypsin treatment of microsomes. It was, however, decreased by the same treatment of deoxycholate-treated microsomes. On the other hand, the apparent molecular size of NADPH-cytochrome c reductase was reduced by trypsin without prior deoxycholate treatment. Thyroid peroxidase of microsomes did not bind to Concanavalin A Sepharose. Thyroglobulin added exogenously was not iodinated by microsomes, but endogenous thyroglobulin, which had been associated with microsomes, was iodinated. Similar results were obtained with rough microsomal membranes prepared from crude microsomes by sucrose density gradient centrifugation. These results suggest that thyroid peroxidase is oriented toward the luminal side of the microsomal vesicles.

Animals↗
PubMed1983

Peripheral K cells in Graves' disease and Hashimoto's thyroiditis in relation to circulating immune complexes.

A plaque assay was employed to quantify the number of peripheral K cells in autoimmune thyroid diseases. The percentages of peripheral K cells determined were 3.1 +/- 2.2% and 3.3 +/- 1.3% (mean +/- SD) in forty-one patients with Graves' disease and twenty-two patients with Hashimoto's thyroiditis, respectively, which were significantly lower than 5.3 +/- 2.5% in 22 normal subjects. Circulating immune complexes (ICs) were measured by three different methods. The positivity of these ICs was investigated in relation to the level of peripheral K cells. The percentage of peripheral K cells in the patients positive for ICs was found to be lower than that in the patients negative for any ICs. Moreover, it was observed that the number of detectable K cells from a normal subject was significantly decreased by incubating the K cells with sera of patients positive for ICs. These results suggest that the decrease of peripheral K cells in the patients with autoimmune thyroid diseases may be due to saturation of K cell Fc receptors by binding ICs.

Adolescent↗
PubMed1983

An enzyme-linked immunoassay for the measurement of circulating immune complexes.

A solid-phase, enzyme-linked immunoassay for the quantitation of circulating immune complexes was developed using C1q. In this assay, the immune complexes in human serum are adhered to C1q-coated polystyrene tubes and then detected by a beta-D-galactosidase-conjugated monovalent fragment (Fab') of rabbit (anti-human IgG, gamma chain-specific) IgG. The enzyme activity bound to the tubes was determined fluorometrically using 4-methylumbelliferyl-beta-D-galactopyranoside as substrate. The sensitivity of this assay was as little as 100 ng/tube, which corresponded to 6 micrograms/ml when 16.7 microliter of serum was assayed. The specificity of the assay was demonstrated by the following observations: (1) absence of cross-reaction of monomeric IgG; (2) non-detectability of immune complex in most of the sera from normal subjects; (3) parallelism of the standard curve with dilution of reference serum. The precision of the assay was proved by the demonstration of sufficient within-assay coefficients of variation (10.7-14.9%).

Animals↗
PubMed1983

Marked reduction of ANAE reactivity of T cells in the lymphoid tissues of monkeys treated with multiple high doses of human beta interferon.

We report a histological study of the organs and tissues of cynomolgus monkeys which had received multiple high doses of human beta interferon (IFN). There were dense aggregations of small lymphocytes without the acid alpha-naphthyl acetate esterase (ANAE) reactivity in the thymus-dependent areas (TDA) of lymphoid tissues, indicating that the T mu cells of IFN-treated monkeys had lost this characteristic enzyme. This change of T mu cells in the tissues was not accompanied by a change in circulating T mu cells.

Animals↗
PubMed1983

Macromolecular alkaline phosphatase and an immunoglobulin G that inhibited alkaline phosphatase in a patient's serum.

Macromolecular alkaline phosphatase (EC 3.1.3.1) was found in the serum of a patient suffering from myasthenia gravis (adult type II) complicated with thymoma, and was shown by immunoelectrophoresis to be bound to immunoglobulins A and G (IgG). Placental alkaline phosphatase, complexed with either the patient's serum or IgG purified from the patient's serum, remained at the origin on electrophoresis, with significant loss of activity. Intestinal alkaline phosphatase, complexed with either the patient's serum or the patient's IgG, migrated to a position similar to that of the macromolecular alkaline phosphatase in the patient's serum on electrophoresis. About 50% of the placental alkaline phosphatase activity was inhibited with 0.1-0.2 g of the patient's IgG per liter, but 6.93 g of the IgG per liter was required for about 20% inhibition of the intestinal alkaline phosphatase activity. The complex of intestinal alkaline phosphatase with the patient's IgG was fairly heat stable. From these results, we concluded that the macromolecular alkaline phosphatase in the patient's serum consisted of intestinal alkaline phosphatase and IgG that was specific for placental alkaline phosphatase.

Adult↗
PubMed1983

The detection of fibrinogen (or FgDP)-protamine sulphate complex in vitro and in the circulating blood.

In order to clarify the pathophysiological significance of the fibrinogen (FgDP)-protamine complex, the interaction between fibrinogen and protamine sulphate was studied in vitro and in vivo. Using the electrophoretic technique, an attempt was made to determine whether the complex could be detected in vitro and in the circulating blood. From the present studies, it has been clarified in in vitro experiments that the slower mobility of two peaks on immunoelectrophoresis represented a complex of protamine sulphate and fibrinogen (FgDP). Furthermore, the peak of slower mobility on crossed immunoelectrophoresis appeared in the circulating blood of a rabbit receiving administration of excess protamine sulphate. That is, in vivo experiments showed that a complex of protamine sulphate and fibrinogen (FgDP) could be detected in the circulating blood.

Animals↗
PubMed1982

One- and two-electron oxidations of tyrosine, monoiodotyrosine, and diiodotyrosine catalyzed by hog thyroid peroxidase.

Stopped flow experiments were carried out with purified hog thyroid peroxidase (A413 nm/A280 nm = 0.42). In the steady state of oxidations of L- and D-tyrosines, N-acetyltyrosinamide, and monoiodotyrosine, thyroid peroxidase existed in the form of Compound I, the primary catalytic intermediate of peroxidase in its reaction with H2O2. Kinetic results led us to conclude that thyroid peroxidase catalyzes two-electron oxidations of these molecules. In the steady state of oxidation of diiodotyrosine, on the other hand, the enzyme was found in the form of compound II at pH 7.4, but in the form of compound I at pH 5.5. The result implies that the mechanism of diiodotyrosine oxidation varied from a one-electron to a two-electron type as the pH decreased. The selection of mechanisms of oxidation appears to be peculiar to thyroid peroxidase; horseradish peroxidase and lactoperoxidase catalyzed only one-electron oxidations of these five donor molecules. Rate constants for rate-limiting steps in the reactions of these donor molecules with the three peroxidases were measured by overall kinetic and stopped flow kinetic methods.

Animals↗
PubMed1982

A sandwich enzyme immunoassay of human growth hormone in serum using anti-human growth hormone IgG-beta-D-galactosidase conjugate.

A sandwich enzyme immunoassay for the measurement of human growth hormone in serum was developed using antibody IgG-beta-D-galactosidase conjugate and antibody IgG-coated silicone rubber rods. The sensitivity of the assay was 4 pg/tube, which corresponded to 0.4 ng/ml, when 10 microliters of serum was used. The assay system showed little cross-reaction to other human pituitary hormones including prolactin, thyroid-stimulating, follicle-stimulating and luteinizing hormones and human chorionic gonadotropin. The coefficients of variation for different levels of serum hGH ranged from 5.8-9.3% and 6.7-14.5%, for within and between run, respectively. The regression equation and coefficient for correlation to radioimmunoassay were gamma (EIA) = 1.06 kappa (RIA) -0.22 and 0.97 (n = 46), respectively.

Animals↗
PubMed1982

Reactions of purified hog thyroid peroxidase with H2O2, tyrosine, and methylmercaptoimidazole (goitrogen) in comparison with bovine lactoperoxidase.

Stopped flow experiments were carried out with purified hog thyroid peroxidase (A413 nm/A280 nm = 0.42). It reacted with H2O2 to form Compound I with a rate constant of 7.8 X 10(6) M-1 s-1. Compound I was reduced to Compound II by endogeneous donor with a half-life of 0.36 s. Compound I was reduced by tyrosine directly to the ferric enzyme with a rate constant of 7.5 X 10(4) M-1 s-1. Tyrosine could also reduce Compound II to the ferric enzyme with a rate constant of 4.3 X 10(2) M-1 s-1. Methylmercaptoimidazole accelerated the conversion of Compound I to Compound II and reacted with Compound II to form an inactivated form, which was discernible spectrophotometrically. The reactions of thyroid peroxidase with methylmercaptoimidazole quite resembled those of lactoperoxidase, but occurred at higher speeds. The absorption spectra of thyroid peroxidase were similar to those of lactoperoxidase and intestinal peroxidase, but obviously different from those of metmyoglobin, horseradish peroxidase, and chloroperoxidase. Similarity and dissimilarity between thyroid peroxidase and lactoperoxidase are discussed.

Animals↗