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T Kotani

Publications and source records attributed to T Kotani.

At least 181 records · Page 10Linked to original sources
PubMed1989

Characterization of rat and human steroid sulfatases.

Rat and human steroid sulfatases were purified from liver and placenta, respectively, by the same procedure. The rat and human enzymes were solubilized with Triton X-100, and purified by immunoaffinity chromatography with a monoclonal antibody showing high binding activities to both the enzymes. They were further purified by high-pressure anion-exchange chromatography to compare their structural and catalytic properties. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that both enzymes had a molecular weight of 62,000. The enzymes had similar amino acid compositions and amino-terminal amino acid sequences. Significant differences of the optimum pH, Michaelis constant and maximum velocity were observed between these enzymes. The optimum pH of each enzyme varied from 6.0 to 8.0, depending on substrates and with or without Triton X-100. In detergent-free media, steroid sulfates competitively inhibited the ability of these enzymes to hydrolyze 4-nitrophenyl sulfate. In media containing Triton X-100, on the other hand, the inhibition types of the steroid sulfates on the hydrolyzing activities of the rat and human enzymes were noncompetitive- and mixed-types, respectively.

Amino Acid Sequence↗
PubMed1989

Thyroglobulin-mediated one- and two-electron oxidations of glutathione and ascorbate in thyroid peroxidase systems.

The one- or two-electron oxidation of thyroglobulin by the thyroid peroxidase system was found to be regulated by the iodine content of thyroglobulin. The catalytic intermediate of thyroid peroxidase observed at steady state of the reaction was Compound I and II when the iodine content in thyroglobulin was 0.2 and 0.7%, respectively, apparent rate constants for the rate-limiting steps being estimated at 4.7 x 10(7) and 4.8 x 10(4) M-1 S-1. The thyroglobulin-mediated oxidation of GSH occurred by way of two-electron transfer at 0.2% iodine content and by way of one-electron transfer at 0.7% iodine content. The spin-trapping experiment with 5,5-dimethyl-1-pyrroline-N-oxide showed that glutathione radicals were formed in the latter reaction but not in the former. In the reactions of thyroid peroxidase, the one- and two-electron oxidations of ascorbate were also mediated by 0.2 and 0.7% iodine thyroglobulins, respectively. The reactions were analyzed and mimicked with the use of p-cresol and p-acetaminophenol as a mediator in the reactions of lactoperoxidase and thyroid peroxidase.

Acetaminophen↗
PubMed1989

cDNA-directed expression of human thyroid peroxidase.

A human thyroid peroxidase cDNA, hTPO-1 [(1987) Proc. Natl. Acad. Sci. USA 84, 5555-5559], was expressed in human Hep G2 cells using a vaccinia virus cDNA-expression system. When examined by immunoblot analysis, the level of hTPO-1 protein expression reached a maximum approx. 24 h after infection and remained at a similar level up to 72 h post-infection. The expressed protein was enzymatically active as measured by guaiacol oxidation. Monoclonal antibody-assisted immunoaffinity column chromatography was used for partial purification of vaccinia-expressed hTPO-1, resulting in more than 300-fold higher specific activity and a measurable difference spectrum of the hTPO-1 (Fe3+)-CN complex.

Blotting, Western↗
PubMed1989

Structure of the human thyroid peroxidase gene: comparison and relationship to the human myeloperoxidase gene.

All exons of the human thyroid peroxidase gene were cloned from phage and cosmid libraries and sequenced, including 2599 base pairs of upstream DNA. The gene contains 17 exons and covers at least 150 kilobase pairs of chromosome 2. The transcription start site was identified by both S1 mapping and primer extension; a typical TATA box was found 25 bases upstream of the putative start site. A comparison of the gene structures of thyroid peroxidase and a granulocyte protein, myeloperoxidase, revealed that the positions of the 3rd through 11th exon-intron junctions in thyroid peroxidase coincide exactly with those of the 2nd through 11th exon-intron junctions in myeloperoxidase except the 7th myeloperoxidase junction, that does not have any counterpart in thyroid peroxidase. The amino acid codon separation pattern in each junction is well conserved between both enzymes. Four exons, unique to thyroid peroxidase, are located at the 3' end of the gene (exons 13-16), each of which encompasses a different protein module. Three of these modules, representing exons 13, 14, and 15, bear significant similarities to C4b-beta 2 glycoprotein, the EGF-LDL receptor, and a typical transmembrane domain, respectively. The genes coding for these modules were probably fused to an ancestral peroxidase gene to generate the present thyroid peroxidase gene. The data suggest that intron loss, and/or insertion, and exon shuffling have played important roles in the evolution of the thyroid peroxidase gene.

Amino Acid Sequence↗
PubMed1989

Resonance Raman characterization of hog thyroid peroxidase. An SERRS study.

Resonance Raman (RR) spectra of hog thyroid peroxidase (TPO) were observed for the first time and compared with those of lactoperoxidase (LPO) and horseradish peroxidase (HRP). Since TPO purified by monoclonal antibody-assisted immunoaffinity chromatography was strongly fluorescent, the surface enhancement technique using Ag colloid adsorption was used for the oxidized form, but ordinary RR spectra could be obtained for the reduced form. The RR spectra of TPO were distinct from those of HRP in both the oxidized and reduced states and indicated the presence of six-coordinated iron-protoporphyrin.

Animals↗
PubMed1989

A monoclonal antibody to rat liver arylsulfatase C and its application in immunohistochemistry.

We purified arylsulfatase C from rat liver microsomes and prepared a monoclonal antibody (P42C2) to the purified enzyme. By SDS-PAGE and immunoblotting analysis using P42C2, the molecular weight of the purified enzyme and of the enzyme in liver and kidney microsomes were estimated at 62,000 daltons. P42C2 caused little inhibition of arylsulfatase C activity, and was bound only slightly to liver microsomes. Localization of arylsulfatase C was studied at the light and electron microscopic level by the indirect immunoperoxidase method using P42C2. In rat liver, arylsulfatase C was detected mainly in the hepatocytes, and less frequently in endothelial cells, Kupffer's cells, and Ito's cells. In rat kidney, strong staining was observed in the straight portions of the proximal tubules. The podocytes, interstitial cells, endothelial cells, and epithelial cells of Henle's thin limbs were stained faintly. By electron microscopy, arylsulfatase C was found localized on the membranes of the endoplasmic reticulum and nuclear envelopes in these cells. These immunohistochemical findings agree with the localization demonstrated by an enzyme-histochemical method which we had previously developed.

Animals↗
PubMed1989

[Autoantigens in autoimmune thyroid diseases].

Human thyroid peroxidase (TPO) was found to bind with both IgG and IgM from patients with autoimmune thyroid diseases. Furthermore, binding of IgG to microsomes was inhibited by TPO. Patients' IgG that passed through an antigen column packed with TPO-coupled Sepharose 4 B beads lost their hemagglutinating activity in microsome hemagglutination test. Various strains of mice were immunized with porcine TPO emulsified in complete Freund's adjuvant. Both C 57 BL/6 and C 57 BL/10 mice showed mononuclear cell infiltration in the thyroid with very high incidences of thyroiditis. The strains of mice showing thyroiditis were very different from high responder strains that were immunized with murine thyroglobulin. Cloning of human TPO cDNA was attempted to deduce the primary structure of human TPO. Using three kinds of oligonucleotides as probes, two cDNAs, 2.8 and 2.4 kb, were selected from a cDNA library constructed from mRNA purified from Graves' disease thyroid. Sequencing of cDNAs revealed that shorter cDNA lacked 171 nucleotides at the middle portion of the longer cDNA. Primer extension analysis of mRNA indicated that the full length cDNA of human TPO consists of 3,048 nucleotides and its open reading frame codes 933 amino acids.

Animals↗
PubMed1989

[Measurement of coefficient of variation of R-R interval in ECG for patients with impotence].

The coefficient of variation (CV = SD/mean x 100) of the R-R interval in electrocardiogram (ECG) was measured as an indication of parasympathetic nerve function in patients with impotence. Of 762 patients 104 (13.6%) had a CV value lower than 2.0. The factors which led to the low CV value were evident in 50 patients (48%), i.e. diabetes mellitus in 26, side effects by antiulcer, antidepressant or antihypertensive agent in 16, renal failure in 1, alcoholism in 2 and age over 70 years old in 4. The possible factors of the low CV value, such as neurogenic, thyroid, cardiovascular and endocrine disorders were found in 12 patients (11.5%). In the remaining 42 patients (40.4%), CV value was decreased by unknown factors. Measurement of CV of R-R interval in ECG was safe and useful as a screening test for patients with impotence to detect the dysfunction of the parasympathetic nerve system.

Adult↗
PubMed1989

[Measurement of penile brachial index (PBI) in patients with impotence].

PBI was measured in 555 patients chosen at random from those who visited our impotence clinic between October 1984 and September 1986. Of 555 patients with impotence 29 (5.2%) showed a low PBI level of less than 0.6. On the basis of carefully taken history, risk factors causing the disturbance of penile blood flow were found in 13 (44.8%) of these 29 patients. They included diabetes mellitus, hypertension, surgery of pelvic organ, vertebral injury, vascular disorder of brain and gangliosympathectomy. Although penile angiography is necessary to make the definite diagnosis of vasculogenic impotence, hospitalization is usually required to perform the examination. Measurement of PBI is useful as a screening test to detect vasculogenic impotence at an outpatient clinic, because this examination is noninvasive and technically easy.

Adolescent↗
PubMed1988

Immunoelectron microscopy of the hormonogenic sites of the thyroglobulin molecule.

Hog thyroglobulin was treated with the monoclonal anti-thyroglobulin antibody, 16.3.2, which recognizes the thyroxine residues of thyroglobulin. Size-exclusion high-performance liquid chromatography of the immune complexes indicated that there were 3.5 immunoreactable thyroxine residues/mol of 19 S thyroglobulin. The immune complexes were negatively stained and observed under an electron microscope. Under antigen excess conditions, the complexes were demonstrated as branched chains in which thyroglobulin molecules were linked by antibody molecules. When the antibody was in excess, we found many single thyroglobulin molecules each connecting to 2-3 antibody molecules. Based on the morphology of the immune complexes, we conclude that thyroxine residues are formed preferentially on the molecular surface and localized near both ends of the elongated molecule of 19 S thyroglobulin and possibly at one side of the middle of the molecule.

Animals↗
PubMed1988

[Chlamydia trachomatis--detection of inflammatory diseases of the fallopian tubes].

The prevalence of Chlamydia trachomatis in tubal swabs obtained by laparoscopy was investigated in a prospective study. Specimens were collected from women with salpingitis (group 1), tubal infertility (group 2) and from controls, which were considered as being not infected and subjected to laparoscopy for other reasons (group 3). In the period 12/1980-8/1984 chlamydial infection was diagnosed, by means of the McCoy tissue culture method, in group 1 in 19% (16/85), in group 2 in 4% (10/250) and in group 3 in 0% (0/122). In a second period until 6/1985, positive chlamydial cultures were seen in 2% (1/44), 2% (2/110) and 0% (0/75) respectively, however, a simultaneously used IFT (Micro Trak, Syva Merck) was negative in all cases. In the third period until 12/1986, positive chlamydial cultures were obtained in 12% (9/77), 1% (1/137) and 4% (4/90) respectively. The sensitivity of a simultaneously performed EIA (Chlamydiazyme, Abbott) in women with salpingitis was 67%, the specificity was 97%. The EIA seemed to provide an alternative diagnostic method to culture when evaluating tubal specimens. In an additional trial, chlamydial complement fixation test was positive in 26% of a total of 90 women with tubal infertility (and negative tubal culture) vs. 6% in a control group of pregnant females (p less than 0.001). In conclusion, the reported results support the etiologic role of Chlamydia trachomatis as an important agent in salpingitis as well as in tubal infertility.

Bacteriological Techniques↗