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Biomedical subjects

N Shimojo

Publications and source records attributed to N Shimojo.

At least 127 records · Page 7Linked to original sources
PubMed1989

Test-strip method for measuring lactate in whole blood.

We have developed a dry reagent strip system for measuring lactate in whole blood. The test strip contains lactate oxidase (no EC number assigned), horseradish peroxidase (EC 1.11.1.7), and N-ethyl-N-(2-hydroxy-3-sulfopropyl)-m-toluidine. The system is designed to measure with a reflectometer the color that developed in the test strip, although the lactate concentration can be estimated without the reflectometer. The between-run coefficients of variation for controls at three concentrations were 2.9-5.3%. The lactate concentrations in blood samples from healthy subjects before and after exercise correlated well (r = 0.97) with the results measured by the comparison method with the use of lactate oxidase. This dry reagent strip system provides a convenient and rapid test for measuring blood lactate in clinical and sports medicine.

Horseradish Peroxidase↗
PubMed1988

Plasma fructosamine assay in children with insulin-dependent diabetes mellitus.

Plasma concentrations of fructosamine, an indicator of glycated plasma proteins, were measured in non-diabetic children and children with insulin-dependent diabetes mellitus (IDDM) to see if they also correlate with glycemic control in children as well as in adults. Non-diabetic children aged less than 4 yr had significantly lower plasma fructosamine than non-diabetic children aged 4 or more. Total plasma protein in these children was slightly lower or close to that of older children. There was no difference in fructosamine between non-diabetic children aged 4 or more and healthy adult subjects. Plasma fructosamine in children with IDDM was twofold that of age-matched controls. In children with IDDM, correlations between fructosamine and HbAI (r = 0.799) or HbAIc (r = 0.841) were high. The measurement of plasma fructosamine, which is practical in children because of the small sample volume needed and no influence of HbF, is useful in the management of children with IDDM.

Adolescent↗
PubMed1988

Mutations in the alpha 2 helix of HLA-A2 affect presentation but do not inhibit binding of influenza virus matrix peptide.

Previous studies have suggested that MHC class I molecules bind and present peptides to CTL in a manner that is analogous to the presentation of peptides by class II molecules to Th. Crystallographic studies of HLA-A2 have led to the assignment of a putative peptide binding site that is bordered by two alpha helices consisting of residues 50-84 and 138-180. In this study, we have investigated whether residues in the alpha 2 helix are involved in the binding and/or presentation of a peptide to CTL. We have generated CTL to type A influenza virus by stimulation of human PBL with a synthetic peptide from the influenza A virus matrix protein (M1 residues 57-68) in the presence of rIL-2. Such HLA-A2.1-restricted influenza virus-immune CTL do not recognize infected HLA-A2.3+ targets. A2.1 and A2.3 differ by three amino acids in the alpha 2 domain: Ala vs. Thr at position 149, Val vs. Glu at position 152, and Leu vs. Trp at position 156. Site-directed mutants of the A2.1 gene that encode A2 molecules that resemble A2.3 at positions 149, 152, and 156 have been constructed, transfected into human cells, and assayed for their ability to present the M1 peptide. The results demonstrate that most, but not all, A2.1-restricted M1-peptide-specific CTL fail to recognize M1 peptide-exposed transfectants with certain single amino acid substitutions at positions 152 and 156. In contrast, M1 peptide-exposed transfectants that express A2 molecules with an Ala----Thr substitution at position 149 were recognized by all CTL tested, but they exhibited an apparent difference in the kinetics of peptide binding. These results indicate that amino acid substitutions at positions 152 and 156 of the putative peptide binding site of the A2 molecule can affect presentation without eliminating binding, and indicate that the failure to recognize complexes between the peptide and the mutant A2 molecules is due to different TCR specificities and not to the failure to bind the peptide.

Adult↗
PubMed1988

Induction of renal cytochrome P-450 in hepatic microsomes of diabetic rats.

We purified two forms of cytochrome P-450 which was induced in hepatic microsomes of diabetic male rates treated with streptozotocin. One of these corresponded to P-450j. The other form, designated P450 DM-2, had a minimum molecular weight 53000 and a CO-reduced absorption maximum at 452 nm. The P450 DM-2 efficiently catalyzed the omega- and (omega-1)-hydroxylation of lauric acid, but was not efficient in metabolizing aminopyrine, 7-ethoxycoumarin, aniline, N-nitrosodimethylamine, or testosterone. The NH2-terminal sequence of P450 DM-2 was identical to that of P450 K-5, the major renal cytochrome P-450. Both forms gave very similar electrophoretic patterns of proteolytic digests. P450 DM-2 and P450 K-5 are closely related forms.

Amino Acid Sequence↗
PubMed1988

Biochemical effects of combined gases of nitrogen dioxide and ozone. II. Species differences in lipid peroxides and antioxidative protective enzymes in the lungs.

Changes in lipid peroxide (thiobarbituric acid reactant) levels, in the content of non-protein sulfhydryls (NPSH) and total proteins, and in the activities of antioxidative protective enzymes were examined in the lungs of four animal species exposed to a mixture of NO2 and O3 for 2 weeks. Male mice, hamsters, rats and guinea pigs were used. Thiobarbituric acid (TBA) reactant levels were increased significantly in the lungs of mice and guinea pigs, but not in hamsters and rats. NPSH contents were increased markedly in hamsters, mice and rats, but not in guinea pigs. The activities of antioxidative protective enzymes also changed with the exposure. The most characteristic change was the significant increase in glutathione peroxidase (GPx-H2O2) activity in hamsters and rats - species which did not exhibit increases in their TBA reactant levels. The increase in this enzyme activity in mice was significant, but not very large. Furthermore, guinea pigs were genetically deficient in this enzyme, and the increase in glycolytic enzymes for regenerating NADPH was also lowest in guinea pigs. The glutathione S-transferase (GSH-Tase) activity in mice and guinea pigs was decreased by exposure to the combined gases. These results suggest that the increases in lipid peroxide levels in mice and guinea pigs may be due to a lesser ability to regenerate protective reducing substances, such as NPSH and NADPH, than that of hamsters and rats. Induction of protective enzyme activities on exposure to the combined gases was also poor in mice and guinea pigs.

Animals↗
PubMed1988

Antigenic determinants on thyroglobulin: comparison of the reactivities of different thyroglobulin preparations with serum antibodies and T cells of patients with chronic thyroiditis.

To delineate the antigenic determinants on thyroglobulin (Tg) recognized by serum autoantibodies and peripheral blood T cells from patients with chronic thyroiditis, we studied the reactivities of three different Tg preparations, i.e. enzyme-digested Tg fragments, physically or chemically denatured Tg, or Tg with differing iodine contents. Human Tg was digested with staphylococcal V8 protease, and the fragments were separated by high performance liquid chromatography. The autoantibodies reacted with the larger fragments, but their ability to bind to small fragments was limited. On the other hand, T cells reacted similarly with all fragments, regardless of mol wt. The autoantibodies bound little to denatured Tg after its disulfide bonds were destroyed with dithiothreitol or 2-mercaptoethanol, while the reactivity of heat-denatured Tg was partially decreased, and that of Tg denatured with sodium dodecyl sulfate was conserved. Conversely, T cells reacted with Tg denatured by heating or dithiothreitol treatment. These results indicate that autoantibodies recognize mainly a conformational structure of Tg, presumably containing disulfide bonds, whereas T cells recognize the primary structure of Tg. Variations in the iodine content of Tg were not associated with altered reactivity with autoantibodies or T cells. We propose that variations in Tg conformation related to iodination of the molecule do not contribute significantly to its reactivity with autoantibodies and T cells. In addition, T cells reacted with the smaller Tg fragments containing few T3 or T4 residues to a greater extent than they did with larger Tg fragments with the same amount of T3 or T4 as native Tg. Therefore, it appears that the Tg-reactive T cells predominantly recognize determinants on the Tg molecule that are unrelated to hormone-containing sites.

Adolescent↗
PubMed1988

Thyroglobulin and thyroid peroxidase share common epitopes recognized by autoantibodies in patients with chronic autoimmune thyroiditis.

Monoclonal antibodies specific for human thyroid peroxidase (TPO) were prepared by the hybridoma technique using hyperimmune spleen cells from mice immunized with TPO purified from thyroid glands from patients with Graves' disease. Use of the microenzyme-linked immunosorbent assay method revealed that some of the monoclonal antibodies cross-reacted strongly with human thyroglobulin (Tg). Conversely, monoclonal anti-Tg antibodies cross-reacted with TPO, albeit to a lesser degree. Some anti-Tg autoantibodies in serum from patients with chronic autoimmune thyroiditis purified by Tg affinity chromatography bound TPO, and such binding was completely inhibited by Tg. Western blotting experiments revealed that thyroid microsomal 103K proteins recognized by mouse monoclonal and polyclonal anti-TPO antibodies were recognized by some monoclonal anti-Tg antibodies and anti-Tg autoantibodies, and conversely, that 19S Tg was recognized by some monoclonal anti-TPO antibodies. TPO was immunoprecipitated by anti-Tg autoantibodies isolated by Tg affinity chromatography. On the other hand, the specificity for TPO of the anti-Tg autoantibodies was not identical with that of anti-TPO autoantibodies. These cross-reactivities were not due to contamination of TPO with Tg or vice versa, or to contamination of the anti-Tg autoantibody preparations with anti-TPO autoantibodies. Taken together, these data indicate that Tg and TPO share common antigenic determinants and that some of those determinants are recognized by autoantibodies in the serum of patients with chronic autoimmune thyroiditis.

Adolescent↗
PubMed1988

The distribution of total mercury in the brain after the lateral ventricular single injection of methylmercury and glutathione.

We determined the time course of the distribution of total mercury in the brain after the lateral ventricular single injection of methylmercury (Me-Hg) and glutathione (GSH) to confirm our assumption that glutathione have a very important role in methylmercury transport into the brain. The following results were obtained: (1) The transport of methylmercury into the brain was accelerated by glutathione, but was retarded by the surplus glutathione. (2) The increase of glutathione to methylmercury ratio in the dose was tend to uniformize the distribution of total mercury in the brain. These results might suggest that methylmercury is carried with glutathione and transformed into methylmercury-cysteine by gamma-glutamyl transpeptidase (gamma-GTP), and transported into the brain.

Animals↗
PubMed1988

Purification and characterization of diabetes-inducible cytochrome P-450.

A diabetes-inducible form of cytochrome P-450, termed P-450DM, was purified to electrophoretical homogeneity (MW 51,000) by high-performance liquid chromatography from liver microsomes of diabetic rats induced with streptozotocin. The CO-reduced absorption maximum of P-450DM was at 452 nm and the oxidized heme iron appeared to be predominately in the high-spin state as deduced from the Soret maximum at 395 nm. P-450DM was active in aniline hydroxylation and N-nitrosodimethylamine demethylation. The dealkylation activity toward 7-ethoxycoumarin by P-450DM was much enhanced by the addition of cytochrome b5.

Animals↗
PubMed1987

Identification and characterization of a tissue kallikrein in rat skeletal muscles.

A tissue kallikrein was purified from rat skeletal muscle. Characterization of the enzyme showed that it has alpha-N-tosyl-L-arginine methylesterase activity and releases kinin from purified bovine low-Mr kininogen substrate. The pH optimum (9.0) of its esterase activity and the profile of inhibition by serine-proteinase inhibitors are identical with those of purified RUK (rat urinary kallikrein). Skeletal-muscle kallikrein also behaved identically with urinary kallikrein in a radioimmunoassay using a polyclonal anti-RUK antiserum. On Western-blot analysis, rat muscle kallikrein was recognized by affinity-purified monoclonal anti-kallikrein antibody at a position similar to that of RUK (Mr 38,000). Immunoreactive-kallikrein levels were measured in skeletal muscles which have different fibre types. The soleus, a slow-contracting muscle with high mitochondrial oxidative-enzyme activity, had higher kallikrein content than did the extensor digitorum longus or gastrocnemius, both fast-contracting muscles with low oxidative-enzyme activity. Streptozotocin-induced diabetes reduced muscle weights, but did not alter the level of kallikrein (pg/mg of protein) in skeletal muscle, suggesting that insulin is not a regulator of kallikrein in this tissue. Although the role of kallikrein in skeletal muscle is unknown, its localization and activity in relation to muscle functions and disease can now be studied.

Animals↗
PubMed1987

Enzyme-linked immunosorbent assay (ELISA) for IgG antibodies to thyroglobulin.

We used a computer programmed standard IgG curve for computer-assisted quantification of assay results for autoantibodies to thyroglobulin (Tg) by quantitative enzyme-linked immunosorbent assay (ELISA). Specific antibody levels in unknowns were quantified by comparison of their optical density readings with a standard curve of absorbance vs concentration obtained with dilutions of the reference serum. Anti-Tg antibodies were detected in 80% of the patients with chronic thyroiditis and 90% of those with Graves' disease. Anti-Tg antibodies were also detected in 14.3% of the healthy controls. The titer of anti-Tg antibodies detected by tanned red cell hemagglutination correlated well with that detected by ELISA, although, the sensitivity of the ELISA was higher. By our computer-assisted conversion method, the anti-Tg antibody can be readily and reliably quantified and low titer antibodies to Tg can be detected with adequate precision.

Adolescent↗
PubMed1987

Detection of sulfur-containing compounds in control and cadmium-exposed rat organs by high-performance liquid chromatography--vacuum-ultraviolet inductively coupled plasma-atomic emission spectrometry (HPLC-ICP).

Sulfur-containing compounds in biological samples were separated by high-performance gel permeation chromatography and detected with a vacuum-ultraviolet inductively coupled plasma-atomic emission spectrometer. Distribution profiles of sulfur in the supernatants of liver, kidney, spleen, lung, and pancreas of control and cadmium-exposed rats were determined along with cadmium, copper, iron, phosphorus, and zinc profiles. Changes in sulfur distribution were induced by cadmium exposure not only in the metallothionein fraction, but also in the high-molecular-weight protein fraction, indicating the effect of cadmium exposure on diverse endogenous sulfur-containing compounds. Glutathione and taurine also were detected simultaneously as distinct peaks.

Animals↗