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Biomedical subjects

K Katoh

Publications and source records attributed to K Katoh.

At least 235 records · Page 13Linked to original sources
PubMed1988

Purification of the Sendai virus nonstructural C protein expressed in E. coli, and preparation of antiserum against C protein.

An expression plasmid, ptac-C, was constructed by inserting the cDNA of the coding region of the Sendai virus nonstructural C protein downstream of the tac promoter of E. coli expression plasmid ptac12-Bam. A new protein produced in E. coli after induction was purified to near homogeneity. The purified protein was found to be identical with the C protein predicted from the C gene cDNA in molecular weight, isoelectric point, amino acid composition, and the amino acid sequence at the N-terminal of the protein as well as those of several fragments obtained on V8 protease digestion. Antiserum raised against the purified protein specifically reacted with the C protein in infected cells. Using this antiserum, the localization of the C protein in infected cells was examined by immunofluorescence, which revealed that it appeared in the cytoplasm but not in nuclei.

Animals↗
PubMed1988

Developmental regulation of the chicken delta 1-crystallin gene: analysis by transgenesis and gene dissection.

We previously reviewed what we had learned about the regulation of the delta 1-crystallin gene through experiments using gene transfer techniques [Kondoh et al. (1986) Cell Differ. 19, 151-160]. It was concluded then that regulatory genetic elements for the lens-specific expression are associated with the delta 1-crystallin gene, and that these chicken elements properly function in mammalian cells. In the last couple of years, we have made significant progress in the understanding of lens-specific delta-crystallin expression. This is owing to success in transgenesis of mouse with the delta 1-crystallin gene and in functional dissection of the gene which led us to the discovery of an intragenic enhancer as the major determinant for lens-specific expression. In this article, we summarize these recent advances.

Animals↗
PubMed1988

Passage of indigestible particles of various specific gravities in sheep and goats.

1. Eight kinds of indigestible particles with specific gravities (SG) ranging from 0.92 to 1.87 (2 mm diameter and 4 mm length) were injected into the reticulo-rumen of sheep and goats in order to investigate the relation between SG and passage through the gastrointestinal tract, and the difference in passage time in the two species of animals. 2. The percentage of excreted particles significantly increased, while the percentage of ruminated particles in the excreted particles significantly decreased, as the SG increased. 3. The daily and cumulative recovery rates of particles of SG 1.38 were slower in sheep than in goats, while those with SG 0.92 were not different between species. 4. It is concluded that particles with SG higher than 1.27 would pass through the gastrointestinal tract of sheep and goats more quickly than those with SG lower than 1.21.

Animals↗
PubMed1988

Physicochemical properties of charge isomers of recombinant human superoxide dismutase.

Recombinant human Cu2Zn2SOD expressed in Escherichia coli consisted of mainly three isomers with isoelectric points of 5.14 (A), 5.06 (B), and 4.99 (C). Each isomer was isolated by DEAE-Toyopearl chromatography and the physiochemical properties were investigated. No significant differences in chemical and spectrophotometric properties, such as specific activity, metal contents, amino acid composition, and UV and ESR spectra, were found. The result of labeling of free cysteine residues with ABD-F showed the disulfide bond to be formed between 57Cys and 146Cys in every isomer. A few differences were found in the CD spectrum around 260 nm and in the elution patterns on reverse-phase HPLC. The isoelectric points of the three isomers became the same after treatment by reduction and carboxymethylation and even after reduction only, pI of isomers tended to be at the value of component (A). These results suggest that the three isomers are identical in primary structure but slightly different in secondary or tertiary structure. These differences are probably derived from structural alterations around 111Cys.

Amino Acids↗
PubMed1988

Comparison of properties between human recombinant and placental copper-zinc SOD.

The physicochemical properties of purified recombinant human copper-zinc superoxide dismutase (r-hSOD) were compared with those of human placental copper-zinc superoxide dismutase (h-SOD). No differences were found in specific activity, metal contents, amino acid composition, and tryptic peptide map. The spectrophotometric properties including UV, ESR, and CD spectra were also similar. The result of isoelectric gel electrophoresis showed that the difference in isoelectric point (pI) was derived from acetylation of the N-terminal amino acid (alanine) in h-SOD. In SDS-polyacrylamide gel electrophoresis, both SODs showed the same behavior and enzymic activity was retained only under non-reducing conditions. ESR analysis of the denatured enzyme suggested that the high stability was derived from the structure of the active site around copper. Experiments using other metal-substituted SODs (Cu, Co in place of zinc) suggested that zinc contributed to the stability and the unique electrophoretic behavior of the enzyme.

Amino Acid Sequence↗
PubMed1988

Relation between ESR-detectable Cu(II) and superoxide dismutase activity.

The relation between ESR-detectable Cu(II) and Cu,Zn-superoxide dismutase activity was examined. The Cu(II) spin numbers per one unit of SOD were 6.26 X 10(12) (+/- 0.51 X 10(12] spins in several preparations of recombinant human Cu,Zn-SOD, native placental, and erythrocyte SOD. Measurement could be performed over a wide range of pH (4.0-10.0), preferably at temperatures below -40 degrees C. The data obtained by this method correlated well to the results obtained by the method of Fridovich et al. using the xanthine-xanthine oxidase system (correlation coefficient 0.995). The specific activity of SOD was proportional to the Cu(II) content measured by ESR, but not to the total Cu content measured by atomic absorption. This indicates that it is important to measure the Cu(II) content for determining Cu,Zn-SOD activity.

Copper↗
PubMed1988

Subnuclear localization and antitransforming activity of N-myc:beta-galactosidase fusion proteins.

N-myc expression is under stage- and tissue-specific regulation in mammalian development, but its function is totally unknown. We sought agents to block N-myc activity in order to infer from the effect the possible function of N-myc in the apparently complex processes. As candidates for such agents, we tested fusion genes encoding N-myc:beta-galactosidase fusion proteins for their effects on the formation of transformed foci of rat embryo primary fibroblasts as the result of transfection with N-myc and activated H-ras. One of the gene constructs very efficiently antagonized N-myc activity, as assessed by its effect on focus formation, but did not appreciably affect cell viability. The product of this gene was not only targeted to the nucleus but also accumulated in subnuclear loci which may represent the sites where normal N-myc proteins reside. The occurrence of antagonistic effect at a low stoichiometric ratio suggested that the fusion protein gene competed with the N-myc gene in a fashion analogous to a dominant negative mutation.

Animals↗
PubMed1988

Embryonic stem cell-mediated transfer and correct regulation of the chicken delta-crystallin gene in developing mouse embryos.

To study regulation of delta-crystallin expression during ontogeny, we transferred the gene from chicken into developing mouse embryos by first transforming an embryonic stem (ES) cell line of mouse and then producing chimaeric embryos by combining them with normal mouse embryos. Using this technique, genes were transferred into a variety of developing mouse tissues with high efficiency. Two delta-crystallin gene constructs were used: the wild-type gene with 2200 bp of the 5' flanking sequence, shown to be lens-specific in an assay using cultured mouse cells, and a mutant gene with 51 bp of the 5' flanking sequence, lacking the sequence required for expression in lens cells. Five independent lines carrying the former and two lines carrying the latter were employed in producing chimaeras. In the chimaeric embryos having the wild-type gene, delta-crystallin was expressed in the lens and in specific regions of the primitive central nervous system (CNS) as is seen in embryonic expression in the chicken. In adult mouse chimaeras also, expression was restricted to the lens and the CNS, in the pyramidal neurones of the piriform cortex and the hippocampus. delta-crystallin expression in these tissues is due to proper transcriptional regulation, since no expression was observed when chimaeras were produced with the ES lines carrying the mutant gene. The experimental results reported here demonstrate the advantage of ES-cell-mediated gene transfer in the study of embryonic gene regulation, because a number of gene constructs and chromosomal sites can be analysed shortly after embryo manipulation without requiring gene transmission to the next generation.

Animals↗
PubMed1988

Activity and stability of recombinant human superoxide dismutase in buffer solutions and hypothermic perfusates.

The stability of recombinant human superoxide dismutase (r-hSOD) in buffer solutions was studied in solutions at various pH and temperatures. Additionally, we studied the effects of incubation with proteases, serum and two types of hypothermic perfusates. R-hSOD was stable in the pH range of 6-11 and at temperatures up to 80 degrees C for 30 min. R-hSOD activity was not affected by incubation with trypsin, aminopeptidase M or serum for 2 h. R-hSOD activity determined at various temperatures (4-37 degrees C) did not vary remarkably. R-hSOD in hypothermic perfusates was stable at 4-37 degrees C for 24 h.

Buffers↗
PubMed1988

The effects of drug metabolism inducers on the delayed neurotoxicity and disposition of tri-o-cresyl phosphate in hens following a single intravenous administration.

Intravenous (iv) administration of tri-o-cresyl phosphate (TOCP) caused a delayed neurotoxic effect in hens similar to that produced by oral and dermal administration. The iv ED50s for producing ataxia and paralysis were estimated to be 15.9 mg/kg and 31.7 mg/kg respectively. The tissue disposition of unaltered TOCP was determined in hens following a single iv injection of 40 mg/kg of TOCP. One hour (hr) after the injection, the leg muscle contained the highest concentration, 26.99 micrograms/g fresh weight followed by the adipose tissue. Among the nerve tissues, the sciatic nerve had the highest concentration, 9.63 micrograms/g followed by the spinal cord and the brain. Except the adipose tissue and the sciatic nerve, the concentration in all analyzed tissues dropped below 1.0 microgram/g (ml) after 24 hr. An unidentified metabolite appeared in bile taken 1 and 3 hr after the injection. Pretreatment of hens with 3-methylcolathrene (3-MC) and beta-naphthoflavone (B-NF) protected against the TOCP-induced delayed neurotoxicity, whereas phenobarbital (PB) failed to protect against the neurotoxicity. Plasma creatine phosphokinase (CK) activity in paralytic birds increased approximately 4 times of the control or symptomless hens on the 21st day. 3-MC-, B-NF- and PB-treatment depressed substantially the concentration of unaltered TOCP in brain and plasma 1 hr after iv dosing with 40 mg/kg of TOCP. Only B-NF pretreatment lowered the level of TOCP in spinal cord. There was no effect of these inducers on the level of TOCP in sciatic nerve and adipose tissue. B-NF and 3-MC lowered significantly the TOCP level in leg muscle, whereas PB had no such effect. More attention should be paid to the role of TOCP in muscle, especially to the leg muscle, judging from the present toxicological and metabolic studies.

Animals↗
PubMed1988

[An analysis of quadriceps muscle force in boys with Osgood-Schlatter disease].

Osgood-Schlatter disease is thought to be a consequence of microavulsions caused by repeated traction on the tibial tuberosity. For determination of the etiologic factor of this disease, concentric and eccentric force of the quadriceps muscle was measured in the unaffected side and compared with that of normal boys and girls, normal men and women, and patients with cruciate ligament injury of the knee. The relation between the maximum force and its angle velocity was approximated and the ratio of the eccentric force to the concentric force was calculated at 90 deg/sec. In the group with Osgood-Schlatter disease, the eccentric force was especially strong and this group had the highest ratio among all groups. This finding suggests that the increased eccentric force in the extensor mechanism of the knee is an etiologic factor in the disease.

Adolescent↗