Biomedical subjects

S Takeda

Publications and source records attributed to S Takeda.

At least 19 recordsLinked to original sources

PubMed1992

Hepatitis C virus infection in hepatocellular carcinoma. Detection of plus-strand and minus-strand viral RNA.

BACKGROUND: Although serum antibody to hepatitis C virus (anti-HCV) is found in many patients with hepatocellular carcinoma, the actual roles of HCV in carcinogenesis are unknown. METHODS: With reverse transcription followed by the polymerase chain reaction (RT-PCR), HCV RNA was examined in the sera and liver tissues of 16 patients with hepatocellular carcinoma who did not have hepatitis B virus markers, 13 of whom had anti-HCV. RESULTS: In the 13 patients with anti-HCV, the HCV genomic plus-strand RNA was detected more frequently in noncancerous tissues (8 patients, 61.5%) and in sera (6 patients, 46.2%) than in cancerous tissues (3 patients, 23.1%). No viral RNA was found in either sera or tissues in the three patients without anti-HCV. By using a sense primer for the RT in the RT-PCR assay, amplification was attempted of a putative complementary minus-strand RNA that is believed to reflect viral replication in the eight patients with the plus-strand RNA. The minus-strand RNA was found in the noncancerous tissues of six patients; it was not detected in the cancerous tissues. CONCLUSIONS: These results suggest that HCV can infect and replicate predominantly in noncancerous cells but rarely in tumor cells.

Aged↗

PubMed1992

Camptothecin analog (CPT-11)-sensitive human pancreatic tumor cell line QGP-1N shows resistance to SN-38, an active metabolite of CPT-11.

In the course of our study to determine the cross-sensitivity between CPT-11 and its active metabolite, SN-38, we found a SN-38-resistant human pancreatic tumor cell line, QGP-1N, which shows sensitivity to CPT-11. The IC50 of SN-38 was 152 times greater for QGP-1N than for SUIT-2, also a human pancreatic tumor cell line, whose IC50 of CPT-11 was similar to that for QGP-1N. The uptakes of CPT-11 and SN-38 and the intracellular conversion of CPT-11 to SN-38 could not explain the difference in sensitivity. DNA synthesis of QGP-1N cells was inhibited by CPT-11 which did not affect that of SUIT-2, while SN-38 inhibited the DNA synthesis of SUIT-2 at lower concentrations than that of QGP-1N. The inhibition test of topoisomerase I catalytic activity by CPT-11 or SN-38 revealed no difference in the biochemical properties of the topoisomerase I enzymes to the compounds between these two cell lines. These results indicate that CPT-11 should have its own inhibitory effect on DNA synthesis through a yet unknown mechanism in QGP-1N cells, although SN-38 plays an essential role in the antitumor activity of CPT-11 in SUIT-2 cells. In some cases, the antitumor effect of CPT-11 might be consequent not only on SN-38 but also on CPT-11 itself.

Antineoplastic Agents, Phytogenic↗

PubMed1992

A possible regulatory role for conserved promoter motifs in an adult-specific muscle myosin gene from mouse.

The mouse gene encoding a myosin heavy chain (MHC) protein expressed in type IIB fibers of adult skeletal muscle has been cloned and its promoter isolated. A number of DNA sequence motifs are found within the first 2.5 kilobases of the promoter which are similar to motifs present in the promoters of other muscle-specific genes. One sequence located at approximately -940 base pairs corresponds to the motif called MEF1 which has been shown in other muscle genes to bind the myogenic regulatory factors of which MyoD is one example. The MEF1 site of this adult IIB MHC promoter does indeed bind MyoD although this factor is normally thought to be involved in early muscle cell differentiation. The IIB MHC promoter also has several motifs located in the first 200 base pairs which are strikingly conserved between this mouse gene and several chicken skeletal MHC genes. Of these evolutionarily conserved sequences, two motifs rich in A and T residues appear to be major contributors to the muscle-specific transcriptional activity of the mouse IIB MHC promoter when transfected into quail myogenic and non-myogenic cells. These observations suggest an important functional role for these AT-rich sequence motifs in the regulation of genes of the MHC family.

Animals↗

PubMed1992

Functional conversion of the homologous proteins alpha-lactalbumin and lysozyme by exon exchange.

Exons of eukaryotic genes that encode proteins frequently appear to encode structural and/or functional protein units [Gilbert, W. (1978) Nature (London) 271, 501; Blake, C.C.F. (1979) Nature (London) 277, 598]. alpha-Lactalbumin and c-type lysozyme are functionally quite different but structurally highly homologous proteins. Their gene organizations have been shown to be virtually the same and their exon structures are identical. The exon 2 region of hen lysozyme contains most of the amino acid residues that make up its catalytic cleft. In this study, we engineered a hybrid protein in which the exon 2 region of goat alpha-lactalbumin was replaced with that of hen lysozyme. This conferred catalytic activity on the alpha-lactalbumin, which is a nonenzymatic protein in its native structural form.

Animals↗

PubMed1992

RAG-2 expression is not essential for chicken immunoglobulin gene conversion.

Chicken B cells diversify their immunoglobulin genes by gene conversion in the bursa of Fabricius. The avian leukosis virus-induced B-cell line DT40 continues to diversify its immunoglobulin light chain locus by gene conversion during in vitro passage. Since a variable(diversity)joining recombination-activating gene, RAG-2, is specifically expressed in chicken B cells undergoing immunoglobulin gene conversion, it has been suggested that RAG-2 may be involved in the immunoglobulin gene conversion process. We previously reported high ratios of targeted to random integration after transfection of genomic DNA constructs into DT40. This allows us to easily investigate the function of a gene product by gene disruption. We show here that subclones of DT40 maintain the ability to diversify their immunoglobulin light chain locus by gene conversion even after both copies of the RAG-2 coding regions are deleted. These results demonstrate that the RAG-2 product is not required for gene conversion activity in the immunoglobulin light chain locus.

Animals↗

PubMed1992

Differential expression of DNA topoisomerase I gene between CPT-11 acquired- and native-resistant human pancreatic tumor cell lines: detected by RNA/PCR-based quantitation assay.

RNA/PCR quantitation method was developed to determine DNA Topoisomerase I(Topo I)-specific mRNA in order to study its gene expression in CPT-11 sensitive, acquired- or native-resistant human pancreatic tumor cell lines. The results were supported by Northern blotting and Western blotting analyses. Acquired-resistant cells have shown decreased levels of Topo I mRNA, compared with their parental cells. On the contrary, in the wild type cells no correlation was shown between sensitivity and gene expression. On the other, specific Topo I activity of the native resistant cell lines was fairly lower than that of sensitive cell lines, suggesting that immunoreactive Topo I protein contains low levels of active form enzyme which could be targets of CPT-11 in these native-resistant ones. Finally, the different mechanisms might be operative between acquired- and native-resistant tumor cells.

Antineoplastic Agents, Phytogenic↗

PubMed1992

Determination of baicalin and baicalein in rat plasma by high-performance liquid chromatography with electrochemical detection.

A rapid and sensitive method, using electrochemical detection, has been developed for the determination of baicalin and baicalein, the flavonoid of Scutellariae radix, in rat plasma. Following separation by high-performance liquid chromatography, baicalin and baicalein were oxidized at a glassy carbon electrode to permit selective electrochemical detection. Absolute detection limits were found to be 5 ng/ml from 50 microliters of plasma for baicalin and 2 ng/ml from 100 microliters of plasma for baicalein. The resulting assays were suitable for pharmacokinetic studies of baicalin and baicalein in rats.

Animals↗

PubMed1992

Redesign of the substrate-binding site of hen egg white lysozyme based on the molecular evolution of C-type lysozymes.

On the basis of the molecular evolution of hen egg white, human, and turkey lysozymes, three replacements (Trp62 with Tyr, Asn37 with Gly, and Asp101 with Gly) were introduced into the active-site cleft of hen egg white lysozyme by site-directed mutagenesis. The replacement of Trp62 with Tyr led to enhanced bacteriolytic activity at pH 6.2 and a lower binding constant for chitotriose. The fluorescence spectral properties of this mutant hen egg white lysozyme were found to be similar to those of human lysozyme, which contains Tyr at position 62. The replacement of Asn37 with Gly had little effect on the enzymatic activity and binding constant for chitotriose. However, the combination of Asn37----Gly (N37G) replacement with Asp101----Gly (D101G) and Trp62----Tyr (W62Y) conversions enhanced bacteriolytic activity much more than each single mutation and restored hydrolytic activity toward glycol chitin. Consequently, the mutant lysozyme containing triple replacements (N37G, W62Y, and D101G) showed about 3-fold higher bacteriolytic activity than the wild-type hen lysozyme at pH 6.2, which is close to the optimum pH of the wild-type enzyme.

Acetylglucosamine↗

PubMed1992

Antihepatitis C virus status in hepatocellular carcinoma and the influence on clinicopathological findings and operative results.

Antihepatitis C virus (HCV) status was investigated in 100 patients undergoing hepatectomy for hepatocellular carcinoma (HCC) between 1980 and 1989. The clinicopathological findings and operative results, in patients with or without HCV marker, were compared retrospectively. The positivity rate of anti-HCV was 51 per cent. In this group there was a higher mean age, fewer symptoms, raised alanine aminotransferase level, higher 15-min indocyanine green clearance rate and earlier tumour stage compared with the anti-HCV negative group. Positive tumour margins and vascular invasion were seen less frequently in the anti-HCV positive group. HCC with HCV marker showed characteristic features of chronic non-A non-B hepatitis and of HCC originating from liver cirrhosis. There was a better cumulative 1-year survival rate for anti-HCV positive patients, but 3- and 5-year survival rates after hepatectomy were similar in both groups. Although HCV-related HCC had typical features of chronic non-A non-B hepatitis and a relatively early stage of tumour, biological features and operative results were similar with or without the HCV marker.

Adult↗

PubMed1992

Age variation in the upper limit of hearing.

The upper limit of hearing was measured in 6105 otologically normal ears of subjects ranging in age from 5 to 89 years. The results are as follows: in each age group from 5 to 59 years in both sexes, the upper limit of hearing showed an approximately normal distribution if a logarithmic scale was used for the upper limit of hearing axis. The mode of the distribution shifted to a lower frequency with increasing age. Over age 60 years, the distribution became much wider. Standard upper limit age curves were established by calculating 10th, 25th, 50th, 75th and 90th percentiles for each age group. From early childhood where no age variation was recognized in conventional audiometry, deterioration of the upper limit of hearing was already in progress. This deterioration was slight between ages 25 and 39 but at ages over 40 it was accelerated and led to so-called presbycousis. The upper limit of hearing was found to be one of the best parameters for showing the quantitative age-related changes in hearing.

Adolescent↗

PubMed1992

Bethanechol and a G-protein activator, NaF/AlCl3, induce secretory response in Paneth cells of mouse intestine.

Paneth cells located at the bottom of intestinal crypts may play a role in controlling the bacterial milieu of the intestine. Using morphometry to clarify the secretory mechanism of the Paneth cells, we studied the ultrastructural changes in mouse Paneth cells produced following intra-arterial perfusion with Hanks' balanced salt solution containing a cholinergic muscarinic secretagogue (bethanechol), a neuroblocking agent (tetrodotoxin), or a G-protein activator (NAF/AlCl3). Bethanechol (2 x 10(-4) mol/l) induced Paneth-cell secretion. Many Paneth cells massively exocytosed their secretory material into the crypt lumen; the enhanced secretion caused degranulation and vacuole formation. However, tetrodotoxin (2 x 10(-6) mol/l) did not prevent the bethanechol-enhanced secretion by the Paneth cells. NaF (1 x 10(-2) mol/l) and AlCl3 (1 x 10(-5) mol/l) induced massive exocytosis of the Paneth cells; the exocytotic figures were similar to those observed in mice stimulated by bethanechol. G-protein activation was followed by a sequence of intracellular events, resulting in exocytosis.

Aluminum↗

PubMed1992

Symptoms of construction workers exposed to whole body vibration and local vibration.

To study the effects of construction machinery operation on subjective symptoms, a questionnaire survey was carried out among construction machinery operators by a self-reporting method. Subjects were 184 power shovel operators, 127 bulldozer operators, 44 forklift operators as operator groups, and 44 office workers as a control. Their ages were in a range of 30-49 years. The questionnaire contained 20 symptoms referring to fatigue, digestive problems, and upper or lower limbs or back problems. The prevalence rate and symptom characteristics were examined. The dominant symptoms of the operator groups were stiff shoulder, low back pain, and stomach symptoms. The prevalence rate of low back pain was significantly different between forklift operators and controls. No significant differences were found in the symptoms of upper limbs and fingers between operator groups and controls. The prevalence of Raynaud's phenomenon was 0.5%-2.3% in the operator groups and 2.3% in the control group.

Adult↗

PubMed1992

Potentiation of the antitumor activity of 5-trifluoromethyl-2'-deoxyuridine by the use of depot forms of the parent compound.

5-Trifluoromethyl-2'-deoxyuridine (CF3dUrd), an antitumor agent, is known to be short-lived in human plasma. Since its rapid elimination from the bloodstream seems to have descouraged the clinical evaluation of this drug, we explored the potential use of masked derivatives of CF3dUrd as "depot" forms of the parent compound. First, we observed that the toxicity of CF3dUrd against HeLA cells in culture was 10(4) times greater for a 24-h treatment as compared with a 1-h treatment at identical concentrations of the drug, which suggests the importance of using a prolonged treatment period. In fact, the divided dosing of CF3dUrd to L1210-bearing mice was markedly more effective than its single administration. 5'-O-Hexanoyl-, N3-p-butylbenzoyl-, 5'-O-benzyloxy-methyl-, and 3'-O-benzyl-CF3dUrd were found to be effective in maintaining the CF3dUrd concentration in plasma. The oral doses of these agents required to achieve 50% growth inhibition (ED50) in mice bearing sarcoma 180 tumors were 19, 34, 10, and 13 mg kg-1 day-1, respectively, whereas that of CF3dUrd was 63 mg kg-1 day-1. The ED50 values for these compounds were inversely correlated with the residence time of CF3dUrd in plasma. The therapeutic indices of these compounds, calculated as the dose producing a 50% inhibition of body-weight gain (IB50) divided by the ED50 value (1.89, 1.21, 1.40, and 2.15, respectively), were significantly higher than that of CF3dUrd (0.78). Consequently, these depot forms of CF3dUrd, particularly 3'-O-benzyl-CF3dUrd, are expected to be more useful than the parent compound as antitumor agents.

Animals↗

PubMed1992

A rapid and sensitive method for HLA-DRB1 typing by acridinium-ester-labeled DNA probes.

A rapid and accurate detection of HLA class II antigen is essential for transplantation and for the understanding of disease susceptibility. Recent molecular genetic studies have revealed that the number of class II loci and the number of alleles at these loci are greater than had been previously detected. It is, therefore, of great importance to detect these extensive polymorphisms. A great deal of effort has been made on identification of individual HLA class II specificities at the DNA level, called "DNA typing." What seems to be lacking, however, is handling simplicity. Here we accomplished a simple method for HLA-DRB1 typing based on hybridization of acridinium-ester (AE)-labeled DNA probes to amplified DNA. This method is called hybridization protection assay (HPA). By using 13 AE-labeled probes, 20 homozygous B-cell lines and leukocytes from 80 healthy individuals were typed by HPA. The results were completely consistent with those obtained by polymerase chain reaction--restriction fragment length polymorphism. This method is suitable for mass screening because of its procedural simplicity and swiftness.

Acridines↗

PubMed1992

Interaction of DP-TAT-59, an active metabolite of new triphenylethylene-derivative (TAT-59), with estrogen receptors.

DP-TAT-59, (Z)-2-(4-(1-(4-hydroxyphenyl)-2-(4-isopropylphenyl)-1-butenyl) phenoxy)-N, N-dimethylethylamine, has been reported to inhibit estrogen-stimulated growth of MCF-7 cells as well as rat uterus at lower concentrations than the hydroxymetabolite of tamoxifen (4-OH-TAM). In the present study, the growth of mouse Leydig cell tumor, B-1F cells were also more effectively inhibited by DP-TAT-59 than 4-OH-TAM. Additionally, the expression of estrogen responsive element ligated CAT gene transfected into B-1F cells was also suppressed by DP-TAT-59. Thus, the interaction of DP-TAT-59 with estrogen receptor (ER) was characterized and compared with that of 4-OH-TAM using immature rat and bovine uteri. The dissociation constant of DP-TAT-59 to ER of immature rat uterus was 0.24 nM and was similar to that of 4-OH-TAM (Kd = 0.20 nM) and estradiol (Kd = 0.29 nM). Using sucrose density gradients, the sedimentation constant of DP-TAT-59 with bovine uterus was 4.9S, which was similar to that of estradiol (5.1S) and 4-OH-TAM (5.3S). However, the elution profile of the DP-TAT-59-ER complex from a DEAE-Sephadex column was different for both estradiol-and 4-OH-TAM-ER complexes. These results suggest that ER forms different complexes with DP-TAT-59 than estradiol or 4-OH-TAM, while the ER binding affinity of these compounds are similar to each other.

Animals↗

PubMed1992

Isozymes of cyclic-3',5'-nucleotide phosphodiesterases in renal epithelial LLC-PK1 cells.

Metabolism of cAMP and cGMP by the major types (families) of cyclic-3',5'-nucleotide phosphodiesterases (PDE) was studied in confluent renal epithelial LLC-PK1 cells grown in vitro. LLC-PK1 cells mainly contain the cAMP-specific rolipram-sensitive PDE type-IV (PDE-IV), the Ca(2+)-calmodulin dependent PDE type-I and cGMP-specific PDE type-V; all these PDEs are mainly localized in cytosol. Analysis of PDE activities in soluble extract of LLC-PK1 cell homogenate by FPLC ionex chromatography on Mono-Q column also disclosed the presence of low activities of cGMP-stimulated PDE-II and PDE-III. Moreover, activity of PDE-IV was resolved into four distinct chromatographic peaks. The increase of cAMP level in response to incubation of intact LLC-PK1 cells with vasopressin (AVP) was markedly enhanced in the presence of rolipram, but not in the presence of other PDE isozyme-specific inhibitors. Incubation with AVP and atriopeptin (ANP) together resulted in increase in cGMP and a small decrease of cAMP accumulation in LLC-PK1 cells. Results of these studies first show that the LLC-PK1 cells contain all five major types of PDE isozymes where PDE-IV, PDE-I and PDE-V are quantitatively predominant. The rolipram-sensitive PDE-IV, present in several chromatographically distinct forms, appears to be the key PDE isozyme involved in control of cAMP generated in response to stimulation by AVP in LLC-PK1 cells.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1992

Effect of Gomisin A (TJN-101) on liver regeneration.

We studied the effect of TJN-101, a lignan component of Schisandra fruits (Schisandrae fructus), on liver regeneration after partial hepatectomy. TJN-101 was given orally to male Wistar rats 30 min before partial hepatectomy. The mitotic index and the level of DNA synthesis increased after partial hepatectomy and their increase was significantly enhanced by TJN-101. Ornithine decarboxylase (ODC) activity increased in the early stages of liver regeneration and it was also significantly enhanced by TJN-101. Besides, TJN-101 enhanced the increase in hepatic putrescine. These results suggest that TJN-101 stimulates liver regeneration after partial hepatectomy by enhancing ODC activity, which is an important biochemical event in the early stages of liver regeneration.

Animals↗

PubMed1992

A protease inhibitor produced by Streptomyces lividans 66 exhibits inhibitory activities toward both subtilisin BPN' and trypsin.

A proteinaceous protease inhibitor was isolated from the culture broth of Streptomyces lividans 66 by a series of purification steps (salting out by ammonium sulfate, ion-exchange chromatography on DEAE-cellulose, hydrophobic chromatography on Phenyl-Sepharose, and gel-filtration on Sephacryl S-200), and was named S. lividans protease inhibitor (SLPI). The purified SLPI existed in a dimeric form consisting of two identical subunits, each of which was composed of 107 amino acids. SLPI exhibited strong inhibitory activity toward subtilisin BPN'. These features were similar to those of protein protease inhibitors produced by other Streptomyces (SSI family inhibitor). In addition, SLPI was capable of inhibiting trypsin with an inhibitor constant (Ki) of about 10(-9) M. The primary structure of SLPI and location of two disulfide bridges were homologous to those of the other serine protease inhibitors of Streptomyces. The reactive site of SLPI was found to be Arg67-Glu68 from the sequence analysis of cleaved SLPI which was produced by acidification of subtilisin-SLPI complex. An Arg residue at the P1 site was consistent with the trypsin-inhibitory property of SLPI. Sequence comparison with other members of the SSI family revealed that amino acid replacements in SLPI were mainly localized on the surface of the SLPI molecule, and many of the amino acid residues in beta-sheets and hydrophobic core were well conserved.

Amino Acid Sequence↗