Biomedical subjects

H Toyoda

Publications and source records attributed to H Toyoda.

At least 163 records · Page 9Linked to original sources

PubMed1996

Simultaneous determination of dermatan sulfate and oversulfated dermatan sulfate in plasma by high-performance liquid chromatography with postcolumn fluorescence derivatization.

A chemical method for the determination of dermatan sulfate (DS) and oversulfated dermatan sulfate has been developed and applied to the pharmacokinetic study of these polysaccharides in experimental animals. The analytical procedure includes a simple preparation step of administered DS and oversulfated DS from blood plasma, HPLC for the separation and detection of DS and oversulfated DS using an Asahipak NH2P-50 column, fluorometric reaction of the polysaccharides with guanidine in a strong alkaline medium. DS and oversulfated DS were extracted from plasma by treating it with proteinase to remove plasma proteins and recovered with endogenous plasma glycosaminoglycans by ethanol precipitation. Finally, DS and oversulfated DS were analyzed by fluorometric HPLC. The detection limits of DS and oversulfated DS were 10 and 20 ng, respectively. Furthermore, we demonstrated that artificial oversulfation of DS increased its biological half-life after intravenous administration to rats.

Animals↗

PubMed1996

Nucleotide sequence diversity of hypervariable region 1 of hepatitis C virus in Japanese hemophiliacs with chronic hepatitis C and patients with chronic posttransfusion hepatitis C.

Hemophiliac patients with chronic hepatitis C might be exposed to and become infected with multiple hepatitis C virus (HCV) strains by means of frequent use of blood products, even if they are infected with a single subtype of HCV. To test this hypothesis, we analyzed the genetic diversity of hypervariable region 1 (HVR1) of HCV in chronically infected hemophiliacs and in patients with chronic posttransfusion hepatitis with a single HCV inoculation. The diversity of nucleotide sequences in HVR1 of serum HCV RNA was compared between 21 hemophiliacs infected with a single HCV subtype and 16 patients with posttransfusion HCV infection. The number of HCV quasispecies was determined by fluorescence single-strand conformation polymorphism (SSCP) analysis. Direct sequencing was performed to determine the diversity in HVR1. The number of HCV quasispecies in the blood was 5.2 +/- 2.0 clones in hemophiliacs and 4.0 +/- 2.3 clones in posttransfusion patients, a nonsignificant difference (P = .0943). The number of sites at which the nucleotide was not homogenous in all quasispecies was significantly higher in hemophiliacs (13.0% +/- 7.4%) than in posttransfusion hepatitis patients (2.7% +/- 2.8%; P < .0001). In conclusion, there was a high degree of genetic variation in HVR1 of HCV specimens isolated from hemophiliacs compared with posttransfusion patients. These findings indicate the possibility that multiple infections of a single HCV subtype may occur among patients frequently exposed to blood products; single HCV subtypes may therefore derive from multiple origins.

Adult↗

PubMed1996

Methylation status of 5'-regulatory region of tumor necrosis factor alpha gene correlates with differentiation stages of monocytes.

The DNA methylation status of the 5'-regulatory region of human tumor necrosis factor-alpha (TNF-alpha) gene was examined using two human monocytic cell lines representing different differentiation stages: one cell line, THP-1, represents the most differentiated macrophage-like phenotypes and the other, HL-60, represents a least differentiated phenotype. Two restriction enzymes were used to detect methylated sites in the 5'-regulatory region of the TNF-alpha gene. The restriction enzyme Msp I, which recognizes both CCGG (unmethylated) and C(me)CGG (methylated) sequences, whereas Hpa II recognizes only the unmethylated CCGG sequence. Two Msp I sites located at -600 and +200 nucleotides, respectively, from the transcriptional initiation site were unmethylated in DNA from the THP-1, whereas these sites were methylated in the one from the HL-60. Treating the HL-60 cells with 1,25-vitamin D3, ( 1,25-(OH)2D3), which is known to induce monocytic cell differentiation, induce demethylation of the sites, suggesting the methylation status was correlated with the differentiation stages of monocytes.

Base Sequence↗

PubMed1996

High-performance liquid chromatographic analysis of glycosaminoglycan-derived oligosaccharides.

High-performance liquid chromatography of glycosaminoglycan (GAG)-derived oligosaccharides has been employed for the structural analysis and measurement of hyaluronan, chondroitin sulphate, dermatan sulphate, keratan sulphate, heparan sulphate and heparin. Recent developments in the separation and detection of unsaturated disaccharides and oligosaccharides derived from GAGs by enzymatic or chemical degradation are reviewed.

Carbohydrate Sequence↗

PubMed1996

Comparison of serum hepatitis C virus RNA concentration by branched DNA probe assay with competitive reverse transcription polymerase chain reaction as a predictor of response to interferon-alpha therapy in chronic hepatitis C patients.

A study was carried out to assess the correlation between the serum concentration of hepatitis C virus RNA (HCV-RNA) in patients with chronic hepatitis, as measured by competitive reverse transcription polymerase chain reaction (cRT-PCR) and branched DNA probe assay (bDNA), and response to interferon-alpha (IFN alpha) therapy. The serum HCV-RNA concentration was evaluated by both cRT-PCR and bDNA in 54 patients who had received a total dose of 480 MU of IFN alpha. HCV subtypes were also identified in all patients. The measurement of serum HCV-RNA concentration by bDNA correlated significantly with that of cRT-PCR. The concentration of HCV-RNA in subtype 1 patients was significantly higher than that in subtype 2 patients when measured by bDNA, but not when measured by cRT-PCR. The correlation of HCV-RNA concentration between bDNA and cRT-PCR was associated with both subtypes 1 and 2. The difference in serum HCV-RNA concentration between complete and incomplete responders was more significant when measured by bDNA probe assay than by cRT-PCR. Moreover, only 1 of 26 patients with a HCV-RNA concentration of more than 1 x 10(6) eq/ml as measured by bDNA probe assay attained a complete response, while 19 of 28 patients with that of less than 1 x 10(6) eq/ml achieved it. Measurement of serum HCV-RNA concentration by bDNA probe assay was a better predictor of clinical response of IFN alpha therapy than measurement by cRT-PCR.

Adult↗

PubMed1996

Tumor necrosis factor microsatellites define a Crohn's disease-associated haplotype on chromosome 6.

BACKGROUND & AIMS: HLA class II associations have been described in Crohn's disease (CD) and ulcerative colitis (UC) and may be markers for other closely linked genes that are involved in disease pathogenesis. The tumor necrosis factor (TNF) locus, which contains the genes for TNF-alpha and TNF-beta, is located on chromosome 6 within the major histocompatibility complex loci. To investigate potential genetic associations in inflammatory bowel disease at the TNF locus, we studied 75 patients with CD, 73 patients with UC, and 60 ethnically matched controls using microsatellite markers. METHODS: Five TNF microsatellite loci (TNFa, TNFb, TNFc, TNFd, and TNFe) were typed using polymerase chain reaction. RESULTS: A CD-associated allelic combination, TNFa2b1c2d4e1, was found in 24% of patients with CD, 4.1% of patients with UC (P=0.001; odds ratio, 7.4; CD vs. UC), and 6.7% of control subjects (P=0.01; odds ratio, 4.4 CD vs. controls). This TNF haplotype was associated with the previously described HLA-DR1/DQ5 combination in CD. CONCLUSIONS: The TNFa2b1c2d4e1 allelic combination is the strongest genetic risk factor described in CD and, with HLA class II alleles, defines a group of markers on chromosome 6 that extends from HLA class II to upstream of the TNF-beta gene.

Alleles↗

PubMed1996

Long-term administration of natural interferon-alpha in patients with chronic hepatitis C: relationship to serum RNA concentration, HCV-RNA genotypes, histological changes and hepatitis C virus.

To virologically assess the efficacy of interferon therapy in chronic hepatitis C, either 5 or 10 MU/day natural interferon-alpha (IFN alpha) was administered to 57 patients with chronic hepatitis C for 38 weeks. A complete and sustained response (CR-SR), as evidenced by the absence of serum hepatitis C virus (HCV)-RNA during the administration period and at 6 months after the final administration of IFN alpha and normal GPT level at 6 months after final administration, occurred in 42.6% (23/54) of subjects. Liver tissue was histologically evaluated using the histological activity index (HAI) score before and after the administration period. In CR-SR cases, significant improvements (P < 0.01) occurred in periportal necrosis, intralobular necrosis, portal inflammation and total score. A comparison, by HCV genotypes, revealed that CR-SR occurred in 60% (9/15) of subjects with type 2a and 30.3% (10/33) of subjects with type 1b. A comparison by virus concentration revealed that CR-SR occurred in 71.4% (15/21) of those subjects having a virus concentration of < 10(5) copies/mL, but in only 24.2% (8/33) of those having a virus concentration of > 10(5) copies/mL. Analysis by a multiple logistic model revealed a strong correlation between the therapeutic effect of interferon therapy and the pre-administration virus concentration (P = 0.0061) and genotype (P = 0.0015). These results suggest that the pre-administration virus concentration and genotype are both key factors affecting the therapeutic effect of interferon therapy in chronic hepatitis C and that the therapeutic effect of interferon is satisfactorily high, irrespective of virus concentration, in subjects with type 2a HCV, but varies depending on virus concentration in subjects with type 1b.

Aged↗

PubMed1996

Estimation of usefulness of N-butyl-2-cyanoacrylate-lipiodol mixture in transcatheter arterial embolization for urgent control of life-threatening massive bleeding from gastric or duodenal ulcer.

We estimated the usefulness of a mixture of N-butyl-2-cyanoacrylate (NBCA) with lipiodol for transcatheter arterial embolization (TAE) used to control massive bleeding from gastric or duodenal ulcer. Thirty patients who had gastric or duodenal ulcers and massive bleeding that was uncontrollable by endoscopic procedures were included in this study. All patients were subjected to TAE (without NBCA in 23 and with NBCA in seven patients). Coils and/or gelfoam were also used. The achievement of haemostasis, occurrence of rebleeding and the time taken for TAE were compared between patients who received TAE without and with NBCA. Eighteen of 23 patients (78.3%) who received TAE without NBCA and six of seven patients (85.7%) who underwent TAE with NBCA achieved complete haemostasis without rebleeding. The time for TAE was significantly shorter in patients who received NBCA compared with those who did not (P = 0.0095). TAE using NBCA or a combination of NBCA and coils achieved a rapid, complete embolization regardless of vascular distribution or arterial diameter. Thus NBCA is considered to be useful as a secondary embolization material in TAE that is urgently conducted to control massive bleeding from gastric or duodenal ulcers.

Adult↗

PubMed1996

Case report: multiple systemic lymph node metastases from a small hepatocellular carcinoma.

Extrahepatic lymph node metastases from hepatocellular carcinoma (HCC) are usually observed in patients with advanced and poorly differentiated HCC. We encountered a patient with multiple, systemic lymph node metastases from a small HCC (18 mm in diameter), which was nodular and had a capsule at the time of resection (a partial hepatectomy of the postero-inferior subsegment of the right lobe of the liver). Widespread lymphadenopathy resembling malignant lymphoma developed 2 months after surgery. A biopsy specimen from a supraclavicular lymph node revealed metastatic HCC. The patient died 2.5 months after the detection of the lymphadenopathy. Lymph node metastases can occur in small HCC less than 2 cm in diameter and may adversely affect the long-term prognosis of patients with these curatively resectable small HCC.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1996

Compositional analysis of hyaluronan, chondroitin sulfate and dermatan sulfate: HPLC of disaccharides produced from the glycosaminoglycans by solvolysis.

An ion-exchange high-performance liquid chromatography procedure was developed for analysis of mixtures of N-acetyldermosine, N-acetylchondrosine and N-acetylhyalobiuronic acid produced quantitatively by heating dermatan sulfate, chondroitin sulfate and hyaluronan in dimethyl sulfoxide (DMSO) containing 0.1% (v/v) H20 at 80 degrees C for 48 h. These disaccharides were eluted from a TSK gel SAX column using 0.1 M acetic acid containing KCl, and detected fluorometrically by post column derivatization. The eluate was mixed with 2-cyanoacetamide solution and alkaline solution, and heated at 110 degrees C for 4 min. The resultant compounds were detected fluorometrically (Ex. 335 nm and Em. 390 nm). The usefulness and practicality of the present method were verified by applications to the determination of glycosaminoglycans in tissues.

Animals↗

PubMed1996

Chitinous component of the cell wall of Fusarium oxysporum, its structure deduced from chitosanase digestion.

The cell of Fusarium oxysporum was digested with commercial Bacillus chitosanase. The chitosanase produced low molecular weight heterooligosaccharides consisting of GlcN and GlcNAc from the cell wall. A main component of the digestion products was identified as 2-amino-2-deoxy-beta-D-glucopyranosyl- (1-->4)-2-acetamido-2-deoxy-D-glucopyranose. The chitosanase appeared to be more effective than Streptomyces griseus chitinase for cell wall digestion. Moreover, maltose was unexpectedly found in the digestion products, indicating that the cell wall contains alpha-1,4-linked glucan chain as a polysaccharide component.

Acetylglucosamine↗

PubMed1996

Improvement of hemolysis in muscle phosphofructokinase deficiency by restriction of exercise.

A 29-year-old woman with muscle phosphofructokinase (PFK) deficiency had exercise intolerance, painful cramps, elevation of muscle enzyme levels in the serum and compensated hemolysis. After the restriction of exercise, the creatine kinase level and indirect bilirubin level decreased, and the reticulocyte count and haptoglobin level were normalized. It is suggested that the hemolysis which was accelerated by exercise was improved by restriction of exercise.

Adult↗

PubMed1996

Effect of 5-alpha dihydrotestosterone on T-cell proliferation of the female nonobese diabetic mouse.

Nonobese diabetic (NOD) mice develop type I diabetes spontaneously and have been utilized as a model for human autoimmune insulin-dependent diabetes. The disease is caused by the destruction of insulin-producing beta cells in the pancreatic islet of Langerhans by infiltrating inflammatory cells, which are primarily T lymphocytes. The incidence of diabetes in NOD mice is increased in females compared with males, suggesting that sex steroid hormones play an important role in the development of the disease. We therefore investigated the effect of a male steroid, 5-alpha-dihydrotestosterone (5DHT), on disease development, T-cell phenotype, T-cell proliferation, and cytokine profiles in this model. None of the mice that received 5DHT for 120 days (n = 7) developed insulitis, whereas all control mice (n = 8) developed the disease. The percentage of CD4+ T cells in peripheral blood mononuclear cells was markedly decreased in the 5DHT-treated females compared with those in controls (37.1 +/- 4.8 vs 51.3 +/- 9.3, P < 0.02), whereas no significant differences in the percentage of CD8+ T cells were observed between treated and control female mice. Results of a syngeneic mixed lymphocyte reaction (SMLR) also suggested that T cells are major target cells of 5DHT administration. An increased expression of IL-4 mRNA, representing T helper 2 (Th2) T cells, was observed in the SMLR. On the basis of these results, a systemic administration of 5DHT appears to have direct effects on the expansion of Th2 cell populations with subsequent restoration of normal immune responses.

Animals↗

PubMed1996

[The clinical evaluation of hepatic arterial infusion chemotherapy in patients with liver metastases from colorectal cancer employing an implanted port system].

Between 1986 and 1994, 66 patients with unresectable liver metastases from colorectal cancer were treated by repeated hepatic arterial infusion chemotherapy employing an implantable port system. In this study, 44 patients who received intermittent hepatic arterial infusion of high dose 5-FU (1000mg/m2) were analysed according to a response rate, survival rates, developments of extrahepatic lesions, periods of hepatic arterial infusion, and the rates at home. Two cases (4.5%) achieved a complete response (CR) and 27 cases (61.4%) a partial response (PR). The overall one-year and two-year survival rates were 56.7% and 37.8% respectively. The 50% survival time for all patients was 17.2 months. During the course of observation, extrahepatic lesions developed in 16 of the 39 patients (41.0%) and many responders died due to deterioration of extrahepatic lesions. The over all rates at home were more than 85%. Hepatic arterial infusion of the 5-FU at 1000mg/m2 every week showed a high response rate in colorectal cancer patients with hepatic metastases, and the responders showed a low rate of death due to the hepatic metastases. However, in many patients the prognosis-determining-factor was extrahepatic lesions. Thus, countermeasures were necessary for extrahepatic lesions.

Aged↗

PubMed1996

Effect of daily administration period of natural alpha-interferon in patients with chronic hepatitis C.

OBJECTIVE: A randomized, controlled trial was performed to study the effect of the period of daily administration of interferon therapy in patients with chronic hepatitis C. PATIENTS AND METHODS: Sixty-three patients were administered a total dose of 480 MU of natural alpha-interferon, 6 MU each day. In 32 cases, interferon was given daily for 2 wk, followed by intermittent administration (three times a week) for 22 wk. The other 31 patients received daily administration for 8 wk, followed by intermittent administration (twice a week) for 12 wk. The responses were based on the normalization of ALT and the clearance of hepatitis C virus. RESULTS: Although ALT normalized in 37 patients during the administration of interferon, it relapsed after the end of interferon therapy in 20 patients. The other 17 patients achieved sustained normalization of ALT and clearance of hepatitis C virus RNA. Responses did not differ according to the administration period of interferon. CONCLUSIONS: Prolonged daily administration did not improve the responses to interferon therapy when the total dose was the same.

Alanine Transaminase↗

PubMed1996

Differential association of protein Ser/Thr phosphatase types 1 and 2A with the cytoskeleton upon platelet activation.

The association of protein Ser/Thr phosphatase type 1(PP1) and type 2A (PP2A) with the cytoskeleton (Triton X-100 insoluble residue) during human platelet activation was investigated. In unstimulated platelets, 40% of total PP1-like activity was present in the Triton-insoluble cytoskeleton, while only 10% of the total PP2A-like activity was present in this fraction. Stimulation with 1 U/ml thrombin produced a 1.8-fold increase in PP1-like activity and a 7-fold increase in PP2A-like activity, respectively, in the cytoskeletal fraction, under aggregating conditions. Immunoblot analysis revealed that thrombin treatment increased association of PP1 catalytic subunit isozymes (PP1 alpha, PP1 gamma, PP1 delta) and PP2A catalytic subunit with the cytoskeleton, with concomitant decrease of these enzymes in Triton-soluble fractions. The amounts of cytoskeleton-associated PP1 and PP2A depended on the dose of thrombin which could activate platelets. Agonist-induced redistribution of PP1 and PP2A into the cytoskeleton was inhibited by OP-41483 (a prostaglandin I2 analog). Interaction of PP2A with cytoskeletal proteins strongly correlates with aggregation, whereas the association of PP1 with cytoskeleton can be detected upon platelet activation, even in the absence of aggregation. Co-extraction of protein kinase C and myosin light chain kinase with the cytoskeleton eventually translocated to the cytoskeleton, but only during aggregation. These results suggest that differential translocation of PP1 and PP2A to the cytoskeleton is involved in platelet activation, and their association with cytoskeletal proteins may regulate phosphorylation levels together with protein kinases in platelets.

Cytoskeleton↗

PubMed1995

Molecular cloning of mouse epiregulin, a novel epidermal growth factor-related protein, expressed in the early stage of development.

A cDNA clone encoding a novel epidermal growth factor (EGF)-related growth regulator, epiregulin, was isolated from a cDNA library prepared from a mouse fibroblast-derived tumor cell line, NIH3T3/clone T7. The predicted amino acid sequence revealed that the purified epiregulin peptide of 46-amino acids was synthesized as an internal segment of a 162-amino acid putative transmembrane precursor. The structural organization was similar to that of TGF-alpha precursor among the members of the EGF family. Although epiregulin transcript was not detected in several adult normal tissues by Northern blot analysis, approximately 4.8-kb transcript was present in 7-day-old mouse embryo and then diminished to very low or undetectable levels. Our results suggest that epiregulin may play an important role in the regulation of epithelial cell growth during early development.

3T3 Cells↗

PubMed1995

Epiregulin. A novel epidermal growth factor with mitogenic activity for rat primary hepatocytes.

Epiregulin, a novel epidermal growth factor (EGF)-related growth regulating peptide, was purified from conditioned medium of the mouse fibroblast-derived tumor cell line NIH3T3/clone T7. It was a 46-amino-acid single chain polypeptide, and its amino acid sequence exhibited 24-50% amino acid sequence identity with sequences of other EGF-related growth factors. Epiregulin exhibited bifunctional regulatory properties: it inhibited the growth of several epithelial tumor cells and stimulated the growth of fibroblasts and various other types of cells. Epiregulin bound to the EGF receptors of epidermoid carcinoma A431 cells much more weakly than did EGF, but was nevertheless much more potent than EGF as a mitogen for rat primary hepatocytes and Balb/c 3T3 A31 fibroblasts. These findings suggest that epiregulin plays important roles in regulating the growth of epithelial cells and fibroblasts by binding to receptors for EGF-related ligands.

3T3 Cells↗