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Biomedical subjects

K Furihata

Publications and source records attributed to K Furihata.

At least 55 records · Page 3Linked to original sources
PubMed1998

Structures and biosynthesis of aflastatins: novel inhibitors of aflatoxin production by Aspergillus parasiticus.

Two novel inhibitors of aflatoxin production by Aspergillus parasiticus were isolated from the mycelial extracts of Streptomyces sp. MRI142 and termed aflastatin A and B. The structures of aflastatin A (1) and B (5) were elucidated by NMR and chemical degradation experiments. These compounds have a novel skeleton of a tetramic acid derivative with a highly oxygenated long alkyl chain. The incorporation experiments using 13C-labeled acetates, propionate, glucose and glycolate suggested that most of the C2 and C3 units involved in the alkyl chain moiety of aflastatin A were biosynthesized from acetic and propionic acids, but five C2 units in the alkyl chain originated from glycolic acid.

Aflatoxins↗
PubMed1998

[Clinical evaluation of RT-PCR method for detection of HCV-RNA: second report availability of RT-PCR system using PCR internal control for detection of HCV-RNA and the influence of interfering substances on the detection].

The COBAS Amplicor system is an automated diagnostic PCR system which contains a PCR internal control (P. I. C.) template to monitor the amplification. The applicability of COBAS Amplicor HCV was examined using sera of patients with hepatitis. Furthermore, the effects of possible interfering substance (total protein, triglyceride, hemoglobin, glucose, total bilirubin, heparin, lysis reagent including guanidium) on HCV-RNA detection were investigated. The sensitivity of COBAS Amplicor HCV was equivalent to the manual method of Amplicor HCV, moreover all of the results in 54 clinical samples analyzed on both COBAS Amplicor HCV and Amplicor HCV were in agreement. Detection sensitivity of HCV-RNA decreased in the presence of total bilirubin and heparin. Ten and 25mg/dl of total bilirubin affected HCV-RNA detection but did not affect P. I. C. This result suggested that total bilirubin interfered with the protein denature caused by the lysis reagent. Fifteen U/ml of heparin in the sample completely inhibited amplification both of the HCV-RNA and P. I. C. One U/ml of heparin did not affect amplification, but heparinized blood samples should not be used for the detection of HCV-RNA. To examine the effect of possible carry over contamination on the lysis reagent which contains guanidium, various concentrations of lysis reagent in P. I. C. were tested. RT-PCR was inhibited by 1/500 volume contamination of lysis reagent in specimen diluent. Other substances did not affect the sensitivity. Our results indicate that the carryover contamination of lysis reagent cause more "false negative" results than interfering substances in sera. In conclusion, HCV-RNA detection system containing P. I. C., such as COBAS Amplicor HCV, will become a very useful to differentiate "false negative" and "true negative" result.

Bilirubin↗
PubMed1998

[Measurement of fibrinogen binding to platelets by flow cytometry: evaluation method for reflecting platelet activation].

Platelet aggregation, induced by agonist-mediated activation of membrane glycoprotein (GP) IIb/IIIa, and binding of fibrinogen to GPIIb/IIIa, is commonly analyzed using an aggregometer in the clinical laboratories. However, this method has a limitation to get precise results on the samples with small number of platelet (less than 100,000/1) or hyperlipidemia. Recently, flow cytometry has been used to evaluate platelet function due to the detection of fibrinogen binding to activated platelets using fluorescence labeled fibrinogen or anti-fibrinogen antibody. However, the appropriate rule for evaluation of the results has not been established yet. We converted a ratio of fibrinogen binding platelets to a velocity per unit concentration of ADP as follows: a difference of two ratios of fibrinogen binding platelets on neighboring two ADP concentrations was divided by a difference of ADP concentrations. It was considered to be a mean velocity between the two ADP concentrations. We adopted the range of ADP concentration, which gave the maximum velocity, as an index of platelet activation. If the peak of maximum velocity move toward lower or higher ADP concentration, it means hyper- or hypoactivation of the platelets, respectively. The objectivity of this method may make it a useful technique for clinical examination of platelet function.

Adenosine Diphosphate↗
PubMed1998

[Evaluation of enzyme-linked immunosorbent assay for antinuclear antibodies].

Antinuclear antibodies (ANAs) are clinically important indicators of collagen diseases. As corresponding antigens for ANAs vary considerably, patients with collagen diseases usually demonstrate several ANAs coincidentally, making difficult to detect the full spectrum of ANAs in each patient's serum. To design an efficient system for measuring ANAs, an enzyme-linked immunosorbent assay (ELISA) which adsorbs eight kinds of recombinant or purified antigens in each well of a multiwell plate was used and results were compared to those obtained with conventional assays by the fluorescent antinuclear antibodies (FANA), and double immunodiffusion (DID) methods. The positivity rates of 106 sera from patients with collagen diseases and 286 sera from healthy subjects were 92.5% and 5.5%, respectively. Sixty-one of 65 positive sera (93.8%) in the corresponding ANAs positive sera by DID or other conventional assay methods were positive by ELISA. Anti-SSA/Ro antibody could be detected with higher sensitivity by this assay method than with the FANA and DID method, but the sensitivities for anti-Scl-70 antibody and anti-centromere antibody were lower. Application of this ELISA method for measuring ANAs along with the FANA test may be beneficial for diagnosis of collagen diseases.

Adult↗
PubMed1997

Detection of prostate cancer cells circulating in peripheral blood by reverse transcription-PCR for hKLK2.

Two of the human tissue kallikrein family, hK2 and hK3 (prostate-specific antigen), are primarily produced by the prostatic epithelium under the regulation of androgens. In this study, we detected prostate cancer cells that expressed hKLK2 or hKLK3 mRNA in the peripheral blood of patients with prostate cancer using reverse transcription-PCR (RT-PCR). We then demonstrated some differences in characteristics, such as differentiation of cancer cells and response to antiandrogen therapy, between hKLK2 and hKLK3 mRNA-expressing prostate cancer cells. Total RNA was isolated from 41 patients with known prostate cancer, 7 patients with benign prostatic hyperplasia, and 20 normal volunteers. By RT-PCR, hKLK2 mRNA was detected in 7 patients (33%), and hKLK3 mRNA was detected in 17 (81%) of 21 stage D prostate cancer patients. In contrast, all patients with benign prostatic hyperplasia and healthy volunteers were negative. From comparison of the background of the patients positive for hKLK2 and/or hKLK3 mRNA, it became evident that the response to antiandrogen therapy and the expression of hKLK2 mRNA were reciprocally correlated, in contrast with the expression of hKLK3 mRNA. Additionally, our study clearly demonstrated that the detection of hKLK2 mRNA in the peripheral blood was useful for screening patients with certain prostate cancers that did not express hK3. We conclude that taking advantage of the difference between hKLK2 mRNA and hKLK3 mRNA expression is clinically useful for following up prostate cancer patients.

Adenocarcinoma↗
PubMed1997

Arotinoid mofarotene (RO40-8757) up-regulates p21 and p27 during growth inhibition of pancreatic cancer cell lines.

Effective chemotherapy for pancreatic cancer is urgently needed. The anti-proliferative activity of a new retinoid, mofarotene (RO40-8757), was compared with that of other retinoids, such as all trans-retinoic acid, 13-cis retinoic acid and 9-cis retinoic acid, on 9 pancreatic cancer cell lines in relation to the effects on various cell cycle-regulating factors. After treatment with each retinoid, anti-proliferative effect was determined by the MTT method and expression of cell cycle-regulating factors, such as cyclins (D1, E and A), cyclin-dependent kinases (2 and 4), cyclin-dependent kinase inhibitors (p21 and p27) and retinoblastoma protein, was analyzed by Western blotting. Mofarotene showed half-maximal inhibition of cell proliferation at concentrations between 0.14 x 10(-6) and 3.8 x 10(-6) mol/l with little cytotoxicity. In contrast, the other retinoids did not inhibit the growth of all cell lines by over 50% compared to controls. A marked increase in the fraction of cells in G1 phase of the cell cycle was observed after mofarotene treatment; this was associated with marked up-regulation of p21/p27 and a shift of retinoblastoma protein into the hypophosphorylated form. In conclusion, mofarotene inhibits the growth of pancreatic cancer cells by inducing G1-phase cell cycle-inhibitory factors (p21, p27 and hypophosphorylated form of Rb protein) and is considered to be a useful agent for pancreatic cancer treatment.

Alitretinoin↗
PubMed1997

Germline mutations of the APC gene in two Japanese adenomatous polyposis patients.

Germline mutations of the adenomatous polyposis coli (APC) gene have been reported in patients with familial adenomatous polyposis (FAP) and are believed to be an early event in colorectal carcinoma. We report the results of screening for germline mutations of the APC gene in 4 cases of 2 kindreds using non-radioactive PCR-SSCP (polymerase chain reaction-single strand conformation polymorphism) analysis. The mutation in kindred 1 was a 4 bp deletion at codon 849 in exon 15, resulting in a frameshift leading to truncation of the APC gene product. In kindred 2, a transversion of C to G at codon 2038 was observed, resulting in an amino acid change from leucine to valine. In this case, it is possible to screen presymptomatic diagnosis easily and quickly by digestion with restriction enzyme EcoNI.

Adenomatous Polyposis Coli↗
PubMed1997

Vitamin D analogues up-regulate p21 and p27 during growth inhibition of pancreatic cancer cell lines.

To obtain information regarding the growth-inhibitory effect of 1,25-dihydroxyvitamin D3 and its non-calcaemic analogue 22-oxa-1,25-dihydroxyvitamin D3 on pancreatic cancer cell lines, differences in the effects of G1-phase cell cycle-regulating factors were studied in vitamin D-responsive and non-responsive cell lines. Levels of expression of cyclins (D1, E and A), cyclin-dependent kinases (2 and 4) and cyclin-dependent kinase inhibitors (p21 and p27) were analysed by Western blotting after treatment with these compounds. In the responsive cells (BxPC-3, Hs 700T and SUP-1), our observations were: (1) marked up-regulation of p21 and p27 after 24 h treatment with 10(-7) mol l(-1) 1,25-dihydroxyvitamin D3 and 22-oxa-1,25-dihydroxyvitamin D3; and (2) marked down-regulation of cyclins, cyclin-dependent kinases and cyclin-dependent kinase inhibitors after 7 days' treatment. In non-responsive cells (Hs 766T and Capan-1), no such changes were observed. In conclusion, vitamin D analogues up-regulate p21 and p27 as an early event, which in turn could block the G1/S transition and induce growth inhibition in responsive cells.

Antineoplastic Agents↗
PubMed1997

Long-term follow-up of patients with chronic pancreatitis and K-ras gene mutation detected in pancreatic juice.

BACKGROUND & AIMS: Conflicting reports have raised the question whether K-ras gene mutations in chronic pancreatitis are related to the development of pancreatic neoplasm. The aim of the present study was to clarify this issue by surveying patients with chronic pancreatitis and K-ras gene mutation over 4 years. METHODS: K-ras point mutations at codon 12 in duodenal juices obtained during exocrine function tests were examined by enriched polymerase chain reaction-restriction fragment length polymorphism and direct DNA sequence analysis, and long-term follow-up of these patients was performed. RESULTS: K-ras gene mutation was found in 20 of 54 patients (37%) with chronic pancreatitis. The types of mutation were GAT in 7, GTT in 11, and TGT in 2 patients. Pancreatic neoplasm occurred in none of the mutation-positive patients over a mean follow-up period of 78 months. CONCLUSIONS: K-ras gene mutation in patients with chronic pencreatitis did not seem to be related to the development of pancreatic neoplasm during the follow-up period, and analysis of K-ras gene mutation seems of little use for diagnosing pancreatic neoplasm in patients with chronic pancreatitis.

Adult↗
PubMed1997

Trinucleotide repeat polymorphism within exon 5 of the MICA gene (MHC class I chain-related gene A): allele frequency data in the nine population groups Japanese, Northern Han, Hui, Uygur, Kazakhstan, Iranian, Saudi Arabian, Greek and Italian.

We recently identified a trinucleotide repeat polymorphism, (GCT)n, within the transmembrane (TM) segment of the human MHC class I MICA gene (MHC class I chain-related gene A). Five distinct alleles (A4, A5, A5.1, A6, A9) corresponding to 4, 5, 5 with one nucleotide insertion, 6 and 9 repetitions, respectively, have been detected in various HLA-homozygous B cell lines. Here we present allele frequencies for this trimeric short tandem repeat (STR) in 604 unrelated individuals collected from nine human populations (Japanese, Northern Han, Hui, Uygur, Kazakhstan, Iranian, Saudi Arabian, Greek and Italian) determined using the polymerase chain reaction (PCR) combined with fluorescent-based automated fragment detection technology. All alleles were present in each population, but allelic distributions varied from one population to another. No new alleles (such as A7 or A8) were identified. The evolutionary and structural significance of these data as well as the potential application to forensic medicine is discussed.

Alleles↗
PubMed1997

Quantitative measurement of HCV RNA in the serum: a comparison of three assays based on different principles.

Quantitative measurement of hepatitis C virus (HCV) RNA is useful in patients with chronic hepatitis C, especially with interferon treatment. We examined the clinical usefulness of the AMPLICOR monitor assay, a newly developed assay for quantitative measurement, by comparing it with two other assays with different principles. A total of 48 patients with chronic hepatitis C who were treated with interferon-alpha (IFN-alpha) were studied: 19 were complete responders and 29 were non-responders. Hepatitis C virus RNA was measured quantitatively by AMPLICOR, branched DNA (bDNA) probe, and competitive polymerase chain reaction (C-PCR) assays. An internal quantification standard was used in the AMPLICOR assay. A cDNA competitor with a deletion of 15 base pairs in the middle portion was used in the C-PCR method. The concentration of HCV RNA was significantly correlated between the three assays adopted in this study. Sensitivity of assays was 100% by C-PCR, 90% by AMPLICOR and 69% by bDNA assays. The active quantitative range was best with the C-PCR assay and worst with the bDNA assay. The bDNA assay had a tendency to exhibit lower values for patients with serotype 2 than did the other two assays. The predictive rate of the long-term response to IFN-alpha therapy, before its initiation, was over 75% in all three assays. The predictive rate just after completing IFN-alpha therapy was as high as 80% by C-PCR and the AMPLICOR assays, but was low (58%) with the bDNA assay. The handling of the bDNA and AMPLICOR assays was much easier than the C-PCR assay, which required time and skill. These results indicate that the AMPLICOR assay is a simple and reliable method for measuring the serum concentrations of HCV RNA, and thus is suitable for clinical application.

Adult↗
PubMed1997

Resistance to activated protein C and Arg 506 Gln factor V mutation are uncommon in eastern Asian populations.

We investigated the prevalence of the factor V (FV) Arg 506 Gln mutation in healthy subjects from three eastern Asian countries (Japan, n = 270; China, n = 113; and Korea, n = 93) and in 26 Japanese patients showing venous thromboembolic events. The patients were also examined for activated protein C (APC) resistance by using the Coatest APC resistance kit. The FV mutation was investigated by polymerase chain reaction and restricted enzyme digestion with MnlI RFLP assay of the FV gene. None of the patients showed APC resistance, while all subjects examined were homozygous for Arg at position 506 of the FV gene. Our results imply that FV mutation and APC resistance contribute little to venous thrombotic diseases in eastern Asia.

Adult↗
PubMed1997

Carquinostatin B, a new neuronal cell-protecting substance produced by Streptomyces exfoliatus.

Brain ischemia injury is elicited by the excitotoxicity of L-glutamate. Carquinostatin B was isolated from Streptomyces exfoliatus 2419-SVT2 as a potent neuroprotective substance which protests neuronal hybridoma N18-RE-105 cells from L-glutamate toxicity. The structure of carquinostatin B was established principally by NMR studies to be a carbazole derivative with an ortho quinone function.

Animals↗