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Y Oda

Publications and source records attributed to Y Oda.

At least 19 recordsLinked to original sources
PubMed1997

High-performance capillary electrophoresis of hyaluronic acid: determination of its amount and molecular mass.

The amount and the molecular mass of hyaluronic acid (HA) were determined by high-performance capillary electrophoresis. HA was observed at around 15 min by using an untreated fused-silica capillary (75 microns I.D.) of 58 cm length (effective length, 50 cm) at 20 kV in 50 mM phosphate buffer (pH 4.0). Calibration curves showed good linearity from 0.01 mg/ml to 3.3 mg/ml for all HA samples examined. The lower limit of detection by monitoring the absorbance at 185 nm was 1.0 microgram/ml at the signal-to-noise ratio of 5. HA samples were examined in a buffer containing pullulan (PU) as an additive for the matrix formation material. The HA samples showed marked peak-broadening when analyzed in the buffer solution containing PU with a specified molecular mass. The peak broadening was based on the dispersion of the molecular mass of the HA sample analyzed.

Buffers↗
PubMed1997

Simultaneous quantitative determination method for sphingolipid metabolites by liquid chromatography/ionspray ionization tandem mass spectrometry.

Sphingolipid metabolites ceramide, sphingomyelin, sphingosine, psycosine, sphingosylphosphorylcholine, and dimethylsphingosine were separated and simulataneously quantitated by liquid chromatography/ionspray ionization tandem mass spectrometry (LC/MS/ MS). The use of glassware throughout minimized losses due to adsorption and the pretreatment of this method consisted of simple liquid-liquid extraction procedure with a mixture of chloroform and methanol. After separation on a short C18 silica column eluted in a gradient mode, the metabolites were detected by MS/ MS. This assay allows simultaneously quantification of these metabolites over a range of at least 0.1 to 100 ng/ 10(6) cells. The LC/MS/MS analyses took 10 to 15 min per sample and we could examine up to 50 samples per day. We also detected endogenous sphingosine 1-phosphate in HL-60 cells. The utility of the method was demonstrated by examining changes in metabolites levels in HL-60 cells after treatment with sphingomyelinase. It was found that sphingomyelinase from Bacillus cereus may have selectivity for acyl chain length.

Ceramides↗
PubMed1997

Expression of MDR1/p-glycoprotein and multidrug resistance-associated protein in childhood solid tumours.

We evaluated the expression of MDR1/p-glycoprotein in paediatric tumours using reverse transcriptase polymerase chain reaction (RT-PCR), RNA dot blot analysis, and immunohistochemistry on formalin fixed paraffin-embedded material with JSB-1 and C-219 monoclonal antibodies, and compared these three techniques. The expression of multidrug resistance-associated protein (MRP) gene was examined by RT-PCR assay. We studied MDR1/p-glycoprotein and MRP expression in 13 samples from 10 neuroblastoma patients, 11 samples from 10 nephroblastoma patients, 2 rhabdomyosarcomas, 1 adrenocortical carcinoma and 10 benign tumours or tumour-like lesions. Eleven of 13 neuroblastomas, 7 of 11 nephroblastomas, 2 rhabdomyosarcomas, 1 adrenocortical carcinoma, and 7 of 10 benign tumours or tumour-like lesions showed MDR1 PCR products. By RNA dot blot analysis, MDR1 transcripts were detectable in 11 of 34 specimens. Immunohistochemically, we detected positive reaction products for JSB-1 in 26 of 36 samples. There was a significant correlation between the immunoreactivity for JSB-1 and the expression of MDR1 mRNA expression by RT-PCR (P = 0.0001). However, the presence of p-glycoprotein immunostaining does not correlate with the MDR1 expression shown by RT-PCR in every case. As for MRP mRNA expression, 9 of 13 neuroblastomas and 10 of 11 nephroblastomas revealed PCR products.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

p53 gene mutations in soft-tissue sarcomas--correlations with p53 immunohistochemistry and DNA ploidy.

The significance of p53 mutations in a group of 67 soft-tissue tumors was examined using single-strand conformation polymorphism and direct sequencing analysis. Molecular findings were correlated with immunohistochemical detection of the p53 protein and DNA ploidy status. Mutations of the p53 gene were detected in 13 (19.5%) out of 67 cases of soft-tissue tumors. Only three were localized outside the conservative regions of the p53 gene. Six mutations were described for the first time in these tumors. Most of the mutations were point mutations in exons 5-8 and, in one case, a deletion at the 3'-splice site of exon 5 could be demonstrated. There was no significant correlation between the occurrence of p53 mutations and the histological grade, although a high number of mutations were defined in poorly differentiated tumors (grade 3). Molecular finding of a p53 gene mutation and immunohistochemical detection of p53 expression did not correlate, which may be due to the high percentage of nonsense mutations in our study (50%). We confirm that only DNA sequencing allows a unique identification and differentiation of mutations in the p53 gene. Other factors may be responsible for the detection of p53 protein in many cases. Histological grade correlated with aneuploidy. The frequency of mutations observed was in accordance with values quoted in the literature. Generally, p53 mutations and p53 overexpression are more likely to represent a late event in the oncogenesis of soft-tissue tumors.

Adult↗
PubMed1997

Expression of multidrug resistance-associated protein gene in Ewing's sarcoma and malignant peripheral neuroectodermal tumor of bone.

The expression of multidrug resistance-associated protein (MRP) mRNA was examined in ten samples of Ewing's sarcoma of bone (ES) and in one nude mice transplantable ES and two malignant peripheral neuroectodermal tumor (MPNT) cell lines using an RT-PCR assay. MRP mRNA expression was recognized in eight of the ten clinical specimen and in all three cell lines. On the other hand, the expression of multidrug resistance gene (MDR1) was demonstrated in three of the ten clinical samples and all three cell lines. Our results may contribute to elucidation of the mechanism of anti-cancer-drug resistance in this tumor.

ATP-Binding Cassette Transporters↗
PubMed1997

Purification and properties of poly(3-hydroxybutyrate) depolymerase from the fungus Paecilomyces lilacinus D218.

Poly(3-hydroxybutyrate) depolymerase was purified to homogeneity from the culture filtrate of Paecilomyces lilacinus D218 by column chromatography on CM-Toyopearl 650M and hydroxylapatite. The molecular weight of the enzyme was estimated to be 48,000 by SDS-PAGE. Maximal activity was observed near pH 7.0 and 45 degrees C. The Km and Vmax values for PHB were 0.13(mg/ml) and 3750 (U/mg protein), respectively. The enzyme hydrolyzed PHB and p-nitrophenyl fatty acids but not polycaprolactone and triglycerides.

Carboxylic Ester Hydrolases↗
PubMed1997

Pure akinesia manifested neuroleptic malignant syndrome: a clinical variant of progressive supranuclear palsy.

An autopsy case of pure akinesia (PA) is reported. The patient manifested L-dopa-unresponsive akinesia without accompanying rigidity, tremor, eye movement disorder or dementia from the age of 58 years. Brain magnetic resonance T2-weighted imaging at the age of 63 showed high intensity areas in the subthalamic regions, but brain atrophy was not observed. She received amantadine-HCl and L-threo-3,4-dihydroxyphenylserine (L-DOPS) for 5 years. At the age of 66, she died of the severe illness accompanied by consciousness disturbances, hyperthermia, muscle rigidity, abnormal blood pressure and elevated serum enzymes which were derived from the muscle. We considered her condition to be neuroleptic malignant syndrome (NMS). Pathologically the brain revealed degeneration in the subthalamic nucleus, globus pallidus and substantia nigra. Neurofibrillary tangles were detected in the temporal cortex, hippocampus, amygdaloid body and spinal cord, as well as in the basal ganglia, thalamus and brain stem. These findings were consistent with that of progressive supranuclear palsy (PSP); the change in the ventral pons was insignificant, suggesting that PA may have minimum involvement in the ventral pons. The skeletal muscle showed scattered necrosis that was compatible with NMS. As far as we know, this is the first report of NMS accompanied with PA.

Brain↗
PubMed1997

Human choline acetyltransferase mRNAs with different 5'-region produce a 69-kDa major translation product.

Choline acetyltransferase (ChAT, EC 2.3.1.6) is the biosynthetic enzyme for acetylcholine. We have previously shown that multiple ChAT mRNA species with different 5'-noncoding regions are expressed in the rat and mouse. However, the diversity of ChAT mRNA species in human has not completely been elucidated. In this work N1- and N2-type ChAT cDNAs were cloned from a human brain cDNA library and the N-exon located in the human ChAT gene. Polymerase chain reaction analysis indicates that four species of ChAT mRNAs (R-, N1-, N2- and M-types) are produced in human brain and spinal cord. In all human transcripts, the ATG initiation codon in the rat, mouse and pig was replaced by ACG, which does not serve as an initiation codon for translation. In vitro translation and mammalian expression analyses revealed that N1-, N2- and R-type mRNAs give rise to a single 69 kDa enzyme, while M-type mRNA produces both 82 and 69 kDa enzymes. The translation efficiency of M-type mRNA was lower than that of the other mRNA species. Moreover, the translation efficiency of human ChAT mRNAs was considerably lower than that of rat ChAT mRNA, suggesting that the ATG codons for human ChAT are unfavorable for translation initiation compared with the initiation codon for rat ChAT. These results provide rational explanations for the previous reports that human ChAT protein purified from the brain and placenta had 66-70 kDa molecular mass, and that ChAT activity in a single motor neuron of human was far lower than that of other vertebrates. Sequencing of monkey ChAT gene showed that the initiation ATG in rodent ChAT was also replaced by ACA in the monkey.

Alternative Splicing↗
PubMed1997

High frequency of K-ras mutations in human colorectal hyperplastic polyps.

BACKGROUND: Hyperplastic polyps are common benign colorectal polyps, and are thought to have little association with malignant tumours in the colorectum. However, several reports suggest that some hyperplastic polyps may develop into colorectal neoplasms. AIM: To clarify genetic alterations in colorectal hyperplastic polyps. METHODS: Twenty eight colorectal polyps having serrated components were resected from patients endoscopically. The K-ras gene mutations in codons 12 and 13 were analysed by PCR-RFLP. Intranuclear p53 protein was immunostained by the avidin-biotin complex method. RESULTS: A mutation of the K-ras gene was detected in nine (47%) of 19 hyperplastic polyps, and five (56%) of nine adenomas. p53 protein nuclear accumulation was detected immunohistochemically in two (22%) of nine adenomas, but not in any of the hyperplastic polyps. CONCLUSION: Some hyperplastic polyps may be true neoplastic lesions, and could be precursors of malignant neoplasia.

Adenoma↗
PubMed1997

Phase I-II study of irinotecan hydrochloride combined with cisplatin in patients with advanced gastric cancer.

PURPOSE: A dose-escalation study of irinotecan hydrochloride (CPT-11) combined with fixed-dose cisplatin was conducted to determine the maximum-tolerated dose (MTD), dose-limiting toxicities, and objective response rate in patients with advanced gastric cancer. PATIENTS AND METHODS: Twenty-four patients with or without prior chemotherapy were enrolled. All patients were assessable for toxicities and response. On day 1, CPT-11 was administered as a 90-minute intravenous (I.V.) infusion, which was followed 2 hours later by a 120-minute I.V. infusion of cisplatin 80 mg/m2. CPT-11 alone at the same dose was administered again on day 15. The treatment was repeated every 4 weeks until disease progression was observed. The initial dose of CPT-11 was 60 mg/m2, and was escalated in increments of 10 mg/m2 until severe or life-threatening toxicity was observed. RESULTS: The MTD of this combination was CPT-11 80 mg/m2. At this dose level, 16.7% of patients (two of 12) had leukopenia of less than 1,000/microL, 66.7% (eight of 12) had neutropenia of less than 500/microL, and 16.7% (two of 12) had severe diarrhea of grade 4 during the first course. The dose-limiting toxicity was neutropenia. Ten patients achieved a partial response (PR), and the overall response rate was 41.7% among 24 patients (95% confidence interval, 21.9% to 61.4%). CONCLUSION: The recommended dose and schedule is CPT-11 70 mg/m2 on days 1 and 15 and cisplatin 80 mg/m2 on day 1 every 4 weeks. This combination of CPT-11 and cisplatin, considered to be active against advanced gastric cancer with acceptable toxicity, should be further assessed in a phase II study.

Adult↗
PubMed1997

Thyrotropin (TSH) receptor autoantibodies do not appear to bind to the TSH receptor produced in an in vitro transcription/translation system.

An in vitro transcription/translation (TnT) system was used to produce 35S-labeled full-length TSH receptor (TSHR) and TSHR extracellular domain (TSHRex). The interaction of the labeled proteins with TSHR autoantibodies in Graves' sera was then studied using an immunoprecipitation assay. In the assay, 35S-labeled TSHR or TSHRex were incubated with test sera, and any immune complexes formed were precipitated with protein A-Sepharose (in the case of mouse monoclonal antibodies, antimouse IgG-agarose was used). Rabbit antibodies to the TSHR and a mouse monoclonal antibody precipitated as much as 50% of the 35S-labeled TSHR preparations compared with about 2% for normal rabbit serum and 4% for a control monoclonal antibody. However, none of 34 Graves' sera (TSHR autoantibody levels ranging from 14-95% inhibition of [125I]TSH binding) were able specifically to immunoprecipitate 35S-labeled TSHR or TSHRex. These negative findings were confirmed by analysis of the immunoprecipitates on SDS-PAGE followed by autoradiography. Our results indicate that the TnT system is not useful for producing labeled TSHR preparations that can bind TSHR autoantibodies well. This is in contrast to TnT produced 35S-labeled glutamic acid decarboxylase, thyroid peroxidase, and 21-hydroxylase, which react well with their respective autoantibodies. One main difference between these 3 autoantigens and the TSHR is that the receptor is highly glycosylated, and this extensive glycosylation may be of critical importance for correct folding of the receptor. Consequently, the inability of the TnT system to glycosylate proteins could explain in part why TnT-produced 35S-labeled TSHR and TSHRex do not bind TSHR autoantibodies.

Animals↗
PubMed1997

Synthesis and antitumor activity of fused quinoline derivatives. IV. Novel 11-aminoindolo[3,2-b]quinolines.

Indolo[3,2-b]quinoline derivatives (1) having various amine moieties were prepared and their antitumor activities against P388 leukemia in mice were evaluated, for the purpose of gaining an insight into the role of the amine moiety in the antitumor activity and searching for an effective amine moiety. Introduction of a methylene group between the phenyl group and amino or methanesulfonamido group resulted in decrease or loss of activity.

Animals↗
PubMed1997

Localization and quantification of proliferating cells during rat fracture repair: detection of proliferating cell nuclear antigen by immunohistochemistry.

Bilateral femurs of 12-week-old female Sprague-Dawley rats were fractured, and the fractured femurs were harvested 36 h, 3, 7, 10, and 14 days after the fracture. Localization of cell proliferation in the fracture calluses was investigated using immunohistochemistry with antiproliferating cell nuclear antigen (PCNA) monoclonal antibodies. Thirty-six hours after the fracture, many PCNA-positive cells were observed in the whole callus. The change was not limited to mesenchymal cells at the fracture site where the inflammatory reaction had occurred, but extended in the periosteum along almost the entire femoral diaphysis where intramembranous ossification was initiated. On day 3, periosteal cells or premature osteoblasts in the newly formed trabecular bone during intramembranous ossification still displayed intense staining. On day 7, many premature chondrocytes and proliferating chondrocytes were PCNA positive. Endochondral ossification appeared on days 10 and 14, and the premature osteoblasts and endothelial cells in the endochondral ossification front were stained with anti-PCNA antibodies. Quantification of PCNA-positive cells was carried out using an image analysis computer system, obtaining a PCNA score for each cellular event. The highest score was observed in the periosteum early after the fracture near the fracture site. Immunohistochemistry using anti-PCNA antibodies showed that the distribution of proliferating cells and the degree of cell proliferation varied according to the time lag after the fracture, suggesting the existence of local regulatory factors such as growth factors, and that significant cell proliferation was observed at the beginning of each cellular event.

Animals↗
PubMed1997

Neurotoxicity testing of a new bioactive bone cement.

Bioactive bone cement (BABC) is a novel artificial bone cement. It has some noteworthy characteristics that are applicable to neurological surgery. The toxicity of BABC to the nervous system was tested by implanting BABC and polymethylmethacrylate (PMMA) cement as a control at the parietal and the suboccipital regions of the skull in beagles. The auditory brainstem response (ABR) was tested before and after implantation. Sections of the cerebral cortex and the acoustic nerve were examined at 3 and 6 months after implantation. No abnormal ABR was found in any animals. Histological examination of the cerebral cortex and acoustic nerve demonstrated slight gliosis in both the BABC and PMMA cement groups, but no other abnormalities.

Animals↗
PubMed1997

Expression of P-glycoprotein in high grade osteosarcomas with special emphasis on chondroblastic subtype.

The development of chemoresistance is one of the major clinical problems in the therapy of malignant bone tumors in childhood. The expression of membrane-bound P-glycoprotein turned out to be an essential factor in the evidence of resistant tumor cells. To investigate the significance of multidrug resistance in the prognosis of highly malignant osteosarcomas, the immunohistologic expression of P-glycoprotein was investigated in the tumor tissue of 52 patients under special consideration of the histologic subtype. The data were compared with the histologic regression grade in the resection specimen and correlated with clinical data. Formalin-fixed, paraffin-embedded tissue and, additionally, fresh frozen material taken from the primary biopsy were stained using monoclonal antibody JSB1. 29 (55%) of the tumors investigated were P-glycoprotein positive. Considering the response to chemotherapy, no conclusion could be drawn regarding P-glycoprotein expression, regression grade in the resection specimens, and the clinical follow-up. P-glycoprotein was detected in only 52% of the non-responders. A positive reaction was also evidenced in 59% of the patients with high chemosensitivity. A comparison of the histologic subtypes yielded a significant result in the chondroblastic osteosarcomas. 11 of 12 cases showed a strong expression of P-glycoprotein. Most of the cases were non-responders, and using Kaplan-Meier live tables, an unfavorable clinical outcome could be demonstrated. Possibly, chondroblastic tumors have a special position among osteosarcomas because of their differentiation.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

Effects of 2'-substituents of the first deoxyguanosine residue in the recognition sequence of EcoRI restriction endonuclease activity.

The effects of 2'-substituents of the first deoxyguanosine on EcoRI activity were examined using synthetic octadeoxynucleotides D(GG*AATTCC) containing 2'-substituted derivatives (G*), i.e., 2'-fluoro-2'-deoxyguanosine (dGfl), 2'-chloro-2'-deoxyguanosine (dGcl), and guanosine (rG). The overall structures of the octamers were very similar, as shown by CD and UV measurements, although their EcoRI reactivities were very different: 100% in 60 min for d(GGAATTCC) and d(GGflAATTCC), 5% in 24 h for d[G(rG)AATTCC], and no cleavage at all in 24 h for d(GGclAATTCC). However, the kinetics showed the octamers exhibit similar binding-affinity to the enzyme (10(-6)-10(-7) M). 31P-NMR analysis suggested the modified octamers change the phosphate backbone conformation in a duplex, since an unusual downfield-shifted signal in the spectra was commonly observed for the modified octamers at low temperature (i.e., a duplex state), which was shifted upfield at high temperature (i.e., a single strand state). The order of the differences was dGcl > rG > dGfl-containing octamers, coinciding with that of the vdW volume of 2'-substituents (Cl > OH > F) and the cleavage reactivities. These findings suggest that steric hindrance by the 2'-substituents causes of conformational change of the phosphate backbone close to the scissile bond, and then interferes with the EcoRI reaction.

Binding Sites↗