Biomedical subjects

T Hishinuma

Publications and source records attributed to T Hishinuma.

At least 37 records · Page 2Linked to original sources

PubMed1999

Determination of leukotriene E4 in human urine using liquid chromatography-tandem mass spectrometry.

A liquid chromatographic-tandem mass spectrometric (LC-MS-MS) method was developed for the quantitation of urinary leukotriene E4 (LTE4). LTE4 and its internal standard were extracted by solid-phase extraction and analysed using LC-MS-MS in the selected reaction monitoring (SRM) mode. A good linear response over the range of 10 pg to 10 ng was demonstrated. The accuracy of added LTE4 ranged from 97.0% to 108.0% with a mean and SD of 100.6+/-2.4%. We detected LTE4 (63.1+/-18.7 pg/mg creatinine, n=10) in healthy human urine. This method can be used to determine LTE4 in biological samples.

Adult↗

PubMed1999

Effects of long-term supplementation of eicosapentanoic and docosahexanoic acid on the 2-, 3-series prostacyclin production by endothelial cells.

We studied the effects of polyunsaturated fatty, acids such as arachidonic acid [20:4 (n-6)], eicosapentanoic acid [EPA, 20:5 (n-3)], and docosahexanoic acid [DHA, 22:6 (n-3)] on the changes of lipid profiles and prostacyclin production by cultured bovine aortic endothelial cells. The amounts of 6-keto-prostaglandin F1alpha(6-keto-PGF1alpha) and delta17-6-keto-PGF1alpha, non-enzymatic metabolites of prostacyclin (PGI2 and PGI3) in culture medium were measured by gas chromatography/selected ion monitoring. Endothelial cells were supplemented for five passages with arachidonic acid, EPA, or DHA, and the fatty acids of cell lipids and prostacyclin production in cultured medium were quantified. From the fatty acid analysis, the amounts of docosapentaenoic acid [22:5 (n-3)] were significantly increased in EPA-grown cells. In DHA-grown cells, the amounts of EPA were slightly increased compared to control cells. These cells produced similar amounts of PGI2 as the controls, but larger amounts of PGI3 under basal conditions. These findings suggest that EPA, docosapentaenoic acid, and DHA are interconverted to each other, and anti-aggregatory effects of EPA or DHA may be partially due to the stimulation of prostacyclin formation in endothelial cells.

Animals↗

PubMed1999

Microdetermination of the 6,15-diketo-13,14-dihydroprostaglandin F1alpha in human plasma using gas chromatography/selected ion monitoring with [18O]6,15-diketo-13,14-dihydro-prostaglandin F1alpha as an internal standard.

We devised a simple and effective purification method for the microdetermination of 6,15-diketo-13,14-dihydro-prostaglandin F1alpha (DK), a metabolite of prostacyclin (PGI2). [18O]DK was synthesized from the repeated base-catalyzed hydrolysis of methyl ester derivatives in [18O] water to obtain an internal standard for the gas chromatography/selected ion monitoring (GC/SIM) of DK. The methyl ester-methoxime-tert-butyldimethylsilyl ether derivative was prepared, then gas chromatography/selected ion monitoring was carried out by monitoring the ion at m/z 613.4 for DK and at m/z 617.4 for an internal standard. A good linear response over the range of 10 pg to 10 ng was demonstrated. We detected DK to a level of about 297.8 pg/ml in human plasma. This method can be used to determine DK in biological samples.

6-Ketoprostaglandin F1 alpha↗

PubMed1999

Microdetermination of prostaglandin E2 in joint fluid in rheumatoid arthritis patients using gas chromatography/selected ion monitoring.

We devised an effective purification for the microdetermination of prostaglandin E2 (PGE2) in human joint fluid using gas chromatography/selected ion monitoring and determined PGE2 in the joint fluid in rheumatoid arthritis (RA) patients using this method. The methyl estermethoxime-tert-butyldimethylsilyl ether derivative was prepared, then gas chromatography/selected ion monitoring was carried out by monitoring the ion at m/z 566.4 for PGE2 and at m/z 570.4 for the internal standard (PGE2-d4). A good linear response over the range of 10 pg to 50 ng was demonstrated. We detected PGE2 to a level of about 46 pg/ml in the joint fluid of RA patients. The level of PGE2 in RA patients was significantly higher than that in osteoarthritis patients used as controls. Moreover, we measured inflammatory cytokine (IL-1beta, TNFalpha, IL-6 receptor) levels in joint fluid by using enzyme-linked immunosorbent assay. A relationships between the PGE2 level in joint fluid and these cytokines or biochemical data as the indicator of RA disease was not observed. We found that the PGE2 level in each patient was influenced by therapeutic drugs. The PGE2 level in RA patients with non-steroidal anti-inflammatory drugs was lower than with steroids.

Arthritis, Rheumatoid↗

PubMed1999

Urinary thromboxane A2/prostacyclin balance reflects the pathological state of a diabetic.

Levels of the stable urinary metabolites of thromboxane A2 and prostacyclin, 11-dehydro-thromboxane B2 (11-dehydro-TXB2) and 2,3-dinor-6-keto-prostaglandin F1alpha (2,3-dinor-6-keto-PGF1alpha) were measured in diabetics to elucidate the relation between the thromboxane A2/prostacyclin (TX/PGI) balance and pathological states of diabetes mellitus. 11-Dehydro-TXB2 and 2,3-dinor-6-keto-PGF1alpha were derivatized to methyl ester-propylamide-dimethylisopropylsilyl ether and methyl ester-methoxime-dimethylisopropylsilyl ether derivatives, respectively, and applied to a gas chromatography/selected ion monitoring. The TX/PGI ratios of diabetics were higher than those of healthy volunteers, suggesting the hypercoagulative states of this disease. The ratios showed positive correlations with the levels of blood glucose. The levels of hemoglobin A1c and triglyceride were correlated weakly with the ratio. Some of the patients who had relatively low levels of blood glucose also showed high TX/PGI ratios. Furthermore, the ratio increased in the order of the groups 1, 2, and 3; group 1 contained patients who did not take medicine for diabetes, group 2 contained those who took oral hypoglycemic agents, and group 3 contained those who received insulin therapy. These observations indicate that the TX/PGI ratio reflects the pathological conditions of diabetes and is a useful marker, having few different features from other markers that are presently used.

6-Ketoprostaglandin F1 alpha↗

PubMed1999

Monitoring of the thromboxane A2/prostacyclin ratio in the urine of patients with retinal vascular occlusion through the low-dose-aspirin therapy using the gas chromatography/selected ion monitoring method.

We determined the levels of the stable urinary metabolites of thromboxane A2 and prostacyclin, 11-dehydro-thromboxane B2 (11-dehydro-TXB2) and 2,3-dinor-6-keto-prostaglandin F1alpha (2,3-dinor-6-keto-PGF1alpha) in patients with retinal vascular occlusion (RVO) to elucidate the change of the thromboxane A2/prostacyclin (TX/PGI) ratio with this disease and the effect of low-dose-aspirin therapy. 11-Dehydro-TXB2 and 2,3-dinor-6-keto-PGF1alpha were converted to 1-methyl ester-propylamide-9,12,15-tris-dimethylisopropylsilyl ether derivative and 1-methyl ester-6-methoxime-9,12,15-tris-dimethylisopropylsilyl ether derivative, respectively, and applied to a gas chromatography/selected ion monitoring. The average level of 11-dehydro-TXB2 in 30 patients with RVO was 1038 +/- 958 pg/mg creatinine. It was significantly higher than that of 27 healthy volunteers, which was 616 +/- 294 pg/mg creatinine (p < 0.05 with unpaired t-test). However, 2,3-dinor-6-keto-PGF1alpha levels were not significantly different between these two groups. The average ratio of TX/PGI in the RVO patients was 32 +/- 26 and it was significantly higher than that of healthy volunteers, 17 +/- 10 (p < 0.01). Patients with central retinal artery occlusion or branch retinal artery occlusion showed greatly high 11-dehydro-TXB2 levels and TX/PGI ratios, although the number of patients was limited in the current study. After the administration of low-dose aspirin (40 mg/day) for about 1 month, the TX/PGI ratio decreased to around the normal level. Following the levels for up to 10 months, they also remained at the normal level. These observations suggested that the 11-dehydro-TXB2 levels and the TX/PGI ratio reflect the pathological conditions of RVO and are useful markers of the treatment.

6-Ketoprostaglandin F1 alpha↗

PubMed1998

Analysis of urinary 11-dehydrothromboxane B2 in patients with occluded retinal vein using GC/SIM.

We describe variations of 11-dehydrothromboxane B2(11-dehydro-TXB2) levels in human urine samples. Retinal vein occlusion (RVO) is a thrombotic disease in which the retinal vein is blocked by blood aggregations. We considered the possibility that 11-dehydro-TXB2 plays an important role in the formation of RVO. Thus, we determined the 11-dehydro-TXB2 levels in patients with RVO using gas chromatography/selected ion monitoring (GC/SIM) and compared them with those of healthy volunteers. The thromboxane levels in patients with RVO, who did not also have diabetes, were significantly higher than those in healthy volunteers. One cause of RVO may be the variation of thromboxane production. Furthermore, this GC/SIM method can be applied to the prevention and treatment of not only RVO, but also of general thrombosis.

Adult↗

PubMed1997

Microdetermination of the thromboxane B3 gas chromatography-selected-ion monitoring using [18O]thromboxane B3 as an internal standard.

We devised a simple and effective purification for the microdetermination of thromboxane B3 (TXB3), a hydrolysis product of TXA3- [18O2]TXB3 was synthesized by the repeated base-catalyzed hydrolysis of methyl ester derivatives in [18O]water, to obtain an internal standard (IS) for the gas chromatography/selected ion monitoring (GC/SIM) of TXB3. The methyl ester (ME)-methoxime (MO)-dimethylisopropylsilyl (DMIPS) ether derivative was prepared, then GC/SIM was carried out by monitoring the ion at m/z 668 for TXB3 and that at m/z 672 for IS. A good linear response over the range of 10 pg approximately 10 ng was demonstrated. We were able to detect the levels of TXB3 in the medium of human erythroleukemia (HEL) cell cultured with eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA). This method can be used to determine 3-series thromboxane in biological samples.

Calibration↗

PubMed1997

Effects of haloperidol and cocaine pretreatments on brain distribution and kinetics of [11C]methamphetamine in methamphetamine sensitized dog: application of PET to drug pharmacokinetic study.

Repeated administration of methamphetamine (MAP) causes behavioral sensitization in animals. We previously reported that the maximum accumulation level of [11C]MAP in the MAP-sensitized dog brain was 1.4 times higher than that in the control. In behavioral studies, haloperidol (a dopamine D2 receptor antagonist) prevents MAP-induced behavioral sensitization, and cocaine (a dopamine reuptake blocker) has the cross-behavioral sensitization with MAP. In the present study, to elucidate the relation between the MAP-induced behavioral sensitization and the pharmacokinetics of MAP, we investigated the effects of haloperidol and cocaine pretreatments on brain regional distribution and kinetics of [11C]MAP using positron emission tomography (PET). A significant increase of [11C]MAP uptake into the sensitized dog brain was prevented by haloperidol and cocaine pretreatments. These pharmacokinetic changes were not due to the changes in the rate of MAP metabolism. These results suggest haloperidol and cocaine can change the cerebral pharmacokinetic profile of MAP in the behavioral-sensitized dog. The variations of MAP-accumulation may affect the development or expression of MAP-induced behavioral sensitization.

Animals↗

PubMed1997

Alteration of DNA methylation levels in MRL lupus mice.

Recent reports suggest that DNA methylation is involved in the cause of autoimmune disease. We investigated the alteration of DNA methylation levels in lupus strains of mice, MRL/lpr as a model, which develop an age-dependent lymphadenopathy and autoimmune disease. DNA methylation levels of thymus and axillary lymph nodes in 20-week-old MRL/lpr mice, which are in an autoimmune disease state, were lower than those of 4-week-old MRL/lpr mice with no symptoms as yet. No significant changes were observed in MRL/+ strain mice, which seemed normal at least 20 weeks, while DNA methylation levels in the spleen of both strains of mice increased significantly from the age of 4 to 20 weeks. However, no significant changes of DNA methylation levels in peripheral blood were observed with ageing in MRL strains. Moreover, we clarified that administration of 5-azacytidine had a strong effect on longer survival of MRL/lpr mice and reduced DNA methylation levels in the axillary lymph nodes and spleen. The possible relevance of DNA methylation levels to the progression of autoimmune disease is discussed.

Animals↗

PubMed1997

Molecular cloning of the nemA gene encoding N-ethylmaleimide reductase from Escherichia coli.

Using the gene mapping membrane technique, we identified a gene (nemA) that encodes N-ethylmaleimide reductase in Escherichia coli. The open reading frame encodes a polypeptide of 365 amino acids with a molecular mass of 39,514 Da. The deduced amino acid sequence showed a high degree of homology (87% identical) with the pentaerythritol tetranitrate reductase of Enterobacter cloacae and the morphinone reductase of Pseudomonas putida (52% identical).

Amino Acid Sequence↗

PubMed1997

Phenobarbital molecularly imprinted polymer selectively binds phenobarbital.

Molecularly imprinted polymer (MIP) was prepared against phenobarbital using methacrylic acid as the functional monomer and ethylene glycol dimethacrylate as the cross linking monomer. We analyzed the recognition properties of the phenobarbital MIP. In some organic solvents, imprinted polymer showed selective binding to phenobarbital. Two dissociation constants of binding were calculated by Scatchard plot analyses; Kd values were 1.8, 121.7 microM, and the number of binding sites was 8.3, 92.3 mumol/g MIP in toluene-heptane-acetic acid (25 : 75 : 1, v/v), respectively. The relationship between the binding affinity to phenobarbital MIP and the polarity of the solvent system, as well as the structure of the template molecule is also discussed.

Cross-Linking Reagents↗

PubMed1997

4 Pro-R hydrogen of NADPH was abstracted for enzymatic hydride transfer by N-ethylmaleimide-reductase of Yarrowia lipolytica.

We studied the steric course of the reaction catalyzed by the N-ethylmaleimide (NEM) reductase of Yarrowia (Candida) lipolytica (Y. lipolytica), using 4R-[4-2H1]NADPH and 4S-[4-2H1]NADPH as cofactors and N-ethylcitraconimide as substrate. Active substrates and inhibitors of NEM reductase and its subcellular distribution were also investigated to clarify the biochemical properties of this enzyme. NEM reductase catalyzes the reduction of N-ethylmaleimide to N-ethylsuccinimide with NAD(P)H as the cofactor. Several maleimide and cyclopentenone derivatives tested were also active substrates for NEM reductase of Y. lipolytica. Some pyrazolone derivatives, particularly 1-phenyl-5-pyrazolone, were found to be effective inhibitors of NEM reductase. Subcellular localization of NEM reductase was carried out using protoplast formation and differential centrifugation. Ninety-eight percent of the NEM reductase activity was recovered in the cytosolic fraction, indicating that NEM reductase in Y. lipolytica was the cytosolic enzyme. We also determined the stereochemical specificity of the reduction of N-ethylcitraconimide by NEM reductase in Y. lipolytica, showing that 4 Pro-R hydrogen of NADPH was abstracted for enzymatic hydride transfer by NEM reductase, and two hydrogen atoms from NADPH and H2O added to opposite faces of the double bond of N-ethylcitraconimide.

Candida↗

PubMed1997

[Radiotherapy of esophageal cancer: clinical usefulness of new grouping].

A total of 188 cases of nonresected esophageal cancer were categorized into the following groups based on T category. (1987 UICC) and radiological classification (Japanese Society of Esophageal Diseases); Group 1: superficial or early (T2 or less) tumorous type; Group 2: advanced tumorous type, early serrated or early spiral type; Group 3: others. The response to radiotherapy was significantly related to the group. According to the proportional hazard model, the important factors predicting long-term survival were T category, radiotherapy response at the end of treatment, and group. There was a tendency to obtain better local control in group 1 patients than in group 2 patients, but cause-specific survival was the same for both groups (mainly the appearance of distant metastases). Local control was poor in group 3. If the tumor response at 40Gy was excellent in group 2 patients, the main cause of death was distant metastases, much the same as in group 1 patients. However, if response at 40Gy was not excellent, the main cause of death was local failure even though final tumor response was excellent. It seemed that therapy that may improve local control is valuable for group 3 patient, and for group 2 patients whose tumor response at 40Gy was not excellent.

Aged↗

PubMed1997

The effects of unsaturated fatty acids, oxidizing agents and Michael reaction acceptors on the induction of N-ethylmaleimide reductase in Escherichia coli: possible application for drug design of chemoprotectors.

Menadione and dimethyl maleate, Michael reaction acceptors, induced N-ethylmaleimide (NEM) reductase activity in Escherichia coli strain DH5a. Linoleic acid also induced NEM reductase activity, but oleic acid, which is less susceptible to lipid peroxidation than linoleic acid, did not induce NEM reductase activity. In addition, NEM reductase activity was induced by menadione and linoleic acid also in strain DH5, Y1088 and Y1090. Linoleic acid is not a Michael reaction acceptor, but is known to produce Michael reaction acceptors such as alkenals and 4-hydroxyalkenals as a result of free-radical-initiated lipid peroxidation. Thus, our findings suggested that lipid peroxidation was involved in the induction of NEM reductase by linoleic acid. The electrophilic property of Michael reaction acceptors provides the signal for induction of phase II enzymes such as glutathione S-transferase and quinone reductase in mammals. The inducer potency of phase II enzymes has been used to design chemoprotective drugs. Therefore, the inducible nature of this enzyme will serve not only for the elucidation of its physiological function, but also for the evaluation of chemoprotective drugs.

Enzyme Induction↗

PubMed1996

Positron emission tomography study of the alterations in brain distribution of [11C]methamphetamine in methamphetamine-sensitized dog.

We newly prepared a MAP-sensitized dog by repeated MAP treatment and studied the brain distribution of [11C]MAP in the normal and the MAP-sensitized dog using PET. The maximal level of accumulation of [11C]MAP in the sensitized dog brain was 1.4 times higher than that in the control. No difference was found in the metabolism of MAP between the two conditions. The significant increase of [11C]MAP in the MAP-sensitized brain indicates that subchronic MAP administration causes some functional change in the uptake site of MAP. The pharmacokinetic change may, in part, account for behavioral sensitization.

Animals↗

PubMed1996

Analysis of the thromboxane/prostacyclin balance in human urine by gas chromatography/selected ion monitoring: abnormalities in diabetics.

We microanalyzed 2,3-dinor-6-keto-prostaglandin F1 alpha (2,3-dinor-6-keto-PGF1 alpha 1) and 11-dehydrothromboxane B2 (11-dehydro-TXB2, 2) in human urine. Samples containing a [2H4]-analogue as an internal standard were extracted by chromatography using Sep Pak tC18 and silica gel. The compounds were then analysed by means of the lactone ring opening reaction and dimethylisopropylsilylation. The conversion of 1 to 1-methyl ester (ME)-propylamide (PA)-9, 12, 15-dimethylisopropylsilyl (DMIPS) ether derivative and of 2 to 1-ME-6-methoxime (MO)-9, 12, 15-tris-DMIPS ether derivative was followed by gas chromatography/selected ion monitoring (GC/SIM). Interfering substances from the urine matrix were eliminated during GC/SIM analysis using a DB-5 column. We were able to detect 1 (222-1031 pg/mg creatinine) and 2 (18-155 pg/mg creatinine) in human urine. Furthermore, the thromboxane/prostacyclin (IX/PGI) ratio in the urine of diabetics was higher than that of healthy volunteers. This method can be used to determine the TX/PGI balance in human urine.

6-Ketoprostaglandin F1 alpha↗

PubMed1996

Preparation of a monoclonal antibody specific for 5-methyl-2'-deoxycytidine and its application for the detection of DNA methylation levels in human peripheral blood cells.

A monoclonal antibody specific for a modified nucleoside, 5-methyl-2'-deoxycytidine (m5dCyd), was prepared using 5-methylcytidine (m5Cyd)-keyhole limpet haemocyanin (KLH) conjugate, and was characterized. Termed FMC9, the antibody reacts with m5dCyd and slightly with m5Cyd and 5-methylcytosine (m5Cyt) but not with other nucleosides tested in this investigation. FMC-9 was used in an enzyme-linked immunosorbent assay (ELISA) system for the quantitation of m5dCyd levels. Sensitivity was in the picomole range. Methylation levels in peripheral blood cells of healthy donors were determined by inhibition ELISA. The percentage of m5dCyd in peripheral blood cells of 10 healthy donors was 5.08 +/- 0.50%. These results suggest that the inhibition ELISA using FMC9 is useful to monitor m5dCyd levels in the peripheral blood cells.

Adult↗