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S Akiyama

Publications and source records attributed to S Akiyama.

At least 19 recordsLinked to original sources
PubMed1992

Preparation of a fluorescent derivative of benzoylecgonine, and preliminary studies of its application to the analysis of urine.

A sensitive high-performance liquid chromatographic method using 3-bromomethyl-6,7-dimethoxy-1-methyl-2(1H)-quinoxalinone (Br-DMEQ) as a fluorescent labeling reagent is described for the determination of benzoylecgonine (BE) and ecgonine (EC). The Br-DMEQ derivatives of BE and EC were separated on a C18 column and detected at 455 nm with excitation at 370 nm. The detection limits of the proposed method were 18.7 fmol for BE and 12.5 pmol for EC at a signal-to-noise ratio of 3. Relative standard deviations of five replicate measurements were 1.94% (10 pmol) and 2.98% (50 pmol) for BE and 6.3% (250 pmol) and 5.62% (1.25 pmol) for EC. This method was applied to the determination of BE in human urine. BE was extracted from urine by solvent extraction with chloroform-isopropyl alcohol (9:1, v/v) solution. Levels of 2.5.10(-8) M BE in urine (25 pmol/ml) could be determined.

Chromatography, High Pressure Liquid↗
PubMed1992

Endogenous tumor necrosis factor functions as a resistant factor against hyperthermic cytotoxicity.

One of the mechanisms of cytotoxicity by tumor necrosis factor (TNF) is the induction of reactive oxygen molecules. Cells producing endogenous tumor necrosis factor (enTNF) show resistance to the cytotoxicity of exogenous TNF by scavenging the reactive oxygen molecules. The intracellular hydroxyl radical production is also known to be involved in the heat-induced cytotoxicity. In the present study, we therefore examined the possibility that enTNF may act as a protective protein against the heat-induced cytotoxicity in a manner similar to that of exogenous TNF. Heat-sensitive L-M (mouse tumorigenic fibroblast) cells, originally expressing no enTNF, were transfected with a human TNF expression vector to produce enTNF. The stable transfectants showed apparent resistance to heat treatment. Conversely, when HeLa (human uterine cervical cancer) cells, originally producing an appreciable amount of enTNF, were transfected with an antisense TNF mRNA expression vector to inhibit enTNF synthesis, their heat sensitivity was enhanced. Furthermore, L-M cells which were transfected with nonsecretory human TNF expression vector also acquired resistance to heat treatment. In these cells, heat resistance correlated well with expression of enTNF and intracellular levels of manganous superoxide dismutase. These results indicate that enTNF exerts its intracellular protective effect against the heat-induced cytotoxicity by scavenging reactive oxygen with induced manganous superoxide dismutase in a manner similar to that found in cells treated with exogenous TNF.

Animals↗
PubMed1992

Primary structure of the alanine carrier protein of thermophilic bacterium PS3.

Purified alanine carrier proteins were cleaved into peptides either chemically after solubilization in 1,1,1,3,3,3-hexafluoro-2-propanol or proteolytically with lysylendopeptidase. From the amino acid sequence analyses of these peptides, we synthesized a DNA probe and utilized it for successful cloning of a gene encoding the alanine carrier protein (acp gene). The 5'-flanking region was determined by an inverse polymerase chain reaction, and an open reading frame consisting of 1,335 nucleotides was found. The amino acid sequence deduced from the open reading frame consists of 445 amino acids, and all the partial amino acid sequences determined are included in the sequence. Although the calculated M(r) of 47,803 is significantly larger than the apparent M(r) of 42,500 as reported previously (Hirata, H., Kambe, T., and Kagawa, Y. (1984) J. Biol. Chem. 259, 10653-10656), an in vitro translation experiment revealed that the product of the acp gene migrates at a position coinciding with that of the purified alanine carrier. Hydropathy analysis suggests that the protein contains at least 8 hydrophobic segments presumably spanning membrane. A homology search on a database reveals relatively high scores of homology with either the Escherichia coli melibiose carrier or the human Na+/glucose symporter, particularly in the region from Leu246 to Glu286. Furthermore, the region also reveals low but significant similarities to other Na(+)-coupled symporters.

Amino Acid Sequence↗
PubMed1992

Effect of a dihydropyridine analogue, 2-[benzyl(phenyl)amino]ethyl 1,4-dihydro-2,6-dimethyl-5-(5,5-dimethyl-2-oxo- 1,3,2-dioxaphosphorinan-2-yl)-1-(2-morpholinoethyl)-4-(3-nitrophenyl)-3 -pyridinecarboxylate on reversing in vivo resistance of tumor cells to adriamycin.

A newly synthesized dihydropyridine analogue, 2-[benzyl(phenyl)-amino]ethyl 1,4-dihydro-2,6-dimethyl-5-(5,5-dimethyl-2-oxo-1,3,2-dioxaphosphorina n-2-yl)-1- (2-morpholinoethyl)-4-(3-nitrophenyl)-3-pyridinecarboxylate (PAK-200), at 5 microM inhibited the efflux of [3H]vincristine from KB-C2 cells and increased the accumulation of [3H]vincristine in KB-C2 cells to a level similar to that in KB-3-1 cells. PAK-200 inhibited the photoaffinity labeling of P-glycoprotein in KB-C2 membranes by [3H]azidopine. At 5 microM, PAK-200 enhanced the cytotoxic effect of Adriamycin on drug-sensitive KB-3-1 cells, multidrug-resistant KB-8-5 cells, and two human colorectal carcinoma tumor lines, COK-28LN and COK-36LN, by factors of 2, 5, 2, and 3 times, respectively. The calcium antagonistic activity of PAK-200 was about 1000 and 5 times lower than that of another dihydropyridine analogue, nicardipine, and of verapamil, respectively. PAK-200 in combination with Adriamycin completely suppressed the growth of KB-3-1 and COK-36LN and partially suppressed the growth of KB-8-5 but had no significant effect on COK-28LN cells xenografted in nude mice. The level of MDR1 expression of COK-36LN was about 3 times higher than that of COK-28LN, but lower than that of KB-8-5 cells. These results suggest that the interaction of PAK-200 with P-glycoprotein may be partly correlated with the enhancement of the antitumor effect of Adriamycin on xenografted KB-8-5 and COK-36LN cells in nude mice.

Animals↗
PubMed1992

Upstream region of a genomic gene for human mitochondrial transcription factor 1.

A genomic gene for human mitochondrial transcription factor 1 was cloned from a human genomic library and its 5' flanking region was sequenced. No typical TATA and three consensus sequences for potential Sp1 binding site were found in its 5' flanking region of 2 kilobase pairs. There were, at least, four common sequences among some nuclear genes for mitochondrial-related proteins.

Amino Acid Sequence↗
PubMed1992

Angiogenic factor.

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Escherichia coli↗
PubMed1992

Induction of synthesis of manganous superoxide dismutase in L-M(pNTnF) cells carrying an inducible TNF gene.

Based on findings that the cytotoxic effects of tumor necrosis factor (TNF) are closely related to levels of intracellular oxygen radicals, and on the results of TNF gene transfection studies, the hypothesis was made that endogenous TNF (enTNF) acts as a protective factor against exogenous TNF by inducing inhibitors or scavengers of oxygen radicals. In order to test this hypothesis, we investigated the intracellular levels of manganous superoxide dismutase (MnSOD) and glutathione (GSH) in L-M(pNTnF) cells carrying a TNF gene induced by dexamethasone (DM). When L-M(pNTnF) cells were treated with DM they expressed enTNF, and acquired resistance to exogenous TNF. There was no change in the GSH concentration after enTNF induction, but a 1.9- to 3.9-fold increase in MnSOD levels was noted. Our findings suggest that enTNF exerts its protective function against the cytocidal effect of exogenous TNF by inducing MnSOD production.

Animals↗
PubMed1992

Enhancement of lysosomal enzyme activity by recombinant human tumor necrosis factor and its role in tumor cell killing in vitro.

We investigated the effect of recombinant human tumor necrosis factor (TNF) on the lysosomal enzyme activity of various established cell lines in vitro. Incubation of 1 x 10(6) TNF-sensitive mouse tumorigenic fibroblasts (L-M cells) in the presence of TNF (100 U/ml) for 48 h increased the total (the sum of the enzyme activities in the lysosomes and the cytoplasm) acid phosphatase and beta-glucuronidase activities by 3.7- and 4.2-fold, respectively. The same increase was observed even when 1 U/ml of TNF was added to some cultures and no further augmentation occurred at 10 or 100 U/ml. Measurement of total and free enzyme activities showed that TNF stimulation not only enhanced the total intracellular enzyme activity but also accelerated the conversion into free (cytoplasmic) enzyme activity. Addition of a lysosomotropic agent (methylamine) suppressed both the enhancement of lysosomal enzyme activity and the cytotoxicity of TNF. A similar enhancement of lysosomal enzyme activities was also detected in various TNF-sensitive tumor cell lines, and a strong correlation (acid phosphatase: r = 0.836, beta-glucuronidase: r = 0.910) was observed between the enhancement of enzyme activity and sensitivity to TNF. No such increase was detected in TNF-resistant human diploid cells. These results show that TNF induces the activation and release of lysosomal enzymes in TNF-sensitive cells, and suggest that such events may play an important role in TNF-mediated cytotoxicity.

Acid Phosphatase↗
PubMed1992

Mechanism of synergistic cytotoxic effect between tumor necrosis factor and hyperthermia.

We previously reported that recombinant human tumor necrosis factor (rhTNF) and hyperthermia had a synergistic effect against tumors, in vitro and in vivo. We have now investigated the mechanism of this synergy by measuring the lysosomal enzyme activity and hydroxyl radical production of L-M cells treated with rhTNF and/or hyperthermia. A synergistic activation of lysosomal enzyme and the induction of hydroxyl radical production in L-M cells treated with both rhTNF and hyperthermia was observed. A synergistic cytotoxic effect was observed when rhTNF and hyperthermia were combined, and was inhibited by the addition of a reactive oxygen scavenger, dimethyl sulfoxide or bipyridine. The results show that the augmenting effect of hyperthermia on lysosomal enzyme activation and induction of hydroxyl radical production by rhTNF plays an important role in the synergistic cytotoxic effect.

Acid Phosphatase↗
PubMed1992

Polyoxyethylene-modified superoxide dismutase reduces side effects of adriamycin and mitomycin C.

Polyoxyethylene-modified superoxide dismutase (SOD-POE) is a newly developed long-acting superoxide dismutase. Adriamycin (ADR) and mitomycin C (MMC) generate superoxide, which contributes to their cytocidal effects or side effects. We examined whether SOD-POE could prevent the side effects induced by superoxide generated by antitumor agents, and the following results were obtained. SOD-POE did not influence the antitumor effects of ADR and MMC either in vitro or in vivo, but prevented the toxic death of BALB/c, nu/nu male mice caused by overdoses of ADR or MMC. As for its effective sites, SOD-POE prevented a decrease in the specific activity of rotenone-sensitive NADH-ubiquinone oxido-reductase (complex I) in heart muscle mitochondrial respiratory chain function in BALB/c male mice administered 10 mg/kg ADR, and prevented damage to the sarcoplasmic reticulum and mitochondria of mouse heart muscle by ADR as observed by electron microscopy. Furthermore, SOD-POE prevented bone marrow suppression induced by MMC in Donryu rats. The above results suggest that combination chemotherapy with SOD-POE would make it possible to increase the maximum permissible doses of antitumor agents, improving the efficacy of these agents.

Animals↗
PubMed1992

Potentiation of the vincristine effect on P388 mouse leukemia cells by a newly synthesized dihydropyridine analogue, PAK-200.

A newly synthesized dihydropyridine analogue, 2-[benzyl(phenyl)amino]ethyl 1,4-dihydro-2,6-dimethyl-5-(5,5-dimethyl-2-oxo-1,3,2-dioxaphosphorina n-2-yl)-1- (2-morpholinoethyl)-4-(3-nitrophenyl)-3-pyridinecarboxylate (PAK-200), at 1 microM completely reversed the resistance to vincristine in vincristine-resistant P388 mouse leukemia cells (P388/VCR), in vitro. PAK-200 at 2 microM inhibited the efflux of [3H]vincristine from P388/VCR and increased the accumulation of [3H]vincristine in P388/VCR to a level similar to that in P388 cells. P-Glycoprotein in membrane vesicles from P388/VCR cells was photolabeled with [3H]azidopine. The labeling was completely inhibited by 10 microM PAK-200. The calcium antagonistic activity of PAK-200 was about 1000 times lower than that of another dihydropyridine analogue, nicardipine. Experiments with P388 and P388/VCR-bearing mice showed that PAK-200 enhanced the effect of vincristine on both leukemia cells in vivo. These results suggest that PAK-200 interacts with P-glycoprotein and reverses drug resistance in P388 mouse leukemia cells in vitro, and that PAK-200 has an ability to potentiate the effect of vincristine on P388 mouse leukemia cells in vivo.

Animals↗
PubMed1992

Gas phase derivatization of ammonia with 4-fluoro-7-nitrobenzo-2-oxa-1,3-diazole and its application to urease assay.

An ammonia-specific and rapid fluorometric method for determination of ammonia and urease activity was developed. The method is designed to assay ammonia levels or urease activity for the rapid diagnosis of Helicobacter pylori infection. 4-Fluoro-7-nitrobenzo-2-oxa-1,3-diazole was used to derivatize ammonia and 4-amino-7-nitrobenzo-2-oxa-1,3-diazole was analysed by high performance liquid chromatography at an excitation wavelength of 455 nm and an emission wavelength of 520 nm. Derivatization was designed to react with ammonia gas produced in a strong alkaline pH sample. The fluorescent intensity was linear in the range of 0.1-10 mM ammonia per tube when the reaction was carried out for 15 min at 37 degrees C. Urease activity, judged as the amount of ammonia production from urea, could be measured at 25 ng per tube (S/N = 1.5) with Jack bean meal urease. Because of its rapidity, this assay is potentially superior to the current standard method in use in clinical settings.

4-Chloro-7-nitrobenzofurazan↗
PubMed1992

Serum tumor necrosis factor activity in inflammatory bowel disease.

Serum tumor necrosis factor (TNF) levels in 33 patients with inflammatory bowel disease (IBD) were measured by using a sensitive enzyme immunoassay. Four of five Crohn's diseases (CD) and nine of twenty eight ulcerative colitis (UC) had elevated levels of serum TNF. In active CD or UC, a greater fraction of patients studied had significantly increased serum TNF levels (3/3 for CD and 8/11 for UC). Production of TNF by peripheral blood monocytes when stimulated by lipopolysaccharide was also increased in these patients and correlated with their serum TNF levels. These results suggest that TNF may have some pathoetiological meaning in IBD.

Adult↗
PubMed1992

[Chronic myelogenous leukemia with blastic crisis in which expression of P-glycoprotein was associated with resistance to chemotherapy].

A 55-year-old woman with chronic myelogenous leukemia developed a lymphoid blast crisis (BC) 10 months after diagnosis. By using immunoblotting with a monoclonal antibody against P-glycoprotein (P-gp) C219, her leukemia cells from the first and 3rd crises were shown to be negative for the P-gp, while the cells of the 4th crisis were detected to have a high level of P-gp. This patient did not respond to chemotherapy with several anti-cancer agents in the 4th crisis, although complete remission was achieved in the first, second and third crises after administration of agents including vincristine and prednisolone. Therefore the expression of P-gp in the 4th BC might have been closely related to the resistance to chemotherapy.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1992

[Prophylactic use of antibiotics for fever following fiberoptic bronchoscopy and bronchography].

To evaluate the effect of bacampicillin hydrochloride on fever following fiberoptic bronchoscopy and bronchography, we conducted multi-institutional randomized study. In bronchographic examinations, the rise of body temperatures in the bacampicillin group (0.82 +/- 0.13 degrees C: mean +/- SE) was significantly smaller than that in the control group (1.39 +/- 0.25 degrees C) on the second day of examination. Bacterial infection may contribute to the rise of temperature on the day following bronchography, but no pneumonia or sepsis was observed. There was no differences in the rise of body temperature on the first, third or fourth day. In fiberoptic bronchoscopic examinations, there was no difference between the two groups. We conclude that there is no clinical indication of the value of prophylactic use of antibiotics in either fiberoptic bronchoscopy or bronchography.

Adult↗
PubMed1991

Precolumn fluorescence tagging reagent for carboxylic acids in high-performance liquid chromatography: 4-substituted-7-aminoalkylamino-2,1,3-benzoxadiazoles.

Four new 2,1,3-benzoxadiazole amine reagents having different functional groups at the 4- and 7-positions, [4-nitro-7-N-piperazino-2,1,3-benzoxadiazole (NBD-PZ), 4-(N,N-dimethylaminosulphonyl)-7-N-piperazino-2,1,3-benzoxad iazole (DBD-PZ), 4-(N,N-dimethylaminosulphonyl)-7-N-cadaverino-2,1,3-benzoxad iazole (DBD-CD) and ammonium 7-N-piperazino-2,1,3-benzoxadiazole-4-sulphonate (SBD-PZ)] were synthesized as fluorogenic tagging reagents for carboxylic acids in high-performance liquid chromatography. The reagents, except SBD-PZ, reacted with carboxylic acid at room temperature in the presence of activation agents to produce fluorescent adducts. The maximum wavelengths of arachidic acid tagged with DBD-PZ, DBD-CD and NBD-PZ were 569 nm (excitation, 440 nm), 561 nm (excitation, 437 nm) and 541 nm (excitation, 470 nm), respectively. Among various activation agents tested [diethyl phosphorocyanidate (DEPC), diphenyl phosphoroyl azide (DPPA), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC)-pyridine, 2.2'-dipyridyl disulphide-triphenylphosphine (Mukaiyama A) and 2-chloro-1-methylpyridinium iodide-triethylamine (Mukaiyama B)], DEPC and Mukaiyama A were more effective than the others. When the piperazino reagents (DBD-PZ and NBD-PZ) were used as the tagging reagents, the derivatization reaction in the presence of Mukaiyama A was faster than that in the presence of DEPC. Although the reaction in the presence of Mukaiyama A was completed after 30 min, an unknown peak derived from the activation agent appeared on the chromatograms. The fluorescence peak intensities were compared in the presence of DEPC. The order of the fluorescence peak areas obtained after reaction for 6 h in the presence of DEPC was DBD-PZ greater than DBD-CD greater than NBD-PZ. Thirteen saturated free fatty acids (FFAs) derivatized with DBD-PZ (or DBD-CD) and DEPC (or Mukaiyama A) in acetonitrile were separated completely by linear gradient elution on a reversed-phase ODS column. Eight drugs (ibuprofen, indomethacin, dinoprost, prostaglandin E1, dehydrocholic acid, ursodesoxycholic acid, hydrocartisone succinate and prednisolone succinate) were also tagged with DBD-PZ in the presence of DEPC and separated by isocratic elution. The detection limits (signal-to-noise ratio = 3) of FFAs tagged with DBD-PZ were in the range 3.2-4.7 fmol, whereas those of drugs were in the range 3.9-14 fmol.

Anti-Inflammatory Agents↗