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Biomedical subjects

T Tsuchiya

Publications and source records attributed to T Tsuchiya.

At least 343 records · Page 19Linked to original sources
PubMed1994

Assessment of contact sensitivity of four thiourea rubber accelerators: comparison of two mouse lymph node assays with the guinea pig maximization test.

Contact sensitivity of four thiourea rubber accelerators, diphenylthiourea (DPTU), dilaurylthiourea (DLTU), dibutylthiourea (DBTU) and diethylthiourea (DETU), was evaluated by a new sensitive mouse lymph node assay (SLNA) and the murine local lymph node assay (LLNA). The results of the SLNA and LLNA were compared with the data of the guinea pig maximization test (GPMT). In the LLNA and SLNA, the sensitizing activity was measured as a function of draining lymph node activation following application of the test chemicals. Of these four thioureas, three (DETU, DBTU and DPTU) were not classified as skin sensitizers in the LLNA. The SLNA successfully detected the sensitivity of all thioureas tested. This result indicated that the SLNA was, in these cases, more sensitive than the LLNA for identification of contact allergens. The order of sensitization potential observed from the SLNA was DPTU (greatest), DLTU, DBTU and then DETU (least). The predictions of sensitizing potential and the order of the sensitizing capacity of four thioureas by the SLNA and the GPMT are very similar.

Administration, Topical↗
PubMed1994

Contact sensitivity of and cross-sensitivity between 2-(2'-hydroxy-5'-methylphenyl)benzotriazole (Tinuvin P) and 2-(2'-hydroxy-3'-tert-butyl-5'-methylphenyl)-5-chlorobenzotriazole (Tinuvin 326) evaluated by lymph node cell proliferation and ear swelling response in mice.

Contact sensitivity of and cross-sensitivity between 2-(2'-hydroxy-5'-methylphenyl)benzotriazole (Tinuvin P, TP) and 2-(2'-hydroxy-3'-tert-butyl-5'-methylphenyl)-5-chlorobenzotriazole (Tinuvin 326, T326) were investigated in BALB/c mice. Mice received an intradermal injection and subsequent topical application of the test chemical. Following final application, auricular lymph node cell (LNC) proliferation was measured. In another experiment, the appearance of contact sensitivity was assessed by increases in ear thickness following challenge of the test chemical. Application of TP to mice caused a significant increase in LNC proliferation. Challenge of these mice with TP resulted in an increase in ear thickness. However, TP-sensitized mice did not show an increase in ear thickness by challenge with T326. Under the same conditions, application of T326 failed to induce either LNC proliferation or ear swelling response. The induction of LNC proliferative response was closely related to the ear swelling response by challenge. Following intradermal injection of TP, topical application of T326 to mice did not induce LNC proliferation. A significant ear swelling response was not induced by first challenge with TP or T326 but induced by rechallenge with TP. Topical exposure of mice to TP following injection of T326 failed to induce LNC proliferation or any ear responses by challenge with T326 and TP. These data suggest that TP is a sensitizer, while T326 is not a sensitizer, and these chemicals do not cross-sensitize.

Administration, Topical↗
PubMed1994

Renal and hormonal responses to repeated treatment with enalapril in non-azotemic cirrhosis with ascites.

Since a single dose of the angiotensin-converting enzyme inhibitor enalapril was shown to cause natriuresis in cirrhosis in a previous study, we investigated whether repeated doses of this substance would sustain a favorable renal effect in cirrhosis. Ten milligrams of enalapril maleate were administered once a day for 8 days to ten patients with non-azotemic cirrhosis and ascites. Enalapril reduced blood pressure significantly at 4 to 12 h (systolic blood pressure) and 2, 6, and 8 h (diastolic blood pressure) on day 2, compared to pretreatment (day 0) values, but this depressor effect decreased on day 8. No change in heart rate could be detected. Enalapril significantly suppressed serum angiotensin-converting enzyme activity and plasma aldosterone concentration (p < 0.001 to 0.01), which were elevated prior to treatment, with pretreatment values of 25.8 +/- 1.8 IU/l for serum angiotensin-converting enzyme activity and 241 +/- 67 pg/ml for plasma aldosterone concentration. This drug caused a 12 to 24% increase (p < 0.05 to 0.01) in mean daily urinary volume and a 40 to 54% increase (p < 0.001 to 0.01) in mean daily urinary sodium excretion from the respective pretreatment baselines during the 8-day period. Creatinine clearance was improved (p < 0.05) by the treatment, with mean improvement values from 24 to 34% above the pretreatment value of 47.4 +/- 4.3 ml/min.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1994

Enhancement of serine-sensitivity by a gene encoding rhodanese-like protein in Escherichia coli.

When cells of Escherichia coli are grown on lactate (or other carbon sources), an addition of serine to the medium causes growth inhibition. This growth inhibition is caused by inhibition by serine of homoserine dehydrogenase I, which is involved in threonine-isoleucine biosynthesis [Hama, H., Sumita, Y., Kakutani, Y., Tsuda, M., & Tsuchiya, T. (1990) Biochem. Biophys. Res. Commun. 168, 1211-1216]. We have cloned and sequenced genes which enhance the serine-sensitivity. Two open reading frames were found and designated as sseA and sseB. Introduction of either sseA or sseB gene, or both, into E. coli cells enhanced the serine-sensitivity. The sseA gene elicited stronger enhancement than sseB. The deduced amino acid sequence of SseA showed considerable similarity with that of bovine liver rhodanese, which catalyzes sulfur transfer from thiosulfate. We observed a twofold increase in rhodanese activity in E. coli cells harboring a plasmid carrying the sseA gene. The position of sseA in the genetic map is around 52'. However, sseA is different from cysM, which codes for O-acetylserine sulfhydrylase-B, an enzyme catalyzing sulfur transfer from thiosulfate to O-acetylserine, the map position of which is also around 52'.

Amino Acid Sequence↗
PubMed1994

Properties of the Na+/H+ antiporter in Vibrio parahaemolyticus.

The properties of the Na+/H+ antiporter in Vibrio parahaemolyticus, a slightly halophilic bacterium, were investigated using everted membrane vesicles. It appears that at least two Na+/H+ antiporters are present, one that is pH-dependent and one that is pH-independent. These two antiporters appear to correspond to the NhaA and NhaB antiporters of Escherichia coli, respectively. It seems that amiloride strongly inhibits the pH-dependent antiporter. Na+ is the best substrate for both of the two V. parahaemolyticus antiporters. Li+ is a poorer substrate and K+ is not a substrate. No K+/H+ antiport activity was detected in membrane vesicles of this organism. The Na+(Li+)/H+ antiport activity greatly increased with an increase in pH of the assay medium. pH did not affect the Km value of the Na+/H+ antiport, but it did increase the Vmax.

Hydrogen-Ion Concentration↗
PubMed1994

Calpain-calpastatin interactions in epidermoid carcinoma KB cells.

We examined the activation of mu-calpain in human epidermoid carcinoma KB cells following a rise in intracellular calcium concentration using antibodies specifically recognizing different activation states of mu-calpain. KB cells possess calpastatin activity in large excess of calpain activity as analyzed by ion exchange HPLC. Stimulation of the cells with a calcium ionophore, ionomycin, caused production of the autolytic intermediate form (M(r) = 78 k) of mu-calpain derived from the preautolysis form (80 k), while the fully autolyzed postautolysis form (76 k) remained below detectable levels at all times. The appearance of the autolytic intermediate paralleled limited proteolysis of the membrane-associated calpastatin fractions (110 k and 106 k); the resulting fragments (68 k and 45 k) were released into the cytosol. Both the production of the autolytic mu-calpain intermediate and the limited proteolysis of calpastatin in cell lysates in the presence of calcium were inhibited by a synthetic calpastatin peptide, indicating that proteolysis of calpastatin was indeed catalyzed by calpain and that the autolytic intermediate may have exerted the proteolytic activity. Furthermore, mu-calpain autolysis and calpastatin degradation, upon ionomycin treatment, were both augmented by epidermal growth factor (EGF). These results suggest that calpastatin serves not only as an inhibitor but also as a substrate for calpain at cell membranes and that intracellular conditions associated with the cell cycle may affect the activation of mu-calpain.

Amino Acid Sequence↗
PubMed1994

Transcriptional attenuation and differential mRNA stability in the regulation of the Escherichia coli melibiose operon.

The organization of the melibiose operon of Escherichia coli is promoter-melA-melB. The amount of the product (alpha-galactosidase) of the first gene (melA) is much larger than that of the product (melibiose permease) of the second gene (melB). Using the chloramphenicol acetyltransferase gene (cat) as reporter, we found that there was an element between melA and melB, which reduced the expression of the downstream gene, melB. This region contained a boxA-like sequence, which is known as a binding site for an attenuation factor, NusA. Northern hybridization analysis revealed that the ratio of melA mRNA and melAB mRNA was comparable with the ratio of the melA and melB products. We also found that the melA mRNA was about 3-fold more stable than the melAB mRNA. Experimental results obtained with a nusAts mutant suggested that the NusA protein is involved in the reduced expression of the melB gene. We conclude that the production ratio of alpha-galactosidase and melibiose permease is regulated at two levels: 1) transcription and 2) mRNA stability.

Base Sequence↗
PubMed1994

The NhaB Na+/H+ antiporter is essential for intracellular pH regulation under alkaline conditions in Escherichia coli.

We isolated a mutant of Escherichia coli which was defective in an Na+/H+ antiporter and grew poorly under alkaline conditions [Ishikawa, T., Hama, H., Tsuda, T., and Tsuchiya, T. (1987) J. Biol. Chem. 262, 7443-7446]. Later, it was concluded that the defective Na+/H+ antiporter in the mutant was the NhaB system, and the nhaB gene was mapped to 25.6 min on the E. coli chromosome [Thelen, P., Tsuchiya, T., and Goldberg, E.B. (1991) J. Bacteriol. 173, 6553-6557]. We found that the NhaB-defective cells cannot grow in a high pH medium. Furthermore, intracellular pH in the mutant cells was almost the same as extracellular pH between 7.9 and 9.1, that is, intracellular pH was not regulated at this pH range. On the other hand, intracellular pH of the wild-type cells was maintained at about 7.6 when the extracellular pH was between 7.6 and 8.5. Thus, the NhaB Na+/H+ antiporter is essential for the regulation of intracellular pH under alkaline conditions in E. coli. Introduction of nhaA gene into the mutant cells increased Na+/H+ antiporter activity, but did not restore the defective growth and defective intracellular pH regulation under alkaline conditions.

Bacterial Proteins↗
PubMed1994

Properties and sequence of the NhaA Na+/H+ antiporter of Vibrio parahaemolyticus.

A gene encoding an Na+/H+ antiporter was cloned from chromosomal DNA of the slightly halophilic marine bacterium Vibrio parahaemolyticus. The host was an Escherichia coli mutant that lacked both of the two major Na+/H+ antiporters, NhaA and NhaB. Untransformed mutant cells were unable to grow in the presence of 0.6 M NaCl or 0.1 M LiCl, but Na+ and Li+ were non-toxic to cells transformed with a plasmid carrying the antiporter gene. Membrane vesicles prepared from the original E. coli mutant did not show any detectable Na+/H+ (and Li+/H+) antiport activity. However, we observed high Na+/H+ (and Li+/H+) antiport activity in membrane vesicles prepared from the transformed cells. The activity increased greatly when the pH of the assay medium was increased from 7.0 and 8.5. This property is very similar to that of the NhaA Na+/H+ antiporter of E. coli. Drastic decreases in Km values for Li+ and Na+ were observed with membrane vesicles prepared from the transformed cells compared with those observed with V. parahaemolyticus vesicles. The amino acid sequence deduced from the nucleotide sequence of the cloned gene showed high homology (59% identity and 87% similarity) with the NhaA Na+/H+ antiporter of E. coli. Thus, we conclude that the gene we cloned and sequenced is the nhaA of V. parahaemolyticus. We also found that several regions of the NhaA protein showed sequence similarity with transport proteins from some other organisms. Such regions seem to be important for Na+ recognition, transport or amiloride binding.

Amino Acid Sequence↗
PubMed1994

Inhibitory effect of calcium channel blockers on growth of pancreatic cancer cells.

Calcium, which binds to calmodulin inside the cells, is an important mediator of various intracellular processes, including cell proliferation. We speculated that blockade of Ca2+ influx into the cells by Ca(2+)-channel blockers, such as phenytoin and verapamil, might affect the Ca(2+)-calmodulin pathway leading to suppression of cell growth. In this study, we examined the effect of phenytoin and verapamil on growth of two human pancreatic cancer cell lines, MIA PaCa-2 and CAV, in vitro and in vivo. Both phenytoin and verapamil inhibited growth of the two cell lines in a dose-dependent fashion. Phenytoin and verapamil each significantly prolonged doubling time of MIA PaCa-2 and the combination of the two drugs acted synergistically. The activity of ornithine decarboxylase, which is a rate-limiting enzyme of the polyamine pathway that is closely related to cell proliferation, was significantly inhibited by both drugs in a time-dependent fashion. Phenytoin, but not verapamil, inhibited growth of MIA PaCa-2 tumors xenotransplanted into nude mice, whereas both phenytoin and verapamil inhibited the growth of CAV tumors. Since phenytoin and verapamil are known to have fewer side effects than conventional antineoplastic drugs, these results suggest their possible use in novel therapeutic strategies.

Calcium Channel Blockers↗
PubMed1994

Demonstration of entrainment and presence of slow conduction during ventricular tachycardia in arrhythmogenic right ventricular dysplasia.

During VT in two cases with arrhythmogenic right ventricular dysplasia, entrainment criteria, constant fusion beats except for the last entrainment beat, progressive fusion, and a localized conduction block associated with interruption of VT, were demonstrated with rapid ventricular pacing performed during VT. Furthermore, a long conduction interval was present during entrainment from the pacing site to the earliest activation site during VT, indicating the presence of a slow conduction area. VT in these cases was, thus, due to reentry with an area of slow conduction within the circuit.

Action Potentials↗
PubMed1994

Contact sensitivity to Tinuvin P in mice.

The contact sensitizing potential of 2-(2'-hydroxy-5'-methylphenyl)benzotriazole (Tinuvin P) was investigated using the murine local lymph node assay (LLNA) and the mouse ear swelling test with some modification in the induction procedure (modified MEST). Topical exposure of the ears to Tinuvin P in acetone:olive oil (AOO) for 3 consecutive days induced no increase in auricular lymph node weight and lymph node cell proliferation. In the LLNA, Tinuvin P was negative, and its sensitizing potential was determined as not strong. In the modified MEST, mice were sensitized with a combination of an intradermal injection of 0.2% Tinuvin P in Freund's complete adjuvant into the shaved abdomen and topical application of 1% Tinuvin P in AOO on both ears for 3 consecutive days. The ear thickness in mice previously treated with Tinuvin P increased significantly on challenge with Tinuvin P. A cross-reaction test between Tinuvin P and various substituted benzotriazole ultraviolet light (UV) absorbers was performed. Challenge with these benzotriazole compounds on the ears of the mice previously sensitized to Tinuvin P showed no increase in ear thickness. These data indicated that Tinuvin P had a contact sensitizing ability, while Tinuvin P-sensitized mice did not cross-react with the other benzotriazole UV absorbers.

Animals↗
PubMed1994

Characterization of a glucose transport system in Vibrio parahaemolyticus.

Cells of a glucose-PTS (phosphoenolpyruvate:carbohydrate phosphotransferase system)-negative mutant of Vibrio parahaemolyticus transport D-glucose in the presence of Na+. Maximum stimulation of D-glucose transport was observed at 40 mM NaCl, and Na+ could be replaced partially with Li+. Addition of D-glucose to the cell suspension under anaerobic conditions elicited Na+ uptake. Thus, we conclude that glucose is transported by a Na+/glucose symport mechanism. Calculated Vmax and Km values for the Na(+)-dependent D-glucose transport were 15 nmol/min/mg of protein and 0.57 mM, respectively, when NaCl was added at 40 mM. Na+ lowered the Km value without affecting the Vmax value. D-Glucose was the best substrate for this transport system, followed by galactose, alpha-D-fucose, and methyl-alpha-glucoside, judging from the inhibition pattern of the glucose transport. D-Glucose itself partly repressed the transport system when cells were grown in its presence.

Biological Transport↗
PubMed1994

Age-dependent increase of neurotensin expression in the ileum.

The proliferative activity of gut mucosa is increased with aging. Neurotensin (NT), a tridecapeptide localized to the distal small bowel, stimulates growth of gut mucosa; the effects of aging on gut NT are not known. Young (4-mo-old), adult (1-yr-old), and aged (2-yr-old) male Fischer 344 rats were killed; the ileal mucosa was scraped, weighed, and extracted for measurement of NT mRNA by slot-blot and Northern hybridization; and the relative NT transcription rate was determined by a nuclear run-on assay. In addition, NT tissue content and plasma levels were determined by radioimmunoassay, and full-thickness sections of ileum were examined for age-dependent alterations of NT endocrine cell (N-cell) number using immunohistochemical staining. Slot-blot and Northern-blot analyses showed that the steady-state levels of NT mRNA were increased threefold in the adult group and eightfold in the aged rats. In addition, NT peptide content and plasma levels were significantly increased in the aged group. The dramatic increases in the abundance of NT mRNA were not associated with increases in either NT transcription or N-cell number. In summary, we have demonstrated an age-dependent increase in the constitutive levels of ileal NT mRNA that appears not to be due to concomitant increases in transcription, suggesting that NT mRNA is increased by mechanisms involving mainly a posttranscriptional process. In addition, we have shown corresponding increases in the levels of both tissue and plasma NT with aging, indicating that expression of NT does not remain the same throughout life but actually increases with aging in the rat.

Aging↗
PubMed1994

Ultrasonographic evaluation of vertebral artery to detect vertebrobasilar axis occlusion.

BACKGROUND AND PURPOSE: We performed the present study to determine whether the site of occlusion in the vertebrobasilar circulation could be identified by the measurement of blood flow velocity in the bilateral vertebral arteries. METHODS: Using color-coded duplex carotid ultrasonography, we measured the time-averaged mean and end-diastolic flow velocities with incident angle correction in 130 bilateral vertebral arteries between the C3 and C6 segments of the spine in 65 stroke patients with or without occlusive diseases in the vertebrobasilar circulation. The site of occlusion was confirmed by cerebral angiography. The subjects included 12 patients with unilateral subclavian artery occlusion (SA group), 11 patients with unilateral occlusion at the origin of the vertebral artery (V1 group), 6 patients with unilateral vertebral artery occlusion before the branching of the posterior inferior cerebellar artery (V2 group), 14 patients with unilateral vertebral artery occlusion after the branching of the posterior inferior cerebellar artery (V3 group), 5 patients with basilar artery occlusion (BA group), 5 patients with unilateral posterior cerebral artery occlusion (PCA group), and 12 patients without any occlusive lesions in the vertebrobasilar circulation (control group). RESULTS: In the control group the mean and end-diastolic blood flow velocities were 25.5 +/- 6.9 cm/s and 16.2 +/- 4.3 cm/s, respectively, and the side-to-side differences of these velocities were 4.8 +/- 5.2 cm/s and 4.7 +/- 4.1 cm/s, respectively. All patients in the SA group demonstrated retrograde flow on the affected side. In the V1 group no flow signals were detected on the occluded side. In the V2 group the mean velocity (7.2 +/- 3.1 cm/s) was lower than in the control group, and the end-diastolic velocity was zero on the affected side. In the V3 group the mean and end-diastolic velocities (11.5 +/- 3.1 cm/s and 5.9 +/- 2.8 cm/s, respectively) on the occluded side were lower than in the control group. Flow velocities on the unaffected side were higher than those on the affected side in the SA, V1, V2, and V3 groups. However, there were no differences in flow velocity between the control, BA, and PCA groups. CONCLUSIONS: Measurement of vertebral artery blood flow velocity may help in localizing the site of occlusion in the subclavian and vertebral arteries.

Aged↗
PubMed1994

Reopening phenomenon in cardioembolic stroke observed by duplex carotid ultrasonography. Case reports.

The authors attempted to detect the reopening of an occluded artery within the carotid axis of acute cardioembolic stroke patients noninvasively by using duplex carotid ultrasonography. The timing of reopening as related to hemorrhagic transformation was also studied. For assessing the reopening phenomenon, quantitative measurement of flow velocity in the bilateral common carotid arteries was repeated in 10 patients who also underwent repeated angiography or autopsy. The ratio of the end-diastolic flow velocity (nonaffected/affected) was related to angiographic or autopsy findings. In 17 patients including the 10 cases mentioned, the occurrence of hemorrhagic transformation was related to the timing of reopening. The hemorrhagic infarction in this study was defined to be a hyperdense area in the basal ganglia on computed tomography. A significant decrease in the ratio of the end-diastolic velocity was observed in 6 patients, in whom the repeated angiography demonstrated reopening of the artery. Of the remaining 4 patients not showing a significant change, only 1 patient was found to have reopening. Reopening was detected in 12 of 17 cases. The timing was 2.5 hours after onset in 1, between days 2 and 5 in 6 cases, and after day 6 in 5 cases. Hemorrhagic infarction was found in 5 of the cases with reopening on days 2-5.

Aged↗
PubMed1994

Studies on the standardization of cytotoxicity tests and new standard reference materials useful for evaluating the safety of biomaterials.

Standard reference materials (SRM) made of polyurethane films containing various amounts of cytotoxic compounds such as zinc diethyldithiocarbamate (ZDEC) and zinc dibutyldithiocarbamate (ZDBC) are proposed. The cytotoxicity indices of five dithiocarbamates and the SRM films obtained by colony assay were compared with those by published standards, i.e., neutral red assay (NF S 90-702), agar diffusion assay (USP XXII), and millipore filter diffusion assay (DIN-V 13 930). Among them, colony assay using 3 cell lines of Balb 3T3, L929 and V79 was found to be most sensitive. The extraction method and direct contact method were proposed as two test methods of colony assay. In the former, culture media after incubation with test materials were used as test solution, and in the latter, cells were directly cultured on test materials. The extraction method gave a linear relationship between the cytotoxic potentials of the SRM and its concentration of ZDEC or ZDBC. The cytotoxic potentials of the SRM correlated well with the thickness of inflammatory layer in rabbit muscle implantation tests of the SRM. The direct contact method was found to be useful to detect weak cytotoxicity because of its high sensitivity. Based on these data, we propose the colony assay and the two kinds of new SRMs as Japanese standard test method for evaluating the cytotoxicity of biomaterials.

Animals↗