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Biomedical subjects

K Hirota

Publications and source records attributed to K Hirota.

At least 217 records · Page 12Linked to original sources
PubMed1996

Redox control of resistance to cis-diamminedichloroplatinum (II) (CDDP): protective effect of human thioredoxin against CDDP-induced cytotoxicity.

Thioredoxin is a small ubiquitous protein with multiple biological functions, including cellular defense mechanisms against oxidative stress. In the present study, we investigated the role of human thioredoxin (hTRX) in the acquisition of cellular resistance to cis-diamminedichloroplatinum (II) (CDDP). The expression and activity of hTRX in Jurkat T cells was dose-dependently enhanced by exposure to CDDP, as determined by immunoblot analysis and insulin reducing assay. Furthermore, chloramphenicol acetyltransferase analysis using the hTRX promoter-reporter gene construct revealed that treatment of Jurkat cells with CDDP caused transcriptional activation of the hTRX gene, which might be mediated through increased generation of intracellular reactive oxygen intermediates. To examine the biological significance of hTRX induction, we established hTRX-overexpressing derivatives of L929 fibrosarcoma cells by stable transfection with the hTRX cDNA. The clones, which constitutively expressed the exogenous hTRX, displayed increased resistance to CDDP-induced cytotoxicity, compared with the control clones. After exposure to CDDP, the control cells showed a significant increase in the intracellular accumulation of peroxides, whereas the hTRX-transfected cells did not. Taken together, these results suggest that overexpressed hTRX is responsible for the development of cellular resistance to CDDP, possibly by scavenging intracellular toxic oxidants generated by this anticancer agent.

Animals↗
PubMed1996

Nested-polymerase chain reaction for the detection of Helicobacter pylori infection with novel primers designed by sequence analysis of urease A gene in clinically isolated bacterial strains.

We have established a highly sensitive semi-nested PCR assay for the detection of H. pylori infection using gastric juice samples, which can be aspirated with disposable nasogastric tubes. The primers targeting H. pylori urease A gene were designed based on the sequence conservation analysis of sixteen H. pylori strains isolated from Japanese patients. The efficacy of the PCR assay, designated as the URA-PCR, was confirmed by in vitro and in vivo assessments. Its sensitivity was 97.5% in the gastric juice samples aspirated from forty patients with proven H. pylori infection, and was significantly higher than that obtained with previously described PCR assays.

Base Sequence↗
PubMed1996

SRB1, a class B scavenger receptor, recognizes both negatively charged liposomes and apoptotic cells.

Scavenger receptors mediate the recognition of a wide range of negatively charged macromolecules such as acetylated low-density lipoproteins (AcLDL). We previously reported the isolation of CHO cell variants expressing a novel type of scavenger receptor, distinct from type I and II macrophage scavenger receptors, that recognize both AcLDL and negatively charged liposomes. In the present study, we found that these isolated CHO cells express class B scavenger receptors (SRB1), which have recently been identified as a new member of the CD36 family of membrane proteins. The isolated CHO cells and CHO transfectants constitutively expressing SRB1 recognized and mediated the endocytosis of cells undergoing apoptosis as well as that of negatively charged liposomes. In contrast, CHO cells constitutively expressing type I or II scavenger receptors were unable to take up these ligands.

Animals↗
PubMed1996

Whole saliva dried on filter paper or diagnosis of HIV-1 infection by detection of antibody IgG to HIV-1 with ultrasensitive enzyme immunoassay using recombinant reverse transcriptase as antigen.

Whole saliva samples collected from HIV-1 seropositive subjects by simple spitting without using any devices were dried on filter paper strips, from which filter paper discs of 3-mm diameter were punched out. The eluates of the discs were subjected to the immune complex transfer enzyme immunoassay for antibody IgG to HIV-1 using recombinant reverse transcriptase of HIV-1 as antigen and a two-site enzyme immunoassay for whole IgG. The signals for antibody IgG to HIV-1 and the amounts of whole IgG obtained with one disc per assay tube were 126-290% of those obtained with 1 microliter of whole saliva samples, provided that filter paper strips were treated with nonspecific rabbit serum prior to drying whole saliva samples and that filter paper discs were tested within a few days after drying whole saliva samples. From these results, diagnosis of HIV-1 infection was indicated to be possible with whole saliva samples dried on filter papers, since the diagnosis was previously shown to be possible with 1 microliter of whole saliva samples. The test for HIV-1 infection with whole saliva samples dried on filter papers was suggested to be useful for various purposes.

Acquired Immunodeficiency Syndrome↗
PubMed1996

Droperidol inhibits tracheal contraction induced by serotonin, histamine or carbachol in guinea pigs.

PURPOSE: Droperidol (D) is effective in the treatment of patients with status asthmaticus. It has been reported that D inhibits the bronchoconstriction induced by serotonin (5-HT) but not that by histamine (H) or acetylcholine. However, haloperidol, another butyrophenone, is known to interact with and inhibit calmodulin, an intracellular Ca(++)-binding protein which is important in the contraction of smooth muscles. The present study was designed to investigate the effects of D on tracheal contractions induced by 5-HT, H or carbachol (C) and to determine the contribution of alpha-adrenoceptors to the relaxant effect of D in vitro. METHODS: Tracheas of female guinea pigs were cut spirally into strips and mounted in water-jacketed organ baths in Tyrode's solution, aerated with a mixture of 95% O2 and 5% CO2 at 37 degrees C. The changes in isometric tension induced by each spasmogen in the strips were measured with a transducer and a polygraph. RESULTS: We found that D inhibited the tracheal contractions induced by 5-HT, H or C in a concentration-dependent manner. At 1.25 x 10(-6) M D blocked the effect of 10(-4) M 5-HT by 44.1 +/- 4.3% and at 2.5 x 10(-6) M by 63.8 +/- 3.8%. Similarly, at 5.0 x 10(-6) M concentration, D blocked the effect of 10(-5) M H by 27.7 +/- 5.3% and at 10(-5) M by 56.2 +/- 2.6%. Furthermore, 5 x 10(-6) M of D reduced the contractions produced by 10(-7) M C by 37.1 +/- 3.0% and 10(-5) M of D by 76.1 +/- 3.2%. The inhibiting effect of D was strongest on contractions induced by 5-HT. Prazosin (10(-6) M) affected neither 5-HT-induced contractions nor the inhibition by D. CONCLUSION: Our data indicate that D partially blocks the contractile responses not only to 5-HT, an effect which would be mediated through a blockade of the 5-HT receptors, but also to H or C, probably through inhibition of calmodulin. Our data support previous reports indicating that droperidol may be an important therapeutic agent in the treatment of patients with hyperreactive airways.

Anesthetics, Intravenous↗
PubMed1996

Intraventricular administration of estradiol modulates rat prolactin secretion and synthesis.

The effect of estradiol (E2) on rat tuberoinfundibular dopaminergic (TIDA) neurons was examined in vivo, employing chronic intraventricular (i.c.v.) infusion technique using an osmotic mini-pump. The activity of TIDA neurons was assessed by the release and synthesis of prolactin (PRL) in the rat pituitary gland and by the changes in the 3, 4-dihydroxyphenylacetic acid (DOPAC) and dopamine (DA) levels and in the DOPAC/DA ratio in the rat hypothalamus. We also examined the [3H] E2 binding in the rat hypothalamus. Ovariectomized female Wistar rats with E2 replacement were treated with daily i.c.v. infusion of 1 microM of E2 or saline vehicle for 1, 3, and 7 days using the Alzet osmotic mini-pump and brain infusion kit. At 1 day of i.c.v. infusion of E2, the serum PRL level was significantly decreased compared with that in the vehicle group. Northern blot analysis of the total RNA isolated from the pituitary glands demonstrated a decrease in the PRL gene transcript level in the E2 group. At 3 days of E2 treatment, however, the serum PRL level was significantly increased compared with that of the vehicle-injected group and Northern blot analysis also demonstrated that the PRL gene transcript level was increased in the E2 group. At 7 days of E2 administration, there were no significant differences between the E2 and vehicle groups in either serum PRL or PRL gene transcript levels. There was a significant increase in the DOPAC/DA ratio after 1 day in the E2 group. However, no significant effects of E2 on this ratio were observed at 3 and 7 days of treatment. The DOPAC concentration in the E2 group was significantly increased at day 1 and significantly decreased at day 3, compared with that of the respective time in vehicle group. At day 7 there was no significant change in DOPAC concentration in either groups. The DA concentrations in the hypothalamus was not changed on any day in either group. Specific [3H] E2 binding was observed in the rat hypothalamus. These data suggest that E2 may have a biphasic effect on the accumulation of PRL gene transcripts and on the PRL secretion in the rat pituitary by first stimulating and then inhibiting the TIDA neuronal activity.

3,4-Dihydroxyphenylacetic Acid↗
PubMed1996

A stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system.

Recently we found that cytokine-induced neutrophil chemoattractant influenced anterior pituitary hormone release in vitro. These observations prompted us to investigate the possibility of the existence of cytokine-induced neutrophil chemoattractant in the hypothalamus. Immunohistochemistry showed that cytokine-induced neutrophil chemoattractant-like immunoreactivity existed in the paraventricular hypothalamic nucleus, the supraoptic nucleus, both the internal and the external layers of the median eminence and the posterior pituitary. Since the paraventricular hypothalamic nucleus plays a pivotal role in response to stressful stimuli, we examined the effect of a single episode of immobilization stress on cytokine-induced neutrophil chemoattractant messenger RNA expression in the paraventricular hypothalamic nucleus. Immobilization stress induced strong hybridization signals of cytokine-induced neutrophil chemoattractant messenger RNA in the parvocellular and magnocellular subdivision of the paraventricular hypothalamic nucleus within 15 min, and cytokine-induced neutrophil chemoattractant-like immunostaining intensity in the posterior pituitary started to increase around the periphery of the posterior lobe at 30 min after stress and extended to the whole lobe at 1 h after stress. The increase in the serum cytokine-induced neutrophil chemoattractant in response to stress showed a kinetically biphasic pattern. A first phase occurred within 15 min which may be due to an immediate release of stored cytokine-induced neutrophil chemoattractant in the neurohypophysis, since hypophysectomy completely blocked this phase. A second phase may reflect the release of newly synthesized cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus and/or peripheral cytokine-induced neutrophil chemoattractant, since hypophysectomy could not reduce this phase. These data suggest that cytokine-induced neutrophil chemoattractant in the paraventricular hypothalamic nucleus was immediately synthesized in response to stress, and then released into the peripheral blood via the hypothalamo-neurohypophysial system, revealing the presence of a stress-sensitive chemokinergic neuronal pathway in the hypothalamo-pituitary system.

Animals↗
PubMed1996

Transactivation of an inducible anti-oxidative stress protein, human thioredoxin by HTLV-I Tax.

Adult T-cell leukemia derived factor (ADF)/human thioredoxin (TRX), which has thiol reducing and radical scavenging activities, plays an essential role on cellular protection against oxidative stress and cell death. TRX itself is induced by various oxidative stress as well as the Human T-cell lymphotropic virus type I (HTLV-I) Tax protein. To investigate the mechanism of this induction, the promoter region of the TRX gene was analyzed. Chloramphenicol acetyltransferase (CAT) reporter constructs containing the TRX promoter sequences responded to the overexpression of the Tax protein, whereas various oxidative agents activated the TRX promoter through a newly identified oxidative responsive element. Moreover, TRX was translocated from the cytoplasm into the nucleus by ultraviolet irradiation, suggesting its possible role on sensing and transducing oxidative signals.

Cell Line, Transformed↗
PubMed1996

In vivo direct measurement of the bronchodilating effect of sevoflurane using a superfine fiberoptic bronchoscope: comparison with enflurane and halothane.

OBJECTIVES: Although volatile anesthetics have been shown to have spasmolytic effects on constricted airways, most previous studies were performed to assess bronchodilation with indirect methods such as measurement of airway resistance, whose reliability is controversial. As the authors have developed a new direct method using a superfine fiberoptic bronchoscope (SFB) and confirmed the accuracy of this method, this study assessed the spasmolytic effect of sevoflurane (S) as compared with enflurane (E) and halothane (H). DESIGN: Open-labeled, randomized study. SETTING: The study protocol was approved by the Animal Care Committee in a school of medicine. PARTICIPANTS: Twenty-two mongrel dogs. INTERVENTIONS: The dogs were anesthetized with pentobarbital, IV, paralyzed with pancuronium, and the lungs were mechanically ventilated. The endotracheal tube had an additional lumen to insert the SFB (outer diameter 2.2 mm). The tip of the SFB was located between a second and third bronchial bifurcation to continuously monitor the bronchial cross-sectional area (BCA) of third or fourth generation bronchi. The BCA was printed out by videoprinter at the end of expiration and was calculated on a Macintosh IIci computer using an NIH (National Institutes of Health) image program. Bronchoconstriction was produced with histamine (HA: 10 micrograms/kg + 500 micrograms/kg/hr). In the first protocol, 4 mongrel dogs were used to assess the accuracy of the new method. BCA was measured 30 and 60 minutes after the placement of the SFB and also 30 and 60 minutes after the start of the histamine (HA) infusion. Arterial blood (4 mL) was collected at 30 and 60 minutes after placement of the SFB to measure the plasma concentration of catecholamines with gas chromatographic mass spectrometry. In the second protocol, 18 dogs were randomly allocated to 3 groups of 6 each: group S, group E, and group H. Thirty minutes after the start of the HA infusion, the dogs subsequently inhaled the following concentrations of S, E, or H for 15 minutes: 0.2, 0.4, 0.8, and 1.6 minimum alveolar concentrations. The BCA was assessed before and 30 minutes after the start of HA and at the end of the inhalation period of each concentration. MEASUREMENTS AND MAIN RESULTS: In the first study, no significant differences in the BCA were found between 30 and 60 minutes after the placement of the SFB and between 30 and 60 minutes after the start of HA. The plasma concentrations of norepinephrine and epinephrine changed from 138 +/- 45 and 162 +/- 18 to 188 +/- 48 and 136 +/- 18 pg/mL, respectively. In the second study, all volatile anesthetics significantly increased the BCA in a dose-dependent manner. There was no significant difference among the three groups. CONCLUSION: As the BCA did not change in the first study, the tip of the SBF could be well fixed, and the 30-minute interval is long enough for the HA infusion to produce stable bronchoconstriction. The plasma catecholamine concentrations suggest that the pentobarbital anesthesia was deep enough to inhibit the direct irritant effect of the endotracheal tube and the SFB on the airway. This new direct method indicates that S dilates HA-constricted proximal airway with the same potency as E and H.

Anesthetics, Inhalation↗
PubMed1996

Relaxant effect of ketamine and its isomers on histamine-induced contraction of tracheal smooth muscle.

The mechanism by which racemic (R(+/-)) ketamine relaxes airway smooth muscle is unclear and there is no information on the differential effects of ketamine and its isomers. In this study, we have examined the spasmolytic effect of R(+/-) ketamine and its isomers S(+) and R(-) ketamine and the role of intracellular calcium and opioid receptors in R(+/-) ketamine-induced relaxation. The tension of isolated guinea pig tracheal strips was measured isometrically with a force displacement transducer and contraction elicited with histamine 10(-5) mol litre-1. In histamine-preconstricted strips, the two ketamine isomers (4.5-18.0 x 10(-4) mol litre-1) produced equipotent relaxation. A subthreshold dose of each isomer of ketamine (10(-4) mol litre-1) which alone did not relax histamine-induced contraction (S(+), P < 0.01; R(+/-), P < 0.01; R(-), P < 0.05) significantly potentiated adrenaline 1.25-5.0 x 10(-9) mol litre-1-induced relaxation (potency: S(+) > R(+/-) > R(-)). Increase in extracellular Ca2+ (1.8-14.4 x 10(-3) mol litre-1) significantly reduced R(+/-) ketamine-induced relaxation. S(-) Bay K 8644, at concentrations up to 2.0 x 10(-6) mol litre-1, partially antagonized R(+/-) ketamine-induced relaxation whereas at 10(-5) mol litre-1 or higher it potentiated the response. Naloxone 1.5-6.0 x 10(-6) mol litre-1 did not affect the relaxation caused by R(+/-) ketamine. We conclude that although both ketamine isomers produced equipotent spasmolytic effects on airway smooth muscle precontracted with histamine, they differed in their ability to potentiate the relaxing effect of adrenaline. S(+) ketamine produced the greatest potentiation. Changes in intracellular Ca2+ level secondary to a reduction in the L-type Ca2+ current may partially mediate the spasmolytic effect of R(+/-) ketamine.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1996

I.v. anaesthetic agents inhibit dihydropyridine binding to L-type voltage-sensitive Ca2+ channels in rat cerebrocortical membranes.

Previous studies have implicated the neuronal L-type voltage-sensitive Ca2+ channel (VSCC) as a target site for i.v. anaesthetic agents. It is unclear if these agents interact with the L-channel alpha-subunit 1,4-dihydropyridine (DHP) binding site. In this study, we have examined the interaction of thiopentone, pentobarbitone, ketamine, etomidate, propofol and alphaxalone, and the non-anaesthetic barbiturate, barbituric acid, with the DHP binding site on rat cerebrocortical membranes. Binding assays were performed in 1-ml volumes of Tris-HCl 50 mmol litre-1, pH 7.4, for 90 min at room temperature containing 200 micrograms of membrane protein with [3H]PN200-110 as a radiolabelled DHP. Non-specific binding was defined in the presence of nifedipine 10(-5) mol litre-1. The interaction of i.v. anaesthetics was determined by displacement of [3H]PN200-110 0.2 nmol litre-1. All i.v. anaesthetics showed some interaction with the DHP binding site. The concentrations of anaesthetic producing 25% inhibition of specific binding (corrected for the competing mass of [3H]PN200-110), K25 were (mumol litre-1): thiopentone 48 (SEM 2), pentobarbitone 95 (7), propofol 40 (2), etomidate 25 (2), alphaxalone 17 (3) and ketamine 198 (16). Barbituric acid was ineffective. With the exception of ketamine, there was a significant correlation between K25 and peak serum concentration during anaesthesia (P = 0.033) and serum concentrations on wakening (P = 0.018), suggesting that the L-channel DHP binding site may be a target for i.v. anaesthetic agents.

Anesthetics, Intravenous↗
PubMed1996

I.v. anaesthetic agents do not interact with the verapamil binding site on L-type voltage-sensitive Ca2+ channels.

In this study we have examined if the i.v. anaesthetic agents thiopentone, pentobarbitone, ketamine, etomidate, propofol and alphaxalone interact with the verapamil binding site on L-type voltage-sensitive Ca2+ channels in rat cerebrocortical membranes. Binding assays were performed in 1 ml volumes of Tris HCl 50 mmol litre-1, pH 7.4, for 90 min at 20 degrees C, with cerebrocortical membranes (200 micrograms of protein), the verapamil binding sites of which were radiolabelled with [3H]verapamil. Non-specific binding was defined in the presence of verapamil 10(-6) mol litre-1. The interaction of i.v. anaesthetics was determined by displacement of [3H]verapamil 0.2 nmol litre-1. The mean concentrations of anaesthetic producing 25% inhibition of specific binding (corrected for the competing mass of [3H]verapamil), K25, were (mmol litre-1): thiopentone 0.68 (SEM 0.14); pentobarbitone 1.22 (0.13); propofol 0.66 (0.10); etomidate 0.24 (0.03); alphaxalone 0.19 (0.02); and ketamine 0.75 (0.04). These concentrations exceeded those seen during anaesthesia and suggest that the neuronal verapamil binding site may not be an important target for i.v. anaesthetic agents.

Anesthetics, Intravenous↗