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Biomedical subjects

K Shibata

Publications and source records attributed to K Shibata.

At least 181 records · Page 10Linked to original sources
PubMed1998

GATA-1 inhibits the formation of notochord and neural tissue in Xenopus embryo.

The expression of GATA-1, which encodes for a hemopoietic transcription factor, initiates at gastrula stage in the Xenopus embryo (1). In order to examine a possible function of GATA-1 in dorso-ventral patterning of mesoderm and ectoderm derivatives, the synthesized RNA of GATA-1 was overexpressed in embryonic cells to assess its biological effects. In the embryos injected with GATA-1 RNA in the dorsal marginal zone at 4-cell stage, dorsal epidermis did not cover the vegetal cells so that the gastrulation was not completed. The same dose of GATA-1 RNA injected into ventral marginal zone did not influence the development, and GATA-2 RNA transcribed from the same vector had little effect, suggesting that this phenomenon is physiologically important. The morphological and immunohistochemical studies revealed that notochord and neural tissue were mostly eliminated in the embryos or the dorsal marginal zone explants after injection of GATA-1 RNA. GATA-1 also inhibited neurogenesis in animal cap explants, which was induced by the injection with noggin RNA. Northern blot analysis using dorsal marginal zone explants showed, however, that only a slight amount of alpha-globin message was induced, and cardiac alpha-actin message was retained. Therefore, GATA-1 did not convert completely the dorsal phenotype to the ventral one. Furthermore, the injection of GATA-1 RNA didnot alter the expression of early dorsal and ventral markers at the onset of gastrulation. These results suggest that GATA-1 is an potential inhibitor of the dorsalization and the neurogenesis, but it affects on the specification of dorsal tissues in relatively later steps.

Actins↗
PubMed1998

The increase in angiotensin type-2 receptor mRNA level by glutamate stimulation in cultured rat cortical cells.

The changes in the angiotensin type-2 (AT2) receptor mRNA level during glutamate neurotoxicity in cultured rat cortical cells are examined to assess the possible involvement of AT2 receptor in cell injury. The day 10-14 cortical neurons were exposed to glutamate at a toxic concentration of 100 microM for 15 min. The viability of the culture was reduced by 60% after 24 h. AT2 receptor mRNA was then increased 2-fold after exposure to glutamate, while the maximum increase was observed in a dose-dependent manner (50-1000 microM) 3 h after glutamate stimulation. AT2 receptor binding also increased 3-12 h after glutamate exposure. The results suggest that the increase in the AT2 receptor preceded to some extent the insult of the cell after exposure. The increase in the mRNA level was suppressed by MK-801, N-methyl-D-aspartate (NMDA) receptor antagonist, thus indicating the possible involvement of NMDA receptor. The increase in the mRNA level was also antagonized by N-nitro L-arginine methyl-ester, a nitric oxide synthase inhibitor. The hemoglobin, a nitric oxide trap, inhibited the increase in the mRNA level. These results suggest that the increase in the mRNA level is associated with the nitric oxide synthesis by glutamate exposure. The viability of cortical cells after glutamate stimulation was partially restored by the AT2 receptor antagonist and by the antisense oligonucleotide for the AT2 receptor. The present results thus suggest that the AT2 receptor may in some way be related to one of the processes in cell injury caused by glutamate.

Angiotensin II↗
PubMed1998

[Expertise in occupational health nursing (II)--Report on the 17th UOEH International Symposium].

Three hundred and fifty-two OHNs and collaborative persons from 17 countries and the ILO assembled at UOEH to participate in the 17th UOEH International Symposium to commemorate the inauguration of the UOEH School of Health Sciences from October 20 to 22, 1997. The main theme of this Symposium was "Occupational Health Nurse (OHM) Expertise." The Symposium was opened by greetings from Dr. Akira Koizumi, followed by a message from ILO presented by Dr. G.H. Cappee, head of the Medical Section Occupational Safety & Health Branch, ILO. Dr. Bonnie Rogers of North Carolina University, Chapel Hill, and President of AAOHN, lectured on the main theme of the Symposium, and Dr. M.A. Fingerhut gave a lecture on "Partnership in Occupational Safety and Health." At the general sessions, 5 themes were discussed.--1) Partnership in Occupational Health, 2) Education and Training of OHNs, 3) Intervention/Behavior to Disease Prevention and Health Promotion, 4) How to Integrate Computing into Occupation Health Nursing and 5) Cost/Benefit Effectiveness of Occupational Health Nursing. For all 5 sessions, there were keynote lectures, oral and poster presentations. This report reviews the lectures by Dr. B. Rogers on "Expertise in Occupational Health Nursing," by Dr. Kuchinski on "Education and Training of Occupational Health Nurses in The US," the presentation by Mrs. J. Fanchette, DIUST/GIT Service de Pathologie Professionnelle Hospital Civil, France, on "Evaluation of An Interuniversity Diploma Course Occupational Health Nurse Qualification," by Dr. K. June, Soonchunhyang Univ., Korea, on "Transition of Occupational Health Nursing Education in Korea" and finally, the lecture by Dr. F. Mitsuhashi on "Revisions in the Industrial Safety and Health Law and Expected Models for Occupational Health Nursing Staff."

International Cooperation↗
PubMed1998

Selective separation of trypsin from pancreatin using bioaffinity in reverse micellar system composed of a nonionic surfactant.

Selective separation of trypsin from a mixture involving many kinds of contaminating proteins, i.e., pancreatin, was achieved using trypsin inhibitor immobilized in the reverse micelles, which were composed of a nonionic surfactant, tetra-oxyethylene monodecylether. To determine the efficient operations throughout the whole separation process we examined the operating conditions, which affect the immobilization efficiency of trypsin inhibitor and also the forward and backward extractions of trypsin. Fifty percent of the recovery of trypsin from pancreatin was realized with no loss of activity of the recovered trypsin.

Animals↗
PubMed1998

An improved method for the detection of changes in brain extracellular glutamate levels.

We developed a method for in vivo real-time monitoring of the concentration of extracellular glutamate ([Glu]e) in the brain under anoxic conditions. A dialysis electrode (Sycopel Int., UK) was employed as a sensing device to measure the concentration of glutamate by enzyme amperometry, and an electron mediator, ferrocene, was introduced into the electrode together with glutamate oxidase. The ferrocene was covalently conjugated with a high molecular weight molecule, bovine serum albumin, to avoid outward diffusion through the dialysis membrane. With this set-up, the amperometric response was independent of the pO2 around the electrode in vitro up to 400 microM glutamate. Using this method, we investigated the dynamics of [Glu]e in the rat striatum during anoxia. [Glu]e increased rapidly at 102+/-5.4s (n = 6) after the start of nitrogen inhalation. The increase continued for about 30 s, and then [Glu]e decreased. The peak value of delta[Glu]e was 141+/-37 micro M. [Glu]e subsequently underwent another gradual increase, reaching 213+/-69 microM at 15 min after the start of nitrogen inhalation. This distinct biphasic profile was reproducible. We conclude that this method is very useful for monitoring [Glu]e in the brain under low pO2 conditions.

Animals↗
PubMed1998

Identification and cloning of human G-protein gamma 7, down-regulated in pancreatic cancer.

Differentially expressed genes between normal and cancer tissues of the pancreas were investigated using differential display. Consequently, we identified a fragment cDNA that was expressed in the normal tissue but was rarely expressed in the cancer tissue. This cDNA was screened in cDNA library prepared from the normal pancreatic tissue by rapid amplification of cDNA ends (5'RACE). 859 bp of cDNA was cloned and sequenced, and the inferred amino acid sequence was found to encode a G protein gamma subunit with 98% homology to cow G protein gamma 7 and complete homology to human G protein gamma 7. The decreased expression of the G protein gamma 7 was confirmed by Northern blot assay in twelve pancreatic malignancies which included nine duct cell carcinomas, two cystoadenocarcinomas and one blastoma. Reverse transcriptase (RT)-polymerase chain reaction (PCR) assay showed no expression of G protein gamma 7 in five of six pancreatic carcinoma cell lines and two pancreatic cancer tissues. Immunohistochemical analysis also displayed positive staining in the normal tissue but no staining in the cancer tissue. The findings demonstrated that the reduced or suppressed expression of human G-protein gamma 7 may play an important role in pancreatic carcinogenesis.

Amino Acid Sequence↗
PubMed1998

A bifunctional hybrid molecule of the amino-terminal fragment of urokinase and domain II of bikunin efficiently inhibits tumor cell invasion and metastasis.

Urinary trypsin inhibitor (UTI) inhibits efficiently tumor cell invasion and the formation of metastasis. The anti-metastatic effect is dependent on the COOH-terminal domain II of UTI [UTI-(78-136)-peptide]. To develop a molecule that binds with high affinity to the urokinase (uPA) receptor (uPAR) on tumor cell surfaces, a bifunctional hybrid molecule [uPA-(1-134)-UTI-(78-136)] consisting of the uPAR-binding NH2-terminal fragment [UTI-(78-136)-peptide] of uPA at the NH2-terminus of UTI-(78-136)-peptide was produced in Escherichia coli by genetic engineering. The purified hybrid protein inhibited trypsin and plasmin 2-3-fold less effectively than UTI-(78-136)-peptide and was found to bind to human tumor cells via uPAR, which was confirmed by cell binding and competition experiments. Using a modified Boyden chamber and an artificial basement membrane, Matrigel, it was found that the hybrid protein is very effective at inhibiting invasion by uPAR-expressing human tumor cells. Sensitivities of tumor cells towards the anti-invasive effect of uPA-(1-134)-UTI-(78-136) correlated with the density of uPAR on human tumor cells. Furthermore, in the spontaneous metastasis model, the hybrid protein inhibited the formation of lung and/or lymphatic metastasis by human ovarian carcinoma and choriocarcinoma cells. The hybrid protein was much more effective than uPA-(1-134)-peptide, UTI-(78-136)-peptide, or UTI. We conclude that this approach extends the possibility of applying recombinant protein for therapeutic use in inhibition of human tumor cell metastasis.

Amino Acid Sequence↗
PubMed1998

Effects of quinidine and verapamil on human cardiovascular alpha1-adrenoceptors.

BACKGROUND: The antiarrhythmic drugs quinidine and verapamil are known to block alpha1-adrenoceptors (alpha1ARs). Alpha1ARs are a heterogeneous family of three subtypes (alpha1A, alpha1B, and alpha1D), and little is known about the effects of quinidine and verapamil on the different human alpha1AR subtypes. METHODS AND RESULTS: Reverse transcriptase-polymerase chain reaction showed that all alpha1AR subtypes are expressed in both human heart (atrium and ventricle) and the mesenteric artery. Pharmacological profiles of quinidine and verapamil actions on the alpha1AR subtypes were characterized with Chinese hamster ovary cells stably expressing cloned human alpha1AR subtypes. Radioligand binding studies showed that quinidine and verapamil had high affinities for all alpha1AR subtypes. Also, both drugs synergistically inhibited alpha1AR-mediated inositol 1,4,5-triphosphate production at the clinical effective concentration range (1 micromol/L quinidine and 0.1 micromol/L verapamil). CONCLUSIONS: The results show that all alpha1AR subtypes are expressed in the human cardiovascular system and that quinidine and verapamil may have a potent, synergistic inhibitory effect on the alpha1ARs. Clinically observed hypotension after quinidine plus verapamil can be explained by their synergistic inhibitory effects on human alpha1ARs.

Adrenergic alpha-1 Receptor Antagonists↗
PubMed1998

Identification and characterization of the cell-associated binding protein for urinary trypsin inhibitor.

Urinary trypsin inhibitor (UTI) inhibits not only tumor cell invasion but also production of experimental and spontaneous metastasis. Cell-binding experiments indicated that human choriocarcinoma SMT-cc1 cells have specific binding sites for UTI on their cell surface. [Kobayashi et al., J. Biol. Chem. 269, 1994, 20,642-20,647]. UTI binding protein (UTIBP) was purified to homogeneity by a combination of UTI-coupled affinity beads, preparative polyacrylamide gel electrophoresis and reverse phase HPLC. This protein is very similar to a truncated form of human cartilage link protein (LP). LP was identified structurally by its apparent molecular mass with and without deglycosylation treatment: Immunologically by the reactivity with anti-UTIBP antibody, and functionally by its ability to bind the NH2-terminal domain of UTI. UTI and UTIBP are distributed uniformly in the cytoplasm and/or over the cell surface of tumor cells and fibroblasts. The level of staining for hyaluronic acid, UTIBP and UTI is much lower in sections digested with hyaluronidase. These results suggest that the cell membrane-derived UTI-associated binding protein is the LP of proteoglycan-hyaluronic acid aggregates, which interacts with hyaluronic acid. Cell-associated LP may play a role in modulating protease activity to the environment close to tumor and fibroblast cell surface.

Chromatography, High Pressure Liquid↗
PubMed1998

Ecabet sodium, a locally acting antiulcer drug, inhibits urease activity of Helicobacter pylori.

In order to clarify the mechanism of the anti-Helicobacter pylori action of ecabet sodium (ecabet), a locally acting antiulcer drug, we evaluated the effects of ecabet on H. pylori urease activity in vitro. H. pylori was cultured and a crude preparation of urease was made. Urea-dependent survival of H. pylori at acid pH was significantly inhibited by ecabet. The urease activity of intact cells and a crude enzyme preparation from H. pylori had two pH optima: pH 4.5-5.0 and 8.0. Ecabet (1-4 mg/ml) concentration dependently inhibited the urease activity of both preparations at pH 5.0, but there was no inhibition at pH 8.0. The enzyme activity was inhibited by ecabet gradually and was not restored by dilution, in contrast to the inhibition elicited by benzohydroxamic acid, a specific and reversible urease inhibitor. These results suggest that irreversible inhibition of H. pylori urease activity contributes to the anti-H. pylori action of ecabet.

Abietanes↗
PubMed1998

T-loop deletion of CDC2 from breast cancer tissues eliminates binding to cyclin B1 and cyclin-dependent kinase inhibitor p21.

The eukaryotic cell cycle is regulated by a highly conserved family of protein kinases, the cyclin-dependent kinases (CDKs). Monomeric free CDKs do not possess enzymatic activity, largely due to the steric hindrance caused by the T-loop at the entrance of the catalytic cleft, making ATP inaccessible to the substrate. Binding of a cyclin, primarily to the NH2-terminal lobe of the CDK that surrounds the PSTAIRE helix, induces a large conformational change in the PSTAIRE helix of the CDK and also causes the T-loop to move out of the way of the catalytic cleft. We identified from breast cancer tissues a novel variant of human CDC2, termed CDC2deltaT, that lacks 171 nucleotides corresponding to 57 amino acids, which compose most of the T-loop. CDC2deltaT was detected in 10 of 14 breast cancer tissues analyzed, whereas it was not detectable in diploid human fibroblast cell lines or in interleukin 2-stimulated normal human lymphocytes. CDC2deltaT protein is unable to complex with cyclin B1 and lacks histone H1 kinase activity. CDC2deltaT also fails to bind to the CDK inhibitor p21. These results indicate that the T-loop not only plays a key role in keeping a free CDK in its inactive state but also in facilitating CDK activation by promoting cyclin binding.

Alternative Splicing↗
PubMed1998

[Expertise in occupational health nursing (I). Report on a visit to US occupational health institutions].

As industries' structures have evolved and diversified recently, the framework of occupational health nursing has been expanded on multidisciplinary bases. On the occasion of the opening of the School of Health Sciences, University of Occupational and Environmental Health, Japan (UOEH), the author was given an opportunity to visit US administrations, academic institutions and some industries to investigate occupational medicine and health nursing in that country. And as the author took part in programming the 17th UOEH International Symposium on Occupational Health Nursing Expertise that took place at the campus of the University in October, 1997, these two events are reported in this paper. The first part of the report is on the visit to the US occupational health professionals including occupational health doctors, nurses, industrial hygienists and other staffs. They have perceived that there has never been more emphasis placed on the significance of occupational health services than in recent years, and that young students have become more interested in occupational health. The second report will be on the 17th UOEH International Symposium--Occupational Health Nursing Expertise--the next time.

Education, Nursing↗
PubMed1998

Both focal adhesion kinase and c-Ras are required for the enhanced matrix metalloproteinase 9 secretion by fibronectin in ovarian cancer cells.

Cell adhesion to the extracellular matrix appears to trigger a cascade of intracellular signalings. We have shown previously that treatment of ovarian cancer cells with peritoneal conditioned medium or purified fibronectin (FN) activated matrix metalloproteinase 9 secretion and, thereby, cancer cell invasion. By use of antisense oligonucleotides to focal adhesion kinase (FAK) and a dominant-negative mutant of ras (S17Nras), we found that both FAK and c-Ras were required for the activation of matrix metalloproteinase 9 secretion by FN. In addition, both antisense oligonucleotides to FAK and S17Nras inhibited mitogen-activated protein kinase activation by FN treatment, suggesting the involvement of mitogen-activated protein kinase in the FN-dependent signaling.

Cell Adhesion Molecules↗
PubMed1998

Characterization and functional expression of the cDNA encoding human brain quinolinate phosphoribosyltransferase.

Mammalian quinolinate phosphoribosyltransferase (QPRTase) (EC 2.4.2.19) is a key enzyme in catabolism of quinolinate, an intermediate in the tryptophan-nicotinamide adenine dinucleotide (NAD) pathway. Quinolinate acts as a most potent endogenous exitotoxin to neurons. Elevation of quinolinate levels in the brain has been linked to the pathogenesis of neurodegenerative disorders. As the first step to elucidate molecular basis underlying the quinolinate metabolism, the cDNA encoding human brain QPRTase was cloned and characterized. Utilizing partial amino acid sequences obtained from highly purified porcine kidney QPRTase, a human isolog was obtained from a human brain cDNA library. The cDNA encodes a open reading frame of 297 amino acids, and shares 30 to 40% identity with those of bacterial QPRTases. To confirm that the cDNA clone encodes human QPRTase, its functional expression was studied in a bacterial host. Introduction of the human cDNA into a QPRTase defective (nadC) E. coli strain brought about an abrupt increase in QPRTase activity and allowed the cells to grow in the absence of nicotinic acid. It is concluded that the cloned cDNA encodes human QPRTase which is functional beyond the phylogenic boundary.

Amino Acid Sequence↗
PubMed1998

Occurrence of free D-aspartate and aspartate racemase in the blood shell Scapharca broughtonii.

Substantial concentrations of D-aspartate were found in several tissues of Scapharca broughtonii together with approximately equal concentrations of L-aspartate. The foot and mantle extracts also contained an aspartate racemase activity. The formation of L-aspartate from the D-enantiomer by the foot extract was apparently slower than the reverse reaction, and this unbalance seemed to be due to the presence of an enzyme activity which rapidly converted L-aspartate to L-alanine. The possible role of D-aspartate in the anaerobiosis was discussed.

Amino Acid Isomerases↗
PubMed1998

Mechanism of acute lung injury caused by inhalation of fabric protector and the effect of surfactant replacement.

OBJECTIVE: To evaluate the role of surfactant in the mechanism and treatment of acute lung injury caused by inhalation of fabric protector. DESIGN: Prospective, randomized study. SETTING: University laboratory. INTERVENTIONS: In vitro experiment: a porcine surfactant suspension (10 mg.ml-1) was exposed to a fabric protector aerosolized with an ultrasonic nebulizer for 1 min. Minimum surface tension (gamma min) was sequentially measured using pulsating bubble equipment. Animal experiment: 14 adult rats were anesthetized with pentobarbital and mechanically ventilated with pure oxygen. Then, all rats inhaled fabric protector aerosolized with the nebulizer for five breaths. Three hours after inhalation, the rats were randomly assigned to two groups: a surfactant group (n = 7), in which surfactant (100 mg.kg-1) was replaced, and a control group (n = 7), in which no substance was given. MEASUREMENTS AND RESULTS: In vitro experiment: exposure to fabric protector aerosol increased the mean gamma min of the surfactant from 1.7 to 19.2 mN.m-1 (n = 5, p < 0.05). Animal experiment; the mean partial pressure of oxygen in arterial blood (PaO2) in all rats decreased from 62.8 to 17.1 kPa at 3 h after inhalation. The PaO2 in the surfactant group increased to 49.8 +/- 11.1 (SD) kPa at 30 min after surfactant replacement (p < 0.05), while the PaO2 in the control group remained below 20 kPa. CONCLUSIONS: Impairment of surfactant is a factor involved in the development of acute lung injury caused by inhalation of fabric protector. Surfactant replacement may be therapeutic for such injuries.

Acetates↗
PubMed1998

Urinary trypsin inhibitor reduces the release of histamine from rat peritoneal mast cells.

We determined the ability of urinary trypsin inhibitor (UTI), which is a Kunitz-type protease inhibitor present in serum and in urine, to inhibit rat peritoneal mast cell (RPMC) mediator release induced by several stimuli. UTI attenuated the immunoglobulin E-mediated release of both preformed (histamine) and newly formed (leukotriene C4) mediators from RPMCs. Inhibition (21%+/-5%) of the anti-IgE-triggered release of histamine was observed after a 30-minute incubation of RPMCs with UTI (5 micromol/L). To investigate the specificity of the UTI effect, we studied the stimulatory activity of phorbol ester (phorbol 12-myristate 13-acetate (PMA)) or calcium ionophore A23187 in control and UTI-treated mast cells. The efficacy of UTI as an inhibitor was dependent on the nature of the stimulus, because histamine release induced by PMA-mediated or calcium ionophore A23187-mediated processes was not inhibited by UTI. A series of structurally distinct protease inhibitors did not inhibit IgE-induced release of mediators from RPMCs. The Kunitz-type protease inhibitors are important in the regulation of RPMC function. In parallel with the UTI-related decrease in anti-IgE stimulatory activity on mediator release, increased microviscosity of membrane lipids could be observed by two independent experiments on fluorescence polarization with diphenylhexatriene (DPH) and on the fluorescence probe fluorescein isothiocyanate-concanavalin A. UTI reduces mediator release by a mechanism-possibly an interruption of the coupling of receptor and effector systems-because UTI acts as an agent to decrease biologic lipid membrane fluidity.

Animals↗