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Biomedical subjects

M Shibuya

Publications and source records attributed to M Shibuya.

At least 181 records · Page 10Linked to original sources
PubMed1997

Effective cross-circulation technique of venoarterial bypass for differential hypoxia condition.

We examined a new technique of cross-circulation (CC) venoarterial bypass (VAB) with femoral arterial perfusion and superior vena cava drainage through a long femoral venous cannula. Six adult mongrel dogs weighing 15 to 20 kg underwent the CC-VAB with oxygenation after introduction of respiratory failure (RF). The flow of the CC-VAB was maintained at half the level of the control cardiac output, and the hemodynamic parameters were monitored. To evaluate hypoxia in the upper body, the arterial partial pressure of oxygen (PaO2 [mm Hg]) in the carotid artery and the venous saturation of oxygen (SvO2 [%]) in the pulmonary artery were measured during control, RF, standard VAB, and CC-VAB conditions. The PaO2 decreased significantly after the introduction of RF (41.7 +/- 12.4), and it returned to normal levels only after CC-VAB (151.2 +/- 24.5, p < 0.05). The SvO2 during CC-VAB (98.6 +/- 2.1) was significantly higher than that during VAB without CC (53.5 +/- 3.4, p < 0.05). These results suggest that this cross-circulation technique could be applied to patients with differential hypoxia during femoral VAB with oxygenation or percutaneous cardiopulmonary support (PCPS).

Analysis of Variance↗
PubMed1997

Significant left ventricular unloading with transaortic catheter venting during venoarterial bypass.

Insufficient unloading of the left ventricle with blood stagnation is a main cause of unsuccessful left ventricular (LV) recovery during percutaneous cardiopulmonary support (PCPS). The purpose of this investigation was to evaluate the effectiveness of transaortic catheter venting (TACV) for LV unloading. Six adult mongrel dogs (mean weight 16.3 kg, range 14-20 kg) underwent venoarterial bypass (VAB) with TACV. Bypass flow ranged from 0.8-1.2 L/min, and TACV flow ranged from 160-240 ml/min. In addition to monitoring the standard hemodynamic parameters, the slope of the LV end-systolic pressure-volume relation (Emax) during transient occlusion of the inferior vena cava, the slope of the LV end-systolic pressure-stroke-volume relation (Ea), the stroke work (SW), the LV pressure-volume area (PVA), and the slope of the SW end-diastolic volume relation, the preload recruitable stroke work (PRSW) were assessed by means of a microtip manometer and a conductance catheter. The LV contractility (Emax) and aortic elastance (Ea) were equivalent in the 2 groups with or without TACV (7.7 +/- 1.1 versus 8.4 +/- 1.5 mm Hg/ml and 8.2 +/- 1.4 versus 7.6 +/- 1.3 mm Hg/ml). Comparing the measurements for the baseline to those for VAB with TACV, the SW was significantly reduced, and the PVA/SW was increased by TACV (1,685 +/- 309 versus 867 +/- 188 x 10(-4) J, p < 0.05 and 1.32 +/- 0.03 versus 1.58 +/- 0.11, p < 0.05, respectively). Furthermore, the PRSW was gradually decreased from the baseline value to the value resulting from VAB with TACV (75 +/- 8 versus 44 +/- 3 x 10(-4) J/ml, p < 0.01). In comparison, the percent reduction of SW between VAB and VAB with TACV tended to be increased by TACV (23.2 +/- 7.2% versus 46.9 +/- 7.7%, p = 0.05). These results suggest that TACV might reduce LV work (SW and PRSW) and might increase the LV energetic charge. In conclusion, TACV would be an adjunctive technique to VAB or PCPS for patients with LV failure.

Angioplasty, Balloon, Coronary↗
PubMed1997

Tyrosine phosphorylation sites at amino acids 239 and 240 of Shc are involved in epidermal growth factor-induced mitogenic signaling that is distinct from Ras/mitogen-activated protein kinase activation.

Epidermal growth factor (EGF) induces tyrosine phosphorylation of the Shc adapter protein, which plays an important role in EGF-stimulated mitogenesis. Shc stimulates Ras/mitogen-activated protein kinase (MAPK) through forming a complex with Grb2 at the phosphorylated tyrosine (Y) residue 317. In this study, we identified novel phosphorylation sites of Shc, at Y239 and Y240. To define the Shc pathway further, we used NIH 3T3 cells expressing the previously characterized mutant EGF receptor (EGF-R) which lacks all known autophosphorylation sites but retains EGF-stimulated mitogenesis with selective phosphorylation of Shc. We constructed wild-type (WT) or mutant Shc cDNAs in which Y317 or/and Y239 and Y240 are replaced with phenylalanine (F) and introduced them into NIH 3T3 cells expressing WT or mutant EGF-R. In the WT EGF-R-expressing cells, the Y239/240/317F Shc, but not Y317F or Y239/240F Shc, decreased EGF-stimulated cell growth. In the mutant EGF-R-expressing cells, Y317F Shc or Y239/240F Shc decreased EGF-stimulated cell growth significantly, though Y317F was a little more potent than Y239/240F. Although cells expressing the Y317F Shc hardly activated MAPK in response to EGF, cells expressing the Y239/240F Shc fully activated MAPK. In contrast, Y239/240F Shc, but not Y317F Shc, reduced the EGF-induced c-myc message. These results suggest that Shc activates two distinct signaling pathways, Y317 to Ras/MAPK and Y239 and Y240 to another pathway including Myc, and that both are involved in EGF-induced mitogenic signaling.

3T3 Cells↗
PubMed1997

In vitro conversion of 2,3-oxidosqualene into dammarenediol by Panax ginseng microsomes.

(RS)-[3-3H]-2,3-Oxidosqualene (1) was converted into (20S)-dammarenediol (2) and not to (20R)-dammarenediol by a microsomal fraction prepared from the hairy root of Panax ginseng. The enzyme activity was highest at pH 6.0 and was not increased by the addition of any detergents. These properties differed significantly from those of other 2,3-oxidosqualene cyclases reported from higher plants and animals.

Cyclization↗
PubMed1997

Molecular cloning of pea cDNA encoding cycloartenol synthase and its functional expression in yeast.

The cDNA encoding cycloartenol synthase [EC 5.4.99.8] has been isolated from pea seedling by an efficient PCR using sets of degenerate primers based on the highly conserved sequences of the known 2,3-oxidosqualene cyclase cDNAs. The obtained cDNA contains a 2271-bp open reading frame and is encoding a predicted protein of 757 amino acids with high homology (81%) to Arabidopsis thaliana cycloartenol synthase. The PCR-amplified open reading frame (ORF) has been inserted into pYES2, an expression vector in yeast, under the control of galactose-inducible promoter. Significant cycloartenol synthase activity has been found in the homogenate of the yeast transformed with the plasmid containing PCR-amplified ORF.

Amino Acid Sequence↗
PubMed1997

[Combined effect of adoptive immunotherapy (AIT) with lymphokine-activated killer (LAK) cells and interleukin-2 (IL-2) and chemotherapy in tumor-bearing mice].

We investigated beneficial effects of AIT with anticancer agents on survival of subcutaneous tumor-bearing mice and suppression of artificial lung metastasis, and optimal schedule of administration of each treatment in vivo. 7-8 weeks old C 57 BL/6 mice were inoculated s.c. with 5 x 10(6) B 16 melanoma cells, or i.v. with 2 x 10(5) B 16-F 10 melanoma cells. Mouse splenocytes were cultured with 3.5 x 10(3) JRU/ml interleukin 2 for 14 days, and induced LAK cells were harvested. Anticancer agents (Cx), CDDP or MMC were given i.p. in mice. 1 x 10(7) or 5 x 10(7) LAK cells were given either s.c. around the tumor or i.v. respectively, and 1.4 x 10(5) JRU/mouse IL-2 was administered s.c. for 6 days after LAK cell injection. Therapy groups were as follows #1: Cx day 3, AIT day 3-8. #2: Cx day 3, AIT day 6-11, #3: Cx day 8, AIT day 3-8. In therapy groups #1 and #2, we observed additive effects of AIT and anti-cancer agents in life-prolongation and suppression of lung metastasis. It was also shown that LAK induction in vivo was augmented by anticancer agents in groups #1 and #2, which might represent one of the mechanisms behind observed additive effects. Furthermore, our results suggest that therapeutic effects of the combination of AIT and anticancer agents depend on the schedule of administration.

Animals↗
PubMed1997

Anisotropic expansion in gypsum-bonded cristobalite investment mold.

Expansion of a gypsum-bonded investment mold was investigated using a fused quartz tube pattern. Two hours after mixing, a fusible alloy was cast into the space around the pattern induced by setting expansion. The other molds were heated up to 700 degrees C, then a dental silver alloy was cast into the space around the pattern induced by total expansion of the mold. Thickness of the castings was measured at every thirty degrees on seven cross-sections. The setting expansion was almost uniform, but the total expansion was not. Especially, a large amount of total expansion was produced toward the open ends of the casting ring and toward a seam of the asbestos liner. The thermal expansion of the mold was anisotropic because the investment was a mixture of calcium sulfate and silica, and was affected considerably by its environment.

Anisotropy↗
PubMed1997

[Adverse drug reaction criteria of the Japan Society for Cancer Therapy].

The Japan Society for Cancer Therapy Committee of Criteria for Efficacy of Cancer Therapy proposed new toxicity grading criteria called "Adverse Drug Reaction Criteria". The basic principles of the JCOG Toxicity Criteria were applied to the Adverse Drug Reaction Criteria of the Japan Society for Cancer Therapy with the following modifications: 1. In Grade 4 toxicity for stomatitis, "requires preventative intubation" was changed to "requires enteral support". 2. In Grade 0 for proteinuria, "none" was changed to "no change". (in addition, the definitions of Grade 1 to 3 toxicity for proteinuria were changed to concentration of proteinuria adapted to Japanese.) 3. The toxicity grading criteria was named Adverse Drug Reaction Criteria of the Japan Society for Cancer Therapy.

Antineoplastic Agents↗
PubMed1996

BCR-ABL induces neurite-like structures and BCR lacking the SH2-binding domain induces cell rounding in PC12 cells.

The activated tyrosine kinase oncoprotein BCR-ABL is responsible for pathogenesis of Philadelphia chromosome-positive human leukemias. Because BCR carries a GAP (GTPase-activating protein) activity toward cytoskeleton-related small GTP-binding proteins, we utilized a neuronal PC12 cell system to test morphogenic potentials of BCR-ABL or BCR. We report here unique morphological phenotypes of PC12 cells expressing either BCR-ABL or a BCR mutant which lacks the SH2-binding domain (BCR Delta162-413). Although MAP kinase was not activated in PC12 cells expressing BCR-ABL, they showed incomplete neurite extensions even in the absence of the nerve growth factor (NGF). Overproduction of BCR Delta162-413 in PC12 cells, on the other hand, induced cell rounding in the absence of NGF. Interestingly, those cells could hardly make terminal differentiation in the presence of NGF and continued to grow without changing their round shape, although NGF receptor as well as MAP kinase appeared to be activated. Interestingly, the botulinum C3 toxin induced neurite-like structures in PC12 cells overexpressing BCR Delta162-413 without NGF.

Animals↗
PubMed1996

A cAMP response element and an Ets motif are involved in the transcriptional regulation of flt-1 tyrosine kinase (vascular endothelial growth factor receptor 1) gene.

The flt-1 gene encodes a transmembrane tyrosine kinase, Flt-1, a receptor for vascular endothelial growth factor. The expression of flt-1 gene is restricted to endothelial cells in vivo. To understand the molecular mechanism underlying endothelial-specific expression of this gene, we studied the functional significance of transcriptional motifs in the 200-base pair region of the human flt-1 gene promoter, which has been identified to confer cell type specificity. By point mutation analysis using chloramphenicol acetyltransferase plasmids in 293E1 cells, which express significant levels of flt-1 mRNA, we found that an Ets motif, E4, at -54 to -51 and a cAMP response element (CRE) at -83 to -76 are involved in the transcriptional regulation of this gene. Disruption of either this CRE or E4 within the promoter sequence of 90 base pairs resulted in a decrease in chloramphenicol acetyltransferase activity of 90%, indicating that co-existence of both of CRE and Ets motif E4 is necessary for transcription of the flt-1 gene. Co-transfection of an expression vector containing c-ets-1, c-ets-2, or c-erg cDNA with this 90-base pair sequence yielded a 5-8-fold elevation of chloramphenicol acetyltransferase activity, further supporting the idea that Ets family protein(s) participates in the regulation of the flt-1 gene. Gel shift assays using nuclear extracts of 293E1 and endothelial cells demonstrated the existence of protein factor(s) that specifically binds to CRE and Ets motif E4, respectively. Taken together, our results strongly suggest cooperation of a CRE and an Ets motif for the function of the flt-1 gene promoter.

Binding Sites↗
PubMed1996

A novel pathway from phosphorylation of tyrosine residues 239/240 of Shc, contributing to suppress apoptosis by IL-3.

Interleukin 3 (IL-3) not only induces DNA synthesis of haematopoietic cells but also maintains their viability by suppressing apoptosis. IL-3 stimulates tyrosine phosphorylation of the Shc adaptor protein and thereby formation of a complex of Shc with Grb2 at phosphorylated tyrosine (Y) residue 317-Shc. This pathway is implicated in Ras/mitogen-activated protein kinase (MAPK) activation towards c-fos gene expression. We examined the possible involvement of Shc in the antiapoptotic activity of IL-3. Conditional overexpression of the Shc SH2 domain, a dominant-negative mutant of Shc, was found to induce apoptosis of IL-3-dependent Ba/F3 cells along with a reduction of c-myc gene expression. Apoptosis was rescued by the exogenously introduced c-myc gene. Since we identify novel tyrosine phosphorylation sites of Shc: Y239 and Y240, their role on cell survival was tested by mutational analysis. Ba/F3 cells expressing mutant Shc Y317F, which is unable to stimulate efficiently the Ras pathway, still showed resistance to apoptosis. However, cells expressing Shc Y239/240F, which is able to stimulate the Ras pathway, were sensitive to apoptosis. In these cells, induction of the c-myc gene was reduced. These findings suggest that a new signalling pathway for cell survival is generated from Y239/240 of Shc to the nuclei involving c-myc gene expression.

Apoptosis↗
PubMed1996

Possible mechanism of the potent vasoconstrictor responses to ryanodine in dog cerebral arteries.

Isolated cerebral (basilar, posterior communicating and middle cerebral) arteries exist in a partially contracted state. To determine the Ca(2+)-buffering function of sarcoplasmic reticulum in the resting state of cerebral arteries, the effects of ryanodine that eliminates the function of sarcoplasmic reticulum, on tension and cellular Ca2+ level were compared in endothelium-denuded strips of the cerebral, coronary and mesenteric arteries of the dog. The addition of ryanodine to strips with basal tone caused a concentration-dependent contraction, which was significantly greater in the cerebral arteries than in the mesenteric or coronary artery. In the presence of 10(-5) M ryanodine, the caffeine (20 mM)-induced contraction was greatly attenuated in these arteries. After washout, the basal tone was greatly elevated in the cerebral arteries. The elevated tone was abolished by 10(-7) M nifedipine. The ryanodine-induced contractions were also abolished by 10(-7) M nifedipine. Nifedipine itself caused a relaxation from the basal tone in the cerebral arteries, suggesting the maintenance of myogenic tone. The basal Ca2+ influx in arteries measured after a 5-min incubation with 45Ca was significantly higher in the basilar artery than in the mesenteric artery. The basal Ca2+ influx was not increased by 10(-5) M ryanodine in either artery. The basal Ca2+ influx was decreased by 10(-7) M nifedipine in the basilar artery, but was unchanged in the mesenteric artery. These results suggest that: (1) the basal Ca2+ influx via L-type voltage-dependent Ca2+ channels was higher in the resting state of the cerebral arteries; (2) the greater part of the higher Ca2+ influx was buffered by Ca2+ uptake into the sarcoplasmic reticulum; and (3) therefore the functional elimination of sarcoplasmic reticulum by ryanodine caused a potent contraction in these arteries. Furthermore, the maintenance of myogenic tone in the cerebral arteries suggests that more Ca2+ enters the smooth muscle cell than the buffering ability of sarcoplasmic reticulum can handle.

Animals↗
PubMed1996

Increased expressions of vascular endothelial growth factor and its receptors, flt-1 and KDR/flk-1, in regenerating rat liver.

Sinusoidal endothelial cells proliferate following hepatocyte regeneration in the liver after partial resection. The expressions of vascular endothelial growth factor (VEGF) and its receptors, flt-1 and KDR/flk-1, were studied by Northern blotting in isolated rat liver cells and 70% resected rat liver. VEGF was expressed in hepatocytes immediately after isolation, and both flt-1 and KDR/flk-1 were expressed in non-parenchymal cells including sinusoidal endothelial cells. The VEGF expression in hepatocytes decreased during primary culture for 48 hr. This expression was maintained at 48 hr of culture by addition of EGF to the medium at 24 hr, increasing thereafter. VEGF, flt-1, and KDR/flk-1 were also expressed in normal rat liver. In 70% resected rat liver, VEGF expression increased with a peak at 72 hr after the operation, followed by expressions of flt-1 and KDR/flk-1 increasing between 72 and 168 hr. These results suggest that VEGF expression increases in regenerating hepatocytes, which may contribute to proliferation of sinusoidal endothelial cells of rat liver following partial resection, probably through flt-1 and KDR/flk-1 receptors upregulated on sinusoidal endothelial cells.

Animals↗
PubMed1996

Molecular cloning and bacterial expression of a cDNA encoding furostanol glycoside 26-O-beta-glucosidase of Costus speciosus.

Furostanol glycoside 26-O-beta-glucosidase (F26G) purified from Costus speciosus rhizomes was digested with endoproteinase, and several internal peptide fragments were obtained. Degenerate oligonucleotide primers based on amino acid sequences of the peptides were used for amplification of F26G cDNA fragments by applying nested polymerase chain reactions to cDNAs from in vitro cultured plantlets of C. speciosus. Using primers based on sequences of the cDNA fragments, the 5'- and 3'-end clones were isolated by rapid amplification of cDNA ends (RACE) methods. Finally, the entire coding portion of F26G cDNA was cloned by using primers designed from sequences of the RACE products. The deduced amino acid sequence of CSF26G1, the protein encoded by the cloned cDNA, consists of 562 amino acids and shows high homology to a widely distributed family of beta-glucosidases (BGA family). Cell-free homogenate of Escherichia coli expressing CSF26G1 cDNA showed beta-glucosidase activity specific for cleavage of the C-26 glucosidic bond of furostanol glycosides.

Amino Acid Sequence↗
PubMed1996

The dimerization property of glutathione S-transferase partially reactivates Bcr-Abl lacking the oligomerization domain.

Bcr-Abl oncoproteins are responsible for the pathogenesis of human leukemias with a reciprocal chromosome translocation t(9;22). The amino-terminal Bcr sequence has a potential to form a homotetramer (tetramer domain), and destructions of the tetramer domain cause a complete loss of biological activities in Bcr-Abl. Here we show that Bcr-Abl in which the tetramer domain is replaced with glutathione S-transferase (GST) with a dimerizing ability (GST/Bcr-Abl-(Delta1-160)) can no longer induce an interleukin-3 (IL-3) independence in Ba/F3 cells or transform mouse bone marrow cells but still retains by 30-40% the ability to transform Rat1 cells. Compared with the wild type Bcr-Abl, autophosphorylation of GST/Bcr-Abl-(Delta1-160) in vivo was reduced by more than 50%. The Grb-2 binding to GST/Bcr-Abl-(Delta1-160) was 50% reduced in Rat1 cells and undetectable in Ba/F3 cells. In Rat1 cells expressing GST/Bcr-Abl-(Delta1-160), phosphotyrosine contents of p62 and Shc were 70% decreased.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Expression of vascular endothelial growth factor, its receptor, and other angiogenic factors in human breast cancer.

Angiogenesis is essential for the growth and metastasis of solid tumors. In this study, we examined gene expression of vascular endothelial growth factor (VEGF); its receptor, flt-1; basic fibroblast growth factor; and transforming growth factors (TGFs) alpha and beta in 18 paired cases of human breast carcinomas and the adjacent nonneoplastic tissues. In all of the paired cases, VEGF expression was markedly increased in the carcinomas. In contrast, an insignificant difference was observed in the expression of flt-1, basic fibroblast growth factor, TGF-alpha, and TGF-beta between the malignant breast tissue and the nonneoplastic counterpart. Immunostaining showed variable VEGF positivity of the malignant cells, whereas the nonneoplastic breast epithelial cells were negative. The findings of this study suggest that VEGF is an important angiogenic factor in human breast cancer.

Angiogenesis Inducing Agents↗