Biomedical subjects

Y Oshima

Publications and source records attributed to Y Oshima.

At least 145 records · Page 8Linked to original sources

PubMed1998

Effect of growth factors on expression of integrin subtypes in microvascular endothelial cells isolated from bovine retinas.

We isolated microvascular endothelial cells from bovine retinas (BREC) by a colony isolation method, and investigated the effects of basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) on growth as well as integrin subtype expression of the cultured BREC. Either of the growth factors stimulated growth of the cultured BREC. In non-stimulated BREC, alpha v beta 3 integrin expression was predominant over the other integrin subtypes tested (alpha v beta 5, alpha 2 beta 1 and alpha 5 beta 1), but no enhancement of the alpha v beta 3 expression occurred by either growth factor stimulation. On the other hand, bFGF addition stimulated alpha v beta 5 and alpha 5 beta 1 expression while VEGF induced the alpha 2 beta 1 expression as well. These results suggest that external stimuli such as bFGF and VEGF alter integrin expression profiles of BREC and probably influence interactions of endothelial cells with extracellular matrix during angiogenesis which must be involved in pathogenesis of ischemic retinal disorders. In this respect, this BREC culture system could be useful to study the mechanism of neovascularization in these disorders.

Animals↗

PubMed1997

Isolation and characterization of mutations affecting expression of the delta9- fatty acid desaturase gene, OLE1, in Saccharomyces cerevisiae.

Expression of the delta9- fatty acid desaturase gene, OLE1, of Saccharomyces cerevisiae is negatively regulated transcriptionally and post-transcriptionally by unsaturated fatty acids. In order to isolate mutants exhibiting irregulation of OLE1 expression, we constructed an OLE1p-PHO5 fusion gene as a reporter consisting of the PHO5 gene encoding repressible acid phosphatase (rAPase) under the control of the OLE1 promoter (OLE1p). By EMS mutagenesis, we isolated three classes of mutants, pfo1, pfo2 and pfo3 positive regulatory factor for OLE1) mutants, which show decreased rAPase activity under derepression conditions (absence of oleic acid). Analysis of the transcription of OLE1 in these pfo mutants revealed that pfo1 and pfo3 mutants have a defect in the regulation of OLE1 expression at the transcriptional level while pfo2 mutants were suggested to have a mutation affecting OLE1 expression at a post-transcriptional step. In addition, four other classes of mutants, nfo1, nfo2, nfo3 and nfo4 (negative factor for OLE1) mutants that have mutations causing strong expression of the OLE1p-PHO5 fusion gene under repression conditions (presence of oleic acid), were isolated. Results of Northern analysis of OLE1 as well as OLE1p-PHO5 transcripts in nfo mutants suggested that these mutations occurred in genes encoding global repressors. We also demonstrated that TUP1 and SSN6 gene products are required for full repression of OLE1 gene expression, by showing that either tup1 or ssn6 mutations greatly increase the level of the OLE1 transcript.

Acid Phosphatase↗

PubMed1997

Crystal structure of PHO4 bHLH domain-DNA complex: flanking base recognition.

The crystal structure of a DNA-binding domain of PHO4 complexed with DNA at 2.8 A resolution revealed that the domain folds into a basic-helix-loop-helix (bHLH) motif with a long but compact loop that contains a short alpha-helical segment. This helical structure positions a tryptophan residue into an aromatic cluster so as to make the loop compact. PHO4 binds to DNA as a homodimer with direct reading of both the core E-box sequence CACGTG and its 3'-flanking bases. The 3'-flanking bases GG are recognized by Arg2 and His5. The residues involved in the E-box recognition are His5, Glu9 and Arg13, as already reported for bHLH/Zip proteins MAX and USF, and are different from those recognized by bHLH proteins MyoD and E47, although PHO4 is a bHLH protein.

Amino Acid Sequence↗

PubMed1997

In vitro effects of a recombinant toxin, mSCF-PE40, targeting c-kit receptors ectopically expressed in small cell lung cancers.

Most small cell lung cancers (SCLCs) ectopically express high levels of the c-kit receptor. We have examined if the receptor can serve as a target for a chimeric toxin, mSCF-PE40 composed of murine stem cell factor (SCF) genetically fused to the N terminus of a modified form of Pseudomonas exotoxin (PE) lacking its cell recognition domain. Selective cytotoxicity was found for human c-kit receptor-negative cells. This agent thus warrants further evaluation for therapy of human CSLCs.

ADP Ribose Transferases↗

PubMed1997

The P-OLE1 gene of Pichia angusta encodes a delta 9-fatty acid desaturase and complements the ole1 mutation of Saccharomyces cerevisiae.

Three PCR-amplified DNA fragments hybridizing with the OLE1 gene encoding delta 9-fatty acid desaturase of Saccharomyces cerevisiae were obtained using, respectively, genomic DNAs of one strain each of Kluyveromyces thermotolerans, Pichia angusta and Yarrowia lipolytica as templates. A gene designated P-OLE1 was cloned from the above fragment of P. angusta and sequenced. An open reading frame of P-OLE1 encodes a 49.6-kDa protein consisting of 451 amino acid residues, which shows high identity (62%) and similarity (89%) to that deduced from the OLE1 nucleotide sequence. Expression of P-OLE1 driven by the S. cerevisiae GAP promoter or its own promoter complemented the ole1 mutation of S.cerevisiae. Transcription of P-OLE1 in the native host was suggested to be partially repressed by oleic acid in the medium, as was that of OLE1 in S. cerevisiae and a similar gene in Y. lipolytica, but that of a similar gene in K. thermotolerans was not.

Amino Acid Sequence↗

PubMed1997

Preliminary X-ray studies of a new crystal form of PHO4-DNA complex.

A new crystal form of PHO4 DNA-binding domain-DNA complex was obtained by the truncation of the N-terminal segment. This diffracts to a higher resolution than any forms reported previously. The present crystal belongs to orthorhombic space group P2(1)2(1)2(1), with unit-cell dimensions a=53.51, b=68.30, c= 108.77 A. The crystal contains one protein dimer-DNA complex in the asymmetric unit.

Journal Article↗

PubMed1997

The homeodomain protein Pho2p binds at an A/T-rich segment flanking the binding site of the basic-helix-loop-helix protein Pho4p in the yeast PHO promoters.

Transcription of the genomic PHO5, PHO81 and PHO84 genes of the PHO regulon requires Pho4p and Pho2p activity, whereas transcription of PHO8 is directed by Pho4p alone. Pho4p binds to two 9-bp motifs, 5'-GCACGTGGG-3' (type 1. e.g. UASp2 of PHO5 and site D of PHO84) and 5'-GCACGTTTT-3' (type 2, e.g. UASp1 of PHO5 and site E of PHO84) in the PHO promoter. Experiments were performed to evaluate the ability of these 9-bp motifs to function as upstream activation sites (UASs) by insertion of various 36-bp fragments bearing the 9-bp motif in a CYC1-lacZ fusion gene. No expression of the lacZ gene was detected with the 36-bp fragment bearing UASp2 of PHO5, whereas similar 36-bp fragments bearing UASp1 of PHO5 and sites D and E of PHO84 showed UAS activity in response to Pi concentration in the medium and to the pho2 mutation. The Pho2p-responsive UASs are flanked by one or two copies of an A/T-rich segment, whereas UASp2 is not. Gel retardation and competition experiments performed using a T7-Pho2p-His chimeric protein showed that Pho2p binds to the 36-bp fragments bearing A/T-rich segment(s) but not appreciably to the 36-bp fragments not bearing such segment(s). Thus, the A/T segments flanking the PHO UASs are Pho2p binding sites and play an important role in PHO regulation.

Base Sequence↗

PubMed1997

Synthesis and cataleptic effects of optically active dihydrohaloperidols and dihydrobromoperidols.

Optically active dihydrohaloperidols and dihydrobromoperidols, the major metabolites of haloperidol and bromoperidol, clinically used as neuroleptic drugs in humans, were asymmetrically synthesized by Jaen's method. The motor effects of all the reduced haloperidol and bromoperidol metabolites were evaluated by the mouse catalepsy test. The results indicate that administration of the optically active dihydrohaloperidols and dihydrobromoperidols as well as haloperidol and bromoperidol can cause significant motor deficits in mice.

Animals↗

PubMed1997

New saxitoxin analogues from the freshwater filamentous cyanobacterium Lyngbya wollei.

Along with decarbamoylsaxitoxin and decarbamoylgonyautoxin-2 and -3, six new saxitoxin analogues were isolated from the freshwater mat-forming filamentous cyanobacterium Lyngbya wollei collected from Guntersville Reservoir on the Tennessee River in Alabama. Their structures were determined by electrospray ionization mass spectrometry and several NMR techniques. Five of the toxins contain an acetyl moiety attached to the side chain, which is the first report of these saxitoxin analogues. In three of the toxins a hydrated ketone at C-12 was reduced to alpha-alcohol. The presence of acetate in the side chain resulted in a sevenfold to 17-fold times decrease in mouse toxicity compared to their carbamoyl counterparts, while the reduction at C-12 resulted in a complete loss of mouse toxicity.

Animals↗

PubMed1997

Isoflurane inhibits endothelium-mediated nitric oxide relaxing pathways in the isolated perfused rabbit lung.

PURPOSE: The role of volatile anaesthetics on nitric oxide (NO)-dependent relaxation is unclear in the pulmonary circulation. We examined the effects of isoflurane on NO-dependent relaxation in isolated perfused rabbit lungs. METHODS: Eighteen rabbit lungs were perfused in a constant-flow recirculation manner. In study 1 (n = 12), acetylcholine (ACh, 4 x 10(-10)-10(-8) M) or nitroglycerine (NTG, 6 x 10(-10)-10(-8) M) was cumulatively injected into the pulmonary artery in the absence or presence of isoflurane (1, 2 MAC). In study 2 (n = 6), ACh was injected as in study 1 in the presence or absence of N omega-nitro-L-arginine methyl ester (L-NAME, 100 microM), an NO synthesis blocker. In all experiments, indomethacin was administered to prevent formation of vasoactive prostanoid metabolites, and the pulmonary vessels were preconstricted with prostaglandin F2 alpha (PGF2 alpha) infused before ACh or NTG injection. The ACh- or NTG-induced relaxation was expressed as % decrease in PGF2 alpha preconstriction. RESULTS: Isoflurane at 2 MAC attenuated the dose-dependent relaxation to ACh at doses of 4 x 10(-9) M and 4 x 10(-8) M from 27.8 +/- 4.3% and 38.8 +/- 5.3% to 17.0 +/- 3.5% and 25.5 +/- 4.9%, respectively (P < 0.05). Isoflurane did not change the dose-dependent relaxation to NTG and L-NAME abolished the ACh-induced relaxation. CONCLUSION: Isoflurane inhibited NO-dependent relaxation in the pulmonary circulation at a site distal to the endothelial cell receptor-mediated responses but proximal to guanylate cyclase activation of vascular smooth muscle. Acetylcholine-induced relaxation in isolated perfused rabbit lungs was regulated primarily by NO.

Acetylcholine↗

PubMed1997

Relationship between isocapnic buffering and maximal aerobic capacity in athletes.

This study was performed to clarify the relationship between isocapnic buffering and maximal aerobic capacity (VO2max) in athletes. A group of 15 trained athletes aged 21.1 (SD 2.6) years was studied. Incremental treadmill exercise was performed using a modified version of Bruce's protocol for determination of the anaerobic threshold (AT) and the respiratory compensation point (RC). Ventilatory and gas exchange responses were measured with an aeromonitor and expressed per unit of body mass. Heart rate and ratings of perceived exertion were recorded continuously during exercise. The mean VO2max, oxygen uptake (VO2) at AT and RC were 58.2 (SD 5.8) ml x kg(-1) x min(-1), 28.0 (SD 3.3) ml x kg(-1) x min(-1) and 52.4 (SD 6.7) ml x kg(-1) x min(-1), respectively. The mean values of AT and RC, expressed as percentages of VO2max, were 48.3 (SD 4.2)% and 90.0 (SD 5.2)%, respectively. The mean range of isocapnic buffering defined as VO2 between AT and RC was 24.4 (SD 4.5) ml x kg(-1) x min(-1), and the mean range of hypocapnic hyperventilation (HHV) defined as VO2 between RC and the end of exercise was 5.8 (SD 3.0) ml x kg(-1) x min(-1). The VO2max per unit mass was significantly correlated with AT (r = 0.683, P < 0.01). In addition, VO2max/mass was closely correlated with both the range of isocapnic buffering (r = 0.803, P < 0.001) and RC (r = 0.878, P < 0.001). However, no correlation was found between VO2max per unit mass and the range of HHV (r = 0.011, NS.). These findings would suggest that the prominence of isocapnic buffering, in addition to the anaerobic threshold, may have been related to VO2max of the athletes. The precise mechanisms underlying this proposed relationship remain to be elucidated.

Adult↗

PubMed1997

The role of cysteine residues in the homeodomain protein Mat alpha 2 in mating-type control of Saccharomyces cerevisiae.

The Mat alpha 2 homeodomain protein plays a pivotal role in the control of cell type in Saccharomyces cerevisiae. The homeodomain in the C-terminal region of Mat alpha 2 functions as a DNA-binding domain and the N-terminal region, containing two cysteine residues at positions 33 and 34, is thought to be involved in formation of Mat alpha 2 homodimers via disulfide bonds. mat alpha 2 mutants, isolated in a previous study, in which haploid-specific genes cannot be repressed by the Matal-Mat alpha 2 heterodimer but a-specific genes can be repressed by the Mat alpha 2 homodimer, were found to produce mutant Mat alpha 2 with a substitution of tyrosine or phenylalanine for Cys33. To clarify the role of Cys33 and Cys34 in the Mat alpha 2 protein, we generated several mat alpha 2 mutants by site-directed mutagenesis which had serine residues in place of these Cys residues. Transforming MATa cells with plasmids carrying these mat alpha 2 (MAT alpha 1+) mutations rendered transformants unable to mate. Northern blot analysis revealed that transcription of the a-specific gene STE2 and the haploid-specific locus RME1 in these transformants is repressed to the same level as in wild-type MATa/MAT alpha cells. We concluded that neither Cys33 nor Cys34 is required for repression of a-specific genes by the Mat alpha 2 homodimer or of haploid-specific genes by the Matal-Mat alpha 2 heterodimer, and therefore suggest that Mat alpha 2 homodimer formation in vivo is not mediated by disulfide linkage.

Cysteine↗

PubMed1997

Effects of L-dopa and bromocriptine on haloperidol-induced motor deficits in mice.

L-3,4-Dihydroxyphenylalanine (L-DOPA), the precursor of dopamine, and bromocriptine, a dopamine D2 receptor agonist, were investigated in haloperidol-induced motor impairments in mice using both catalepsy and pole tests. In catalepsy test, subcutaneous treatment with haloperidol (0.125, 0.25 and 0.5 mg/kg) caused a cataleptic effect in mice in a dose-dependent manner. This cataleptic effect was evident upto 7 hr after haloperidol treatment. In pole test, haloperidol (0.125, 0.25 and 0.5 mg/kg) produced the prolongation of Tturn and TLA as a marker of bradykinesia in mice and the prolongation lasted at least 7 hr after haloperidol treatment. Intraperitoneal co-pretreatment with L-DOPA (400 mg/kg) + carbidopa (10 mg/kg) in mice decreased the catalepsy induced by haloperidol at a dose of 0.125 mg/kg, while co-pretreatment with L-DOPA (200 and 400 mg/kg) + carbidopa (10 mg/kg) dose-dependently decreased the haloperidol (0.125 mg/kg)-induced bradykinesia. The effect of LDOPA + carbidopa in pole test was more pronounced than that in catalepsy test. Intraperitoneal pretreatment with bromocriptine (2 and 4 mg/kg) in mice reduced the catalepsy and bradykinesia produced by haloperidol at a dose of 0.125 mg/kg. The effect of bromocriptine in pole test was relatively similar to that in catalepsy test. Also, co-pretreatment with LDOPA (400 mg/kg) + carbidopa (10 mg/kg) and pretreatment with bromocriptine (2 and 4 mg/kg) significantly decreased the catalepsy induced by haloperidol at a higher dose of 0.5 mg/kg. These results indicate that co-administration with L-DOPA + carbidopa and single treatment with bromocriptine can decrease haloperidol-induced catalepsy and bradykinesia in mice. Furthermore, our study suggests that pole test as well as catalepsy test is of value in the screening of drugs against neuroleptic-induced motor deficits.

Animals↗

PubMed1997

Confirmation of anatoxin-a(s), in the cyanobacterium Anabaena lemmermannii, as the cause of bird kills in Danish lakes.

Cyanobacterial blooms were implicated in bird kills at lakes in Denmark in July 1993 and June-July 1994. These blooms were dominated by Anabaena lemmermannii and were shown to contain a neurotoxin with anticholinesterase activity. In this study, the toxin was isolated by mouse lethality guided column chromatographies from the field sample collected at Lake Knud sø in 1993. Various spectroscopic data indicated that the toxin was anatoxin-a(s), an irreversible anticholinesterase, first reported in Anabaena flos-aquae. Chemical detection of the same toxin in cultured A. lemmermannii also confirmed this species as the cause of the deaths of the wild birds.

Anabaena↗

PubMed1997

Comparative study of intraocular lens implantation through 3.0 mm temporal clear corneal and superior scleral tunnel self-sealing incisions.

PURPOSE: To compare the clinical outcome of phacoemulsification and foldable silicone intraocular lens (IOL) implantation through a 3.0 mm temporal clear corneal incision and 3.0 mm superior scleral tunnel incision. SETTING: Department of Ophthalmology, Yodogawa Christian Hospital, Osaka, Japan. METHODS: Eighty cataractous eyes of 78 patients with pre-existing against-the-rule (ATR) astigmatism were recruited for this prospective, randomized study. The patients were assigned to one of the two groups. Data on uncorrected and corrected visual acuities, keratometry, flare intensity measurement, and central cornea endothelial cell count were evaluated preoperatively and at 2 days, 1 week, and 1 and 3 months postoperatively. RESULTS: Although the pre-existing keratometric cylinder decreased in the temporal clear corneal incision group and increased in the superior scleral tunnel incision group, the amount of cylinder shift was not significantly different. Mean scalar shift of keratometric cylinder in the corneal incision group was 1.19 diopters (D) at 2 days postoperatively, 0.86 D at 1 week, and 0.56 D at 3 months and in the scleral incision group, 1.09 D at 2 days, 0.76 D at 1 week, and 0.65 D at 3 months. Eighty percent of the eyes in each group achieved an uncorrected visual acuity of 20/40 or better from the second day postoperatively. No statistically significant difference in visual rehabilitation or other parameters was noted between the groups throughout the study. Complications including corneal endothelial cell loss and wound incompetence requiring suturing were observed in the temporal clear corneal incision group. CONCLUSIONS: Both incisions offered satisfactory clinical results, but the superior scleral tunnel incision resulted in fewer complications. Minimal corneal keratometric change induced by a 3.0 mm incision was not related to uncorrected visual rehabilitation.

Aged↗

PubMed1997

Fatal GVHD demonstrating an involvement of respiratory muscle following donor leukocyte transfusion (DLT).

A 41-year-old female patient with AML, who relapsed after an allogeneic BMT from her HLA-identical sister, was treated by a donor leukocyte transfusion (DLT). Thereafter, bone marrow aplasia accompanied by the disappearance of leukemic blasts following the GVHD was observed. The patient died of chronic GVHD with respiratory muscle involvement 19 months after the DLT. Although the DLT was considered helpful in suppressing the proliferation of the leukemic cells, it might also have caused the severe GVHD observed in this case. Efforts to separate the lymphocyte clones responsible for GVL from those for the GVHD thus appear to be necessary for the further development of the therapeutic approach, so-called DLT.

Adult↗

PubMed1997

Effect of electric pulses and antiproliferative drugs on cultured bovine retinal pigment epithelial cells.

PURPOSE: The effect of antiproliferative agents with high voltage electric pulses (EP) on the proliferation of retinal pigment epithelial (RPE) cells was investigated. METHODS: Bovine RPE cells were cultured and exposed briefly to an antiproliferative agent: 0.005 to 500 microM of bleomycin (BLM), 0.005 to 500 microM of mitomycin C (MMC) or 0.05 to 5000 microM of 5-fluorouracil (5-FU) with or without high voltage electric pulses (EP: 2,000V/cm, 98 microseconds, 8 pulses). Streptomycin (SM) was used as a negative control. Cell proliferation with or without antiproliferative agents was assessed on day 3. DNA fragmentation was assessed by flow cytometric analysis. RESULTS: Treatment with BLM, MMC or 5-FU inhibited cell proliferation in a dose-dependent manner, with and without EP (p < 0.05). EP enhanced the inhibitory effect of BLM as much as 3,000 times (ID50: BLM without EP; 163.7 microM, BLM with EP; 0.0574 microM, MMC without EP; 132.4 microM, MMC with EP; 26.2 microM, 5-FU without EP; 616.4 microM, 5-FU with EP; 873.8 microM). EP treatment showed an inhibitory effect of BLM on cell proliferation more prominently than BLM alone. The cell numbers of 0.5 microM BLM treatment without EP were 87.4 +/- 11.7% (mean +/- SD), whereas the cell numbers of 0.5 microM BLM with EP were 21.1 +/- 2.16% (p < 0.005). BLM treatment with EP increased the apoptosis like DNA fragmentation in the flow cytometric DNA histogram, showing dominant accumulation in the A0 region, which is the population of DNA fragmentation. (The cell population of the A0 region: control, 3.4%; EP alone, 18.9%; BLM treatment without EP, 32.0%; with EP, 93.8%.) CONCLUSIONS: EP treatment enhances the inhibitory effect of BLM on RPE cell proliferation. The combination of electric pulse and BLM treatment can localize the drug effect and reduce the necessary dose of the antiproliferative agent in comparison to BLM treatment alone.

Animals↗

PubMed1997

The transcriptional activators of the PHO regulon, Pho4p and Pho2p, interact directly with each other and with components of the basal transcription machinery in Saccharomyces cerevisiae.

The transcriptional regulators Pho4p and Pho2p are involved in transcription of several genes in the PHO regulon of Saccharomyces cerevisiae. Genetic evidence with temperature-sensitive pho4 and pho2 mutants suggested that Pho4p and Pho2p interact with each other. Immunoprecipitation experiments showed that Pho4p and Pho2p form a complex on a 36-bp sequence bearing an upstream activation site (UAS) and protein binding assays indicated that these proteins interact directly. DNA-binding experiments with crude extracts prepared from yeast strains expressing T7-PHO4, encoding Pho4p tagged with the T7 epitope, indicated that Pho2p interacts with T7-Pho4p and enhances the binding affinity of T7-Pho4p to the UAS. Protein binding experiments also showed that both Pho4p and Pho2p could bind with the general transcription factors, TBP, TFIIB, and TFIIEbeta, suggesting that the Pho4p-Pho2p complex bound to the UAS activates transcription of the PHO genes by direct interaction with the general transcription factors.

Basal Metabolism↗