Biomedical subjects

E Suzuki

Publications and source records attributed to E Suzuki.

At least 37 records · Page 2Linked to original sources

PubMed2002

High-resolution scanning electron microscopy of immunogold-labelled cells by the use of thin plasma coating of osmium.

The feasibility of plasma coating of a thin osmium layer for high-resolution immuno-scanning electron microscopy of cell surfaces was tested, using Drosophila embryonic motor neurones as a model system. The neuro-muscular preparations were fixed with formaldehyde and labelled with a neurone-specific antibody and 10 or 5 nm colloidal gold-conjugated secondary antibodies. The specimens were post-fixed with osmium tetroxide and freeze-dried. Then they were coated with a 1-2 nm thick layer of osmium using a hollow cathode plasma coater. The thin and continuous coating of amorphous osmium gave good signals of gold particles and fine surface structures of neurites in backscattered electron images simultaneously. This method makes it possible to visualize the antigen distribution and the three-dimensionally complex surface structures of cellular processes with a resolution of several nanometres.

Animals↗

PubMed2001

CD40 ligand promotes priming of fully potent antitumor CD4(+) T cells in draining lymph nodes in the presence of apoptotic tumor cells.

The presence or absence of CD4(+) T cell help can determine the direction of adaptive immune responses toward either cross-priming or cross-tolerance. It has been demonstrated that interactions of CD40-CD40 ligand can replace CD4(+) T cell help and enable dendritic cells to prime cytotoxic T cells. Here, we demonstrate that antitumor reactivity induced in regional lymph nodes (LNs) by s.c. injection of CD40 ligand (CD40L)-transduced tumor (MCA205 CD40L) showed far superior therapeutic efficacy against established brain tumors of a weakly immunogenic fibrosarcoma, MCA205, when adoptively transferred. Coinjection of apoptotic, but not necrotic parental tumor cells with CD40L-expressing tumor cells caused a strong synergistic induction of antitumor reactivity in tumor-draining LNs. Freshly isolated T cells from LNs immunized with apoptotic parental tumor cells and MCA205 CD40L were capable of mediating regression of the parental tumor in vivo. In contrast, T cells derived from LNs immunized without MCA205 CD40L required ex vivo anti-CD3/IL-2 activation to elicit therapeutic activity. On anti-CD3/IL-2 activation, cells from LNs immunized with MCA205 CD40L exhibited superior per cell antitumor reactivity. An in vitro depletion study revealed that either CD4(+) or CD8(+) T cells could mediate therapeutic efficacy but that the antitumor efficacy mediated by CD4(+) T cells was far superior. Cytosolic flow cytometric analyses indicated that priming of CD4(+) cells in LNs draining CD40L-expressing tumors was polarized to the Th1 type. This is the first report that fully potent antitumor CD4(+) T cell priming was promoted by s.c. injection of CD40L-transduced tumor in the presence of apoptotic tumor cells.

Animals↗

PubMed2001

Deoligomerization of siloxanes with dimethyl carbonate over solid-base catalysts.

Hexamethyldisiloxane and hexamethylcyclotrisiloxane were almost completely deoligomerized with dimethyl carbonate over alumina-supported potassium fluoride catalyst to form methoxytrimethylsilane (85% yield) and dimethoxydimethylsilane (94% yield), respectively, and moreover the addition of a small amount of methanol to the hexamethyldisiloxane deoligomerization reaction enhanced the methoxytrimethylsilane yield to 98%.

Journal Article↗

PubMed2001

Endothelial nitric oxide synthase is essential for the HMG-CoA reductase inhibitor cerivastatin to promote collateral growth in response to ischemia.

HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) reductase inhibitors, or statins, are prescribed widely to lower cholesterol. Accumulating evidence indicates that statins have various effects on vascular cells, which are independent of their lipid-lowering effect. Here, we tested the hypothesis that statins may augment collateral flow to ischemic tissues. We induced hind-limb ischemia in wild-type mice and treated them with either saline or cerivastatin. Cerivastatin enhanced the blood flow recovery dramatically as determined by Laser Doppler imaging. The mice treated with saline displayed frequent autoamputation of the ischemic toe, which was prevented completely by cerivastatin. Anti-CD31 immunostaining revealed that cerivastatin significantly increased the capillary density. Endothelial nitric oxide synthase (eNOS) activity was enhanced markedly in the mice treated with cerivastatin. The angiogenic effect of cerivastatin was abrogated in eNOS deficient (eNOS-/-) mice. These results indicate that eNOS is essential for cerivastatin to promote collateral growth in response to ischemia.

Animals↗

PubMed2001

Enantioselective fluorination mediated by cinchona alkaloid derivatives/Selectfluor combinations: reaction scope and structural information for N-fluorocinchona alkaloids.

Cinchona-alkaloid/Selectfluor combinations efficiently fluorinate a variety of carbonyl compounds in a highly enantioselective manner to furnish chiral alpha-fluorocarbonyl compounds. The DHQB/Selectfluor combination is effective for the enantioselective fluorination of indanones and tetralones 1 in up to 91% ee. The first enantioselective syntheses of chiral derivatizing reagents 3 was accomplished with high ee and in high chemical yields by the DHQDA/Selectfluor combination. 3-Fluorooxindoles 7 were prepared with ee up to 83% using the (DHQ)2AQN/Selectfluor or the (DHQD)2PYR/Selectfluor combination. Since the combinations are conveniently prepared in situ from readily available reagents, the present system represents a practical method for enantioselective fluorination. X-ray crystallography and 1H NMR analyses of the cinchona alkaloids/Selectfluor combination have established that the species that mediate this novel reaction are N-fluoroammonium cinchona alkaloid tetrafluoroborates, which adopt open conformations.

Cinchona Alkaloids↗

PubMed2001

Adrenomedullin induces endothelium-dependent vasorelaxation via the phosphatidylinositol 3-kinase/Akt-dependent pathway in rat aorta.

To study the mechanisms by which adrenomedullin (AM) induces endothelium-dependent vasorelaxation, we examined whether AM-induced endothelium-dependent vasodilation was mediated by the phosphatidylinositol 3-kinase (PI3K)/Akt-dependent pathway in rat aorta, because it was recently reported that PI3K/Akt was implicated in the activation of endothelial NO synthase. AM-induced vasorelaxation in thoracic aorta with intact endothelium was inhibited by pretreatment with PI3K inhibitors to the same level as that in endothelium-denuded aorta. AM elicited Akt phosphorylation in a time- and dose-dependent manner. AM-induced Akt phosphorylation was inhibited by pretreatment with a calmodulin-dependent protein kinase inhibitor as well as with PI3K inhibitors. When an adenovirus construct expressing a dominant-negative Akt mutant (Ad/dnAkt) was injected into abdominal aortas so that the mutant was expressed predominantly in the endothelium layer, AM-induced vasodilation was diminished to the same level as that in endothelium-denuded aortas. Finally, AM-induced cGMP production, which was used as an indicator for NO production, was suppressed by PI3K inhibition or by Ad/dnAkt infection into the endothelium. These results suggested that AM induced Akt activation in the endothelium via the Ca(2+)/calmodulin-dependent pathway and that this was implicated in the production of NO, which in turn induced endothelium-dependent vasodilation in rat aorta.

Adrenomedullin↗

PubMed2001

Oxidation of activated carbon and methane using a high-frequency pulsed plasma.

We have demonstrated a new method using a high-frequency pulsed plasma for oxidizing methane and activated carbon powder (1-150 microm) as two substitutes for diesel particulate matter (PM). Methane and activated carbon can be oxidized at a rate of 4-40 and 4-10g-CH(4)/kWh, respectively. A discussion on future development of this technology is given.

Air Pollution↗

PubMed2001

Comparison of the elimination between perfluorinated fatty acids with different carbon chain length in rats.

Elimination in urine and feces was compared between four perfluorinated fatty acids (PFCAs) with different carbon chain length. In male rats, perfluoroheptanoic acid (PFHA) was rapidly eliminated in urine with the proportion of 92% of the dose being eliminated within 120 h after an intraperitoneal injection. Perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA) and perfluorodecanoic acid (PFDA) was eliminated in urine with the proportions of 55, 2.0 and 0.2% of the dose, respectively. By contrast, four PFCAs were eliminated in feces with the proportion of less than 5% of the dose within 120 h after an injection. In female rats, the proportions of PFOA and PFNA eliminated in urine within 120 h were 80% and 51% of the dose, respectively, which were significantly higher compared with those in male rats. There was the tendency that PFCA with longer carbon chain length is less eliminated in urine in both male and female rats. Fecal elimination of PFCAs was not different between PFCAs in female rats and comparable to those in male rats. The rates of biliary excretion of PFCAs in male rats were slower than those in female rats. Sex-related difference in urinary elimination of PFOA was abolished when male rats had been castrated. On the contrary, treatment with testosterone suppressed the elimination of PFOA in urine in both castrated male rats and female rats. The effect of testosterone was in a time- and dose-dependent manner. These results suggest that PFCAs are distinguished by their carbon chain length by a renal excretion system, which is regulated by testosterone.

Animals↗

PubMed2001

Transcriptional activation of the cyclin D1 gene is mediated by multiple cis-elements, including SP1 sites and a cAMP-responsive element in vascular endothelial cells.

In an attempt to examine the mechanisms by which transcriptional activity of the cyclin D1 promoter is regulated in vascular endothelial cells (EC), we examined the cis-elements in the human cyclin D1 promoter, which are required for transcriptional activation of the gene. The results of luciferase assays showed that transcriptional activity of the cyclin D1 promoter was largely mediated by SP1 sites and a cAMP-responsive element (CRE). DNA binding activity at the SP1 sites, which was analyzed by electrophoretic mobility shift assays, was significantly increased in the early to mid G(1) phase, whereas DNA binding activity at CRE did not change significantly. Furthermore, Induction of the cyclin D1 promoter activity in the early to mid G(1) phase depended largely on the promoter fragment containing the SP1 sites, whereas the proximal fragment containing CRE but not the SP1 sites was constitutively active. Finally, the increase in DNA binding and promoter activities via the SP1 sites was mediated by the Ras-dependent pathway. The results suggested that the activation of the cyclin D1 gene in vascular ECs was regulated by a dual system; one was inducible in the G(1) phase, and the other was constitutively active.

Activating Transcription Factor 1↗

PubMed2001

Independent anchoring and assembly mechanisms of INAD signaling complexes in Drosophila photoreceptors.

In Drosophila photoreceptors the multivalent PDZ protein INAD organizes the phototransduction cascade into a macromolecular signaling complex containing the effector PLC, the light-activated TRP channels, and a regulatory PKC. Previously, we showed that the subcellular localization of INAD signaling complexes is critical for signaling. Now we have examined how INAD complexes are anchored and assembled in photoreceptor cells. We find that trp mutants, or transgenic flies expressing inaD alleles that disrupt the interaction between INAD and TRP, cause the mislocalization of the entire transduction complex. The INAD-TRP interaction is not required for targeting but rather for anchoring of complexes, because INAD and TRP can be targeted independently of each other. We also show that, in addition to its scaffold role, INAD functions to preassemble transduction complexes. Preassembly of signaling complexes helps to ensure that transduction complexes with the appropriate composition end up in the proper location. This may be a general mechanism used by cells to target different signaling machinery to the pertinent subcellular location.

Animals↗

PubMed2001

Effects of endocrine disrupting chemicals on the microtubule network in Chinese hamster V79 cells in culture and in Sertoli cells in rats.

The effects of ten endocrine disrupting chemicals, i.e., bisphenol A (BPA), p-nonylphenol (NP), p-octylphenol (OP), p-pentylphenol (PP), butyl benzyl phthalate (BBP), dicyclohexyl phthalate (DCHP), di-n-butyl phthalate (DBP), tetrabutyltin (TBT), tri-n-butyltin chloride (TBC), and di-n-butyltin dichloride (DBD), as well as 17 beta-estradiol (E(2)) as a positive control on the microtubule network in Chinese hamster V79 cells in culture were examined by the indirect immunofluorescence method using anti-beta-tubulin antibody. In the whole-animal system, the effects of BPA, NP, OP, BBP, DBD, and E(2) as well as vinblastine sulfate (VB) as a positive control on microtubules in the cytoplasm of Sertoli cells in rats were examined by electron microscopy. In Chinese hamster V79 cells, TBC and DBD showed higher microtubule-disruptive activity than E(2), while other chemicals had less activity than E(2). The ranking for efficiency on microtubule disruption was (TBC falling dots DBD) > (E(2) = TBT) > (BPA = alkylphenols, NP and OP) >> (phthalate esters, BBP, DHP, and DBP). In rats as a whole-animal system, no disrupting effects on the microtubule network in the cytoplasm of Sertoli cells were observed under any environmental chemicals tested, whereas exposure to VB resulted in marked disruption of the microtubule network. The results of this study suggested that some endocrine disrupting chemicals have disrupting effects on the microtubule network in vitro, but no such effects in vivo.

Animals↗

PubMed2001

Inhibitory role of eosinophils on cell surface plasmin generation by bronchial epithelial cells: inhibitory effects of transforming growth factor beta.

Eosinophilic bronchitis is an essential component of bronchial asthma, and eosinophils play an important role. We studied the effect of eosinophils on cell surface plasmin generation by bronchial epithelial cells, because plasmin is thought to be involved in bronchial tissue repair/remodeling by means of fibrinolysis and the activation of proteases such as matrix metalloproteases. Plasmin was generated from exogenous plasminogen on the cell surface of cultured bronchial epithelial cells, NCI-H292. Transforming growth factor beta (TGF-beta) treatment resulted in reduced cell surface plasmin generation and a large increase in plasminogen activator inhibitor-type 1 (PAI-1) antigen production in NCI-H292 cells, whereas no conspicuous effects were observed with IL-1 beta and TNF alpha treatment (regulators in pulmonary epithelial cells). On the other hand, this cell surface plasmin generation was reduced by co-incubation with Eol-1, an eosinophil cell line. The addition of TGF-beta antisense and anti-TGF-beta antibodies attenuated this adverse effect of Eol-1 cell co-incubation. These data suggest that eosinophils play an inhibitory role on cell surface plasmin generation by bronchial epithelial cells by means of the up-regulation of PAI-1 expression induced by TGF-beta. Therefore, the accumulation of eosinophils in bronchial walls is thought to be involved in bronchial tissue repair/remodeling in asthma through this protease network.

Asthma↗

PubMed2001

Inhibition of macrophage invasion by monoclonal antibodies specific to Leishmania (Viannia) braziliensis promastigotes and characterisation of their antigens.

Monoclonal antibodies that specifically recognise Leishmania (Viannia) braziliensis promastigotes were produced and termed SST-2, SST-3 and SST-4. SST-2 recognises a conformational epitope present in a 24-28 kDa doublet and in a 72 kDa component, as verified by Western blotting. Indirect immunofluorescence showed that the antigen recognised by SST-2 is distributed homogeneously on the parasite surface. SST-3 recognises a flagellar glycoprotein of approximately 180 kDa. The reactivity of this mAb was abolished by sodium m-periodate treatment, indicating that SST-3 reacts with a carbohydrate epitope of the 180 kDa antigen. SST-4 recognises a conformational epitope of a 98 kDa antigen. SST-2, SST-3 and SST-4 were specific to L. (V.) braziliensis promastigote forms. Indirect immunofluorescence did not show reactivity of SST-2 or SST-3 with amastigotes of L. (V.) braziliensis, or with promastigotes of Leishmania (Viannia) panamensis, Leishmania (Viannia) guyanensis, Leishmania (Viannia) naiffi, Leishmania (Viannia) lainsoni, Leishmania (Leishmania) amazonensis, Leishmania (Leishmania) major, or Leishmania (Leishmania) chagasi. We also evaluated the involvement of SST-2, SST-3 and SST-4 antigens in parasite-macrophage interaction. Fab fragments of SST-3 and SST-4 significantly inhibited the infectivity of L. (V.) braziliensis promastigotes to mouse peritoneal macrophages.

Animals↗

PubMed2001

Elevated plasma nitrate levels in depressive states.

BACKGROUND: Previous studies have shown that nitric oxide (NO) synthase inhibitors show preclinical antidepressant-like properties, suggesting that NO is involved in the pathogenesis of depression. The purpose of this study is to examine whether or not NO production increases in depressed patients. METHODS: Plasma nitrate concentrations, an index of NO production, were measured by high-performance liquid chromatography in depressed patients (n=17) and compared with patients suffering anxiety (n=6) and with healthy controls (n=12). RESULTS: Plasma nitrate concentrations were significantly higher in depressed patients than in patients with an anxiety disorder (P<0.05) or in controls (P<0.01). LIMITATIONS: The study group was small. The source of the surplus production of NO in patients with major depressive episode remains unclear. CONCLUSIONS: These results suggest that NO production is increased in depression.

Adult↗

PubMed2001

Effects of intracavernous administration of adrenomedullin on erectile function in rats.

We have reported that adrenomedullin (AM)-induced vasodilation is at least in part nitric oxide (NO)-cGMP-dependent in the rat. Although it is well known that NO is much involved in the erectile function, it is controversial as to whether AM influences the erectile function. Thus, we examined the effects of AM on intracavernous pressure (ICP) during penile erection. The left carotid artery of rats was cannulated to monitor of mean arterial pressure (MAP). Bipolar electrodes were positioned on the cavernous nerve. The right cavernous body was cannulated with a needle connected to a pressure transducer to monitor ICP. Electrical stimulation (ES) increased ICP in a voltage-dependent manner. Elevation of ICP continued during ES. The intracavernous injection of 0.5 nmol AM significantly potentiated ES-induced increases in both maximal developed ICP/MAP and area under the curve (ICP trace; AUC). Since AM slightly lowered MAP, ICP was normalized by MAP. i.v. administration of N(omega)-nitro-L-arginine, a NO synthase inhibitor, markedly decreased AM/ES-induced ICP elevation. However, in the presence of E-4021, a cGMP-specific phosphodiesterase inhibitor, AM further increased both ICP/MAP and AUC. These results suggest that a NO-cGMP pathway is involved in the regulation of AM-induced rat cavernous vasorelaxation.

Adrenomedullin↗

PubMed2001

Polymorphism in RANTES chemokine promoter affects extent of sarcoidosis in a Japanese population.

RANTES, a member of C-C chemokine, is known to be produced at sites of granulomatous reactions in the lung of sarcoidosis. RANTES is a potent eosinophil and lymphocyte attractant with particular preference for CD45RO+ T cells and eosinophils. Polymorphism of the RANTES promoter has recently been shown to be related to allergic and infectious diseases; atopic dermatitis, asthma, and polymyalgia rheumatica. Considering that this might affect sarcoidosis, we studied polymorphism of the RANTES gene in 114 patients with sarcoidosis and 136 healthy control subjects. Their genotypes were determined using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP). Although no difference in the genotype distribution between healthy control subjects and sarcoidosis patients was identified, the difference in the frequencies of the patients with three or more organ involvement was significant (P<0.01) with the frequency of those in AA genotype being elevated (P<0.05). BAL findings in 48 out of 114 patients who underwent bronchoscopy were reviewed. The CD4/8 ratio of lymphocytes in bronchoalveolar lavage fluid in the patients with AA genotype was significantly increased (P<0.05). From the results, we suggest that in RANTES gene polymorphism the homozygous A allele might be a genetic risk factor for extent disease of sarcoidosis.

Bronchoalveolar Lavage Fluid↗

PubMed2001

Characterization of monoclonal antibody MEST-2 specific to glucosylceramide of fungi and plants.

An IgG2a monoclonal antibody anti-glucosylceramide was established and termed MEST-2. High performance thin layer chromatography immunostaining, and solid-phase radioimmunoassay showed that MEST-2 reacts with glucosylceramide from yeast and mycelium forms of Paracoccidioides brasiliensis, Histoplasma capsulatum, and Sporothrix schenckii; from hyphae of Aspergillus fumigatus; and from yeast forms of Candida albicans, Cryptococcus neoformans, Cryptococcus laurentii, and Cryptococcus albidus. Studies on the fine specificity of MEST-2 showed that it recognizes the beta-D-glucose residue, and that the 2-hydroxy group present in the fatty acid is an important auxiliary feature for the antibody binding. It was also demonstrated that phosphatidylcholine and ergosterol modulate MEST-2 reactivity to glucosylceramide, by solid-phase radioimmunoassay. Indirect immunofluorescence showed that MEST-2 reacts with the surface of yeast forms of P. brasiliensis, H. capsulatum and S. schenckii. Weak staining of mycelial forms of P. brasiliensis and hyphae of A. fumigatus was also observed. The availability of a monoclonal antibody specific to fungal glucosylceramide, and its potential use in analyzing biological roles attributed to glucosylceramide in fungi are discussed.

Antibodies, Monoclonal↗

PubMed2001

Characterization of cerebrosides from the thermally dimorphic mycopathogen Histoplasma capsulatum: expression of 2-hydroxy fatty N-acyl (E)-Delta(3)-unsaturation correlates with the yeast-mycelium phase transition.

Cerebroside (monohexosylceramide) components were identified in neutral lipids extracted from both the yeast and mycelial forms of the thermally dimorphic mycopathogen Histoplasma capsulatum. The components were purified from both forms and their structures elucidated by 1- and 2-dimensional nuclear magnetic resonance (NMR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and low energy tandem collision-induced dissociation mass spectrometry (ESI-MS/CID-MS). Both components were characterized as beta-glucopyranosylceramides (GlcCers) containing (4E,8E)-9-methyl-4,8-sphingadienine as the long-chain base, attached to 18-carbon 2-hydroxy fatty N-acyl components. However, while the fatty acid of the yeast form GlcCer was virtually all N-2'-hydroxyoctadecanoate, the mycelium form GlcCer was characterized by almost exclusive expression of N-2'-hydroxy-(E)-delta(3)-octadecenoate. These results suggest that the yeast-mycelium transition is accompanied by up-regulation of an as yet uncharacterized ceramide or cerebroside 2-hydroxy fatty N-acyl (E)-delta(3)-desaturase activity. They also constitute further evidence for the existence of two distinct pathways for ceramide biosynthesis in fungi, since glycosylinositol phosphorylceramides (GIPCs), the other major class of fungal glycosphingolipids, are found with ceramides consisting of 4-hydroxysphinganine (phytosphingosine) and longer chain 2-hydroxy fatty acids. In addition to identification of the major glucocerebroside components, minor components (< 5%) detectable by molecular weight differences in the ESI-MS profiles were also characterized by tandem ESI-MS/CID-MS analysis. These minor components were identified as variants differing in fatty acyl chain length, or the absence of the sphingoid 9-methyl group or (E)-delta(8)-unsaturation, and are hypothesized to be either biosynthetic intermediates or the result of imperfect chemical transformation by the enzymes responsible for these features. Possible implications of these findings with respect to chemotaxonomy, compartmentalization of fungal glycosphingolipid biosynthetic pathways, and regulation of morphological transitions in H.capsulatum and other dimorphic fungi are discussed.

Carbohydrate Conformation↗