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  • 1
    Electronic Resource
    Electronic Resource
    Purification and Characterization of a Trypsin-Like Enzyme with Fibrinolytic Activity Present in the Abdomen of Horn Fly, Haematobia irritans irritans (Diptera: Muscidae) (2000)
    Springer
    The protein journal 19 (2000), S. 515-521 
    Details
    ISSN: 1573-4943
    Keywords: trypsin-like enzyme ; fibrinolytic activity ; protein purification ; hematophagous ; Haematobia irritans irritans
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract This work describes the purification and characterization of a trypsin-like enzyme with fibrinolytic activity present in the abdomen of Haematobia irritans irritans (Diptera: Muscidae). The enzyme was purified using a one-step process, consisting of affinity chromatography on SBTI-Sepharose. The purified protease showed one major active proteinase band on reverse zymography with 0.15% gelatin, corresponding to a molecular mass of 25.5 kDa, with maximum activity at pH 9.0. The purified trypsin-like enzyme preferentially hydrolyzed synthetic substrates with arginine residue at the P1 position. The K m values determined for three different substrates were 1.88 × 10−4, 1.28 × 10−4, and 1.40 × 10−4 M for H-α-benzoyl-Ile-Glu-Gly-Arg-p-nitroanilide (S2222), dl-Ile-Pro-Arg-p-nitroanilide (S2288), and DL-Phe-Pip-Arg-p-nitroanilide (S2238), respectively. The enzyme was strongly inhibited by typical serine proteinase inhibitors such as SBTI (soybean trypsin inhibitor, K i = 0.19 nM) and BuXI (Bauhinia ungulata factor Xa inhibitor, K i = 0.48 nM), and less inhibited by LDTI (leech-derived tryptase inhibitor, K i = 1.5 nM) and its variants LDTI 2T and 5T (0.8 and 1.5 nM, respectively). The most effective inhibitor for this protease was r-aprotinin (r-BPTI) with a K i value of 39 pM. Synthetic serine protease inhibitors presented only weak inhibition, e.g., benzamidine with K i = 3.0 × 10−4 M and phenylmethylsulfonyl fluoride (PMSF) showed traces of inhibition. The purified trypsin-like enzyme also digested natural substrates such as fibrinogen and fibrin net. The protease showed higher activity against fibrinogen and fibrin than did bovine trypsin. These data suggest that the proteolytic enzyme of H. irritans irritans is more specific to proteins from blood than are the vertebrate digestive enzymes. This enzyme's characteristics may be an adaptation resulting from the feeding behavior of this hematophagous insect.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1023/A:1026557600429
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  • 2
    Electronic Resource
    Electronic Resource
    Experimental derivation of Auger transition probabilities from X-ray excited Auger electron spectra in XPS (2000)
    Springer
    Fresenius' journal of analytical chemistry 366 (2000), S. 30-35 
    Details
    ISSN: 1432-1130
    Source: Springer Online Journal Archives 1860-2000
    Topics: Chemistry and Pharmacology
    Notes: Abstract Auger transition probabilities were experimentally derived from dominant XAES and related XPS peaks observed in XPS spectra. Some values of derived probabilities were higher than 1, because of addition or subtraction of background signal from the XAES or/and XPS peak intensity. However, the probabilities obtained are recognized to be useful for practical quantification by XAES and AES.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002160050007
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  • 3
    Electronic Resource
    Electronic Resource
    Construction of an engineered yeast with glucose-inducible emission of green fluorescence from the cell surface (2000)
    Springer
    Applied microbiology and biotechnology 54 (2000), S. 90-96 
    Details
    ISSN: 1432-0614
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: Abstract An engineered yeast with emission of fluorescence from the cell surface was constructed. Cell surface engineering was applied to display a visible reporter molecule, green fluorescent protein (GFP). A glucose-inducible promoter GAPDH as a model promoter was selected to control the expression of the reporter gene in response to environmental changes. The GFP gene was fused with the gene encoding the C-terminal half of α-agglutinin of Saccharomyces cerevisiae having a glycosylphosphatidylinositol anchor attachment signal sequence. A secretion signal sequence of the fungal glucoamylase precursor protein was connected to the N-terminal of GFP. This designed gene was integrated into the TRP1 locus of the chromosome of S. cerevisiae with homologous recombination. Fluorescence microscopy demonstrated that the transformant cells emitted green fluorescence derived from functionally expressed GFP involved in the fusion molecule. The surface display of GFP was further verified by immunofluorescence labeling with a polyclonal antibody (raised in rabbits) against GFP as the first antibody and Rhodamine Red-X-conjugated goat anti-rabbit IgG as the second antibody which cannot penetrate into the cell membrane. The display of GFP on the cell surface was confirmed using a confocal laser scanning microscope and by measuring fluorescence in each cell fraction obtained after the subcellular fractionation. As GFP was proved to be displayed as an active form on the cell surface, selection of promoters will endow yeast cells with abilities to respond to changes in environmental conditions, including nutrient concentrations in the media, through the emission of fluorescence.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s002539900307
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  • 4
    Electronic Resource
    Electronic Resource
    Suplatast tosilate inhibits thymus- and activation-regulated chemokine production by antigen-specific human Th2 cells (2002)
    Oxford, UK : Blackwell Science Ltd
    Clinical & experimental allergy 32 (2002), S. 0 
    Details
    ISSN: 1365-2222
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Background Suplatast tosilate is an anti-allergic agent that suppresses cytokine production by human Th2 cells.Objective We investigated the effects of suplatast tosilate on the production of thymus- and activation-regulated chemokine (TARC) by T cells from allergic patients with asthma.Methods Purified protein derivative (PPD)-specific Th1 cell lines and Dermatophagoides farinae (Der f)-specific Th2 cell lines were established from nine patients with house dust mite-allergic asthma. The effects of suplatast tosilate on mRNA expression of TARC and protein production of TARC from antigen-specific Th1 or Th2 cell lines were investigated after stimulation with relevant antigens or phytohemagglutinin (PHA). In addition, the effects of IL-4, IL-10, and IFN-γ on TARC production by Der f-specific Th2 cell lines in the presence or absence of suplatast tosilate were studied.Results Although PPD-specific Th1 cell lines did not produce TARC after stimulation with PPD antigen or PHA, stimulation of Der f-specific Th2 cell lines with Der f antigen or PHA increased production of TARC. Suplatast tosilate significantly and dose-dependently inhibited production of TARC by Der f-specific Th2 cell lines stimulated with either Der f antigen (76.5% inhibition at 100 µg/mL, P 〈 0.01) or PHA (81.9% inhibition at 100 µg/mL, P 〈 0.01). TARC production by Der f-specific Th2 cell lines was significantly increased only by activation with IL-4 but not with IL-10 or IFN-γ; this increase in TARC production was significantly inhibited by suplatast tosilate (97.5% inhibition at 100 µg/mL, P 〈 0.01).Conclusion Suplatast tosilate inhibits TARC production by human Th2 cells. Therefore, this agent inhibits both Th2 cytokine and Th2 chemokine and may be a useful anti-allergic agent.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2222.2002.01547.x
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  • 5
    Electronic Resource
    Electronic Resource
    Selection of the powdery metal hydride best for producing H− by thermal desorption : The 8th international conference on ion sources (2000)
    [S.l.] : American Institute of Physics (AIP)
    Review of Scientific Instruments 71 (2000), S. 853-855 
    Details
    ISSN: 1089-7623
    Source: AIP Digital Archive
    Topics: Physics , Electrical Engineering, Measurement and Control Technology
    Notes: To find the metal hydride best for producing H− by thermal desorption, the desorption rates of H−, H2 and electron (e−) directly emitted from a powdery sample (∼1 mg) of NaH, LiH, MgH2, CaH2, SrH2, TiH2, ZrH2, KBH4, LiAlH4, or NaAlH4 heated up to ∼1000 K were determined simultaneously with a special system. Theoretical analysis of the experimental data thus achieved yields the following results. (1) Both H− and e− are emitted from those active spots (mainly consisting of alkali or alkali earth metal) created by thermal decomposition (e.g., LiH→Li+H2/2). (2) The active spots are readily destroyed and reconstructed by admission of H2 and by stopping the admission, respectively. (3) The work function (φ) of activated NaH is very low (∼2 eV), but NaH is rapidly depleted owing to its thermal instability. (4) Among the ten hydrides, CaH2 is concluded to be the best as a source material for thermal desorption of H− because activated CaH2 (φ(similar, equals)5 eV) is most stable and strongest in H− (∼10−12 A or 10−10 A/cm2 after mass analysis) around 900 K. © 2000 American Institute of Physics.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1063/1.1150311
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  • 6
    Electronic Resource
    Electronic Resource
    Giant magnetroresistance properties of specular spin valve films in a current perpendicular to plane structure (2001)
    [S.l.] : American Institute of Physics (AIP)
    Journal of Applied Physics 89 (2001), S. 6943-6945 
    Details
    ISSN: 1089-7550
    Source: AIP Digital Archive
    Topics: Physics
    Notes: Conventional and specular spin valve films in a current perpendicular to plane (CPP) structure have been investigated. The specular spin valve film with bottom type structure had two oxidized layers: one in the pinned layer, which was oxidized during an in situ deposition process, and the other in the free layer, which was a naturally oxidized Cu/Ta cap. Both films had increasing resistance, R, and resistance change, ΔR, with decreasing element size. The conventional spin valve film showed a resistance times area product, RA, of 144 mΩ μm2 and a resistance change area product, ΔRA, of 0.7 mΩ μm2 while the specular spin valve film showed RA of 1120 mΩ μm2 and ΔRA of 23 mΩ μm2. The ΔRA of the specular spin valve film was about 33 times larger than that of the conventional spin valve film. The calculated magnetoresistance (MR) ratios, MRSV, of each spin valve film were 1.9% and 2.3%, respectively. We think oxidized layers in the spin valve film caused the specular electron scattering and this lengthened the path of the conduction electrons, enhancing the interfacial and bulk spin dependent scattering. We estimated the output voltage change of the 0.01 μm2 element, the size required for 150 Gb/in.2 recording density, of the specular spin valve film in CPP mode to be 5.3 mV. It was almost six times larger than that of the conventional spin valve film at constant power consumption. Specular spin valve film are advantageous for the CPP structure element for future giant MR sensors. © 2001 American Institute of Physics.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1063/1.1364636
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  • 7
    Electronic Resource
    Electronic Resource
    The temperature dependence of exchange anisotropy in ferromagnetic/PdPtMn bilayers (2000)
    [S.l.] : American Institute of Physics (AIP)
    Journal of Applied Physics 87 (2000), S. 6433-6435 
    Details
    ISSN: 1089-7550
    Source: AIP Digital Archive
    Topics: Physics
    Notes: We have studied the differences in the temperature dependence of exchange anisotropy of PdPtMn/CoFeB, NiFe, and NiFeTa bilayers to understand the role of ordering temperatures. The three different ferromagnetic layers [Curie temperatures: TCNiFeTa(400 °C)〈TCNiFe(570 °C)〈TCCoFeB(1040 °C)] exchange biased by the same antiferromagnet [Néel temperature: TNPdPtMn(600 °C)] showed significantly different behavior: the exchange bias field was monotonically decreasing with temperature for the CoFeB and showed distinct, broad peak for NiFe and NiFeTa at a lower temperature before it decreased above 200 °C. The temperature dependence of exchange anisotropy, calculated from the measured exchange bias field and saturation magnetization, was also different for the three bilayers as a result of the differences in Hua and MFM. The results could be understood by the modified thermal fluctuation aftereffect model, which includes the temperature dependence of the ferromagnetic magnetization and the assumption that the ordering temperature of the ferromagnetic film is different at the interface with antiferromagnet from the rest of the ferromagnetic film due to the exchange coupling with a material with high Néel temperature. © 2000 American Institute of Physics.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1063/1.372729
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  • 8
    Electronic Resource
    Electronic Resource
    Lack of small intestinal ulcerogenecity of nitric oxide-releasing indomethacin, NCX-530, in rats (2001)
    Oxford UK : Blackwell Science Ltd
    Alimentary pharmacology & therapeutics 15 (2001), S. 0 
    Details
    ISSN: 1365-2036
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: To evaluate the intestinal ulcerogenic property of nitric oxide-releasing indomethacin (NCX-530) in the rat, in comparison with indomethacin.〈section xml:id="abs1-2"〉〈title type="main"〉Methods:Animals were given indomethacin or NCX-530 subcutaneously and killed 24 h later for macroscopic examination of the small intestine.〈section xml:id="abs1-3"〉〈title type="main"〉Results:A single administration of indomethacin (10 mg/kg) provoked damage, mainly in the jejunum and ileum, accompanied by an increase in myeloperoxidase and inducible nitric oxide synthase activities as well as bacterial translocation. NCX-530 at an equimolar dose (14.2 mg/kg) caused no gross damage in the small intestine, nor any significant change in inducible nitric oxide synthase and myeloperoxidase activities or bacterial translocation. NOR-3, the nitric oxide donor (6.0 mg/kg), when administered subcutaneously together with indomethacin, significantly prevented the occurrence of intestinal lesions and other mucosal changes. Indomethacin reduced mucus and fluid secretions in the small intestine, while both NCX-530 and NOR-3 enhanced these secretions. NCX-530 reduced the mucosal prostaglandin E2 contents and exhibited an anti-inflammatory action against carrageenan-induced paw oedema, with equal effectiveness to indomethacin.〈section xml:id="abs1-4"〉〈title type="main"〉Conclusion:NCX-530 does not cause intestinal damage, despite inhibiting cyclooxygenase activity. The reduced intestinal toxicity of NCX-530 may be attributable to inhibition of enterobacterial translocation, partly by increasing the mucus and fluid secretions mediated by nitric oxide released from this compound.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2036.2001.00916.x
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  • 9
    Electronic Resource
    Electronic Resource
    Up-regulation of COX-2 by inhibition of COX-1 in the rat: a key to NSAID-induced gastric injury (2002)
    Oxford UK : Blackwell Science Ltd.
    Alimentary pharmacology & therapeutics 16 (2002), S. 0 
    Details
    ISSN: 1365-2036
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: A recent study demonstrated that inhibition of both cyclooxygenase (COX)-1 and COX-2 is required for the development of nonsteroidal anti-inflammatory drug (NSAID)-induced gastric lesions. However, the role of COX-1 or COX-2 inhibition in the pathogenisis of these lesions remains unclear.〈section xml:id="abs1-2"〉〈title type="main"〉Aim:To examine the gastric ulcerogenic properties of selective COX-1 and COX-2 inhibitors in rats and to investigate further the relationship of COX inhibition to various events involved in the process of NSAID-induced gastric lesions.〈section xml:id="abs1-3"〉〈title type="main"〉Methods:Animals were given various COX inhibitors p.o., either alone or in combination, and killed 8 h later. Under the treatment, gastric damage, prostaglandin (PG) E2 content, mucosal permeability, myeloperoxidase (MPO) activity as well as gastric motility were examined.〈section xml:id="abs1-4"〉〈title type="main"〉Results:The nonselective COX inhibitor indomethacin inhibited PGE2 production, enhanced gastric motility, and provoked severe lesions in the stomach, with an increase in mucosal permeability and MPO activity. In contrast, the selective COX-2 inhibitor rofecoxib did not induce any damage in the stomach and had no effect on mucosal PGE2 content. Similarly, the selective COX-1 inhibitor SC-560 also caused no gastric damage, despite inhibiting PGE2 production. The combined administration of SC-560 and rofecoxib, however, provoked gross damage in the gastric mucosa, in a dose-dependent manner for each drug. SC-560, but not rofecoxib, caused marked gastric hypermotility and an increase in mucosal permeability, although an increase in MPO activity was observed only when rofecoxib was coadministered. The normal gastric mucosa expressed COX-1 mRNA and not COX-2 mRNA, but COX-2 mRNA was expressed in the stomach after administration of SC-560 as well as indomethacin but not rofecoxib.〈section xml:id="abs1-5"〉〈title type="main"〉Conclusion:These results suggest that the gastric ulcerogenic properties of NSAIDs are not accounted for solely by COX-1 inhibition, but require the inhibition of both COX-1 and COX-2. The inhibition of COX-1 up- regulates COX-2 expression, and COX-2/PGs may, in turn, counteract the deleterious affects of gastric hypermotility due to COX-1 inhibition.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2036.16.s2.22.x
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  • 10
    Electronic Resource
    Electronic Resource
    The roles of nitric oxide and prostaglandins in alterations of ulcerogenic and healing responses in adjuvant-induced arthritic rat stomachs (2000)
    Oxford, UK : Blackwell Science Ltd
    Alimentary pharmacology & therapeutics 14 (2000), S. 0 
    Details
    ISSN: 1365-2036
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Medicine
    Notes: Aim: To examine alterations of gastric ulcerogenic and healing responses in adjuvant-induced arthritic rats. Methods: Arthritis was induced in male Dark Agouti rats by injection of Freund's complete adjuvant into the right hind paw. Results: The gastric ulcerogenic response to indomethacin was markedly increased in AA rats, depending on the degree of arthritic change. By contrast, HCl/ethanol-induced gastric lesions were significantly suppressed in arthritic rats when compared with normal rats. The increased ulcerogenic response to indomethacin was significantly prevented by L-NAME and antineutrophil serum but not by FR167653, while the reduced ulcerogenic response to HCl/ethanol was significantly prevented by L-NAME and partly by indomethacin or NS-398. On the other hand, the healing of chronic gastric ulcers induced by thermal cauterization was also significantly delayed in arthritic rats when compared with normal rats. This delayed healing of gastric ulcers was affected by neither L-NAME, indomethacin nor FR167653. The gastric mucosa of arthritic rats showed a significant increase in both inducible nitric oxide synthase (iNOS) activity and prostaglandin (PG) E2 contents. Conclusions: Gastric ulcerogenic and healing responses were altered in arthritic rats. The ulcerogenic response to indomethacin was increased while that to HCl/ethanol was decreased. These changes in ulcerogenic responses may both be accounted for by increased production of NO/iNOS, with the latter also being partially related to elevated production of PGs/COX-2. Moreover, the healing of gastric ulcers was also delayed in arthritic rats, but the mechanism was related to neither NO nor PGs.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2036.2000.014s1018.x
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