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N Seno

Publications and source records attributed to N Seno.

At least 55 records · Page 3Linked to original sources
PubMed1987

Separation of isolectins by high-performance hydrophobic interaction chromatography.

High-performance hydrophobic interaction chromatography (HP-HIC) was found to be an effective method for the separation of lectins into isolectin fractions. All of the purified lectins used in this study, Phaseolus vulgaris haemagglutinin (PHA), wheat germ agglutinin (WGA), Ricinus communis agglutinin (RCA), and Arachis hypogaea agglutinin (AHA), were prepared by affinity chromatography. HP-HIC was performed on a column (15 X 2.1 cm) of TSK gel Phenyl-5PW at room temperature. The lectin sample, dissolved in 1.0 or 0.5 M ammonium sulphate in phosphate buffered saline (pH 7.4) (PBS), was applied to the column and eluted with a linear gradient from 1.0 or 0.5 M ammonium sulphate in PBS to 0 M ammonium sulphate in PBS at a flow-rate of 4 ml/min. In the case of RCA, addition of glycerol to the elution buffer resulted in sharper isolectin peaks. PHA, WGA, RCA, and AHA were rapidly separated into 5, 5, 4, and 6 isolectins, respectively.

Arachis↗
PubMed1987

Improved method for the immobilization of heparin.

The optimal conditions for immobilizing heparin through its terminal formyl group were investigated. When Amino Sepharose (1 g) was suspended in 1 ml of phosphate buffer (pH 7) containing 30 mg of heparin and 3 mg of sodium cyanoborohydride, with shaking at room temperature, the maximum immobilization of heparin (10 mg of heparin per gram of wet gel) was reached within 2 days. The Heparin Sepharose thus obtained was stable: no significant loss of the heparin content was observed after storage for 4 months at 4 degrees C. Heparin was also immobilized by the same method with Amino TSK gel G5000PW instead of Amino Sepharose 4B and was successfully applied to the high-performance liquid affinity chromatography of fibronectin and thrombin.

Chromatography, High Pressure Liquid↗
PubMed1986

Characterization of the carbohydrate moiety of Clerodendron trichotomum lectins. Its structure and reactivity toward plant lectins.

Lectins were isolated from fruits and leaves of Clerodendron trichotomum by affinity chromatography on lactamyl-Sepharose. The purified lectins (C. trichotomum agglutinin: CTA) were homogeneous on SDS/polyacrylamide gel electrophoresis, and the carbohydrate moiety was characterized by physicochemical and immunochemical methods. The asparagine-linked oligosaccharides were released by treatment with N-oligosaccharide glycopeptidase (almond, EC 3.5.1.52) of peptic glycopeptides obtained from fruit CTA, and separated by gel filtration and thin-layer chromatography. The structure of the predominant oligosaccharide was determined as Xyl beta 1----2 (Man alpha 1----6)(Man alpha 1----3)Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----3)GlcNAc by high-performance liquid chromatography, sugar analysis and 1H-NMR spectroscopy. The reactivity of the carbohydrate moiety of CTA toward various lectins was studied. Fruit and leaf CTAs were applied to polyacrylamide gel electrophoresis, transferred to nitrocellulose sheets and detected with horseradish-peroxidase-conjugated lectins. Concanavalin A, lentil lectin, pea lectin, Vicia faba lectin and Ulex europeus agglutinin I, but not wheat germ lectin, bound to fruit CTA. The results indicate new binding properties of these plant lectins: a beta-xylosyl residue substituted at C-2 of the beta-mannosyl residue of N-linked oligosaccharide does not affect the binding with mannose-specific lectins, lentil, pea and Vicia faba lectins can bind to N-linked oligosaccharides containing an alpha-L-fucosyl residue attached to C-3 of the asparagine-linked N-acetyl-D-glucosamine residue, and Ulex europeus agglutinin I can bind to the (alpha 1----3)-linked fucose residue of the N-linked oligosaccharide.

Carbohydrates↗
PubMed1986

New role for heparan sulfate: regulator of leukotriene generation in mouse E-mast cells.

In this work, bovine heparan sulfate, pig mucosa heparin and squid chondroitin sulfate-E glycosaminoglycans (GAGs) were compared as to their affect on the synthesis of leukotrienes C4 (LTC4) and B4 (LTB4) as well as on the production of prostaglandin D2 (PGD2) in cultured mouse E-mast cells (E-MC). The maximum percent increase in LTC4 generation in cells treated with 0.2 micrograms heparan sulfate was 52 +/- 3% (mean +/- S.E., n = 5). Whereas 0.5 micrograms of the GAG increased the production of LTB4 by 50% Ten micrograms of heparin slightly increased the LTC4 production (33%) whereas lower doses were found to be ineffective. No significant increase in LTC4 production was demonstrated when the IgE sensitized E-MC were treated with chondroitin sulfate-E GAG prior to the antigen challenge. Neither one of the three GAGs, at the various doses used, affected the antigen induced exocytosis of beta-hexosaminidase from the IgE sensitized E-MC. After 15 min preincubation with heparan sulfate, antigen induced release of PGD2 from 1 X 10(6) sensitized cells was inhibited from 5.4 ng +/- 0.1 ng into 3.5 ng +/- 0.1 ng at 0.5 micrograms/ml GAG. All of the three types of GAGs used were uneffective when the E-MC were activated by calcium ionophore A23187.

Animals↗
PubMed1986

Mammalian kidney lectin.

Lectins that agglutinate intact red blood cells of rabbit, and their endogenous inhibitors, were found in extracts of cow, pig, rat, and human kidneys. Lectins were separated from inhibitors by gel chromatography on a column of Toyopearl HW-75. Adsorption tests with immobilized glycoconjugates and ion-exchange gels revealed that the bovine kidney lectin binds with sialylglycoproteins by sugar-specific interactions but not by nonspecific, electrostatic interactions. The results of the hemagglutination-inhibition tests showed that all of the endogenous inhibitor fractions were active toward all renal lectins tested, and that glycoproteins having sialyl residues, and also heparin, are also good inhibitors of the lectins. However, the hemagglutination-inhibitory activity of human urinary Tamm-Horsfall glycoproteins varied from source to source, and that having the highest inhibitory activity did not always have the highest content of sialic acids. Crude bovine kidney lectin was further purified by ion-exchange chromatography on DEAE-Sephadex A-50. Analysis by SDS-polyacrylamide gel electrophoresis showed that the purified lectin consists of a approximately 63,000 daltons subunit.

Agglutination↗
PubMed1986

Carbohydrate-binding specificity of silkworm lectin.

The binding specificity of a lectin from the hemolymph of silkworm larvae was examined quantitatively by taking advantage of the fluorospectrophotometric properties of the lectin. On excitation at 280 nm, the lectin fraction gave a fluorescence-emission spectrum centered at 336 nm, which was attributable to tryptophan residues. The fluorescence could be completely quenched by the addition of specific saccharides. The affinity constants of the silkworm lectin with specific saccharides were calculated from the changes in intensities of fluorescence-difference spectra induced by the saccharides. The silkworm lectin had the highest affinity for dermatan sulfate and hyaluronic acid, followed by protuberic acid, heparin, and chondroitin sulfate A. Among monosaccharides tested, only D-glucuronic acid and N-acetyl-neuraminic acid induced weak but significant quenching, and their affinity constants were found to be low. These results indicate that the silkworm lectin has a strong affinity for carboxyl groups, especially alpha-L-iduronic acid residues, in the saccharides. In most cases, sulfate groups on the saccharides interfere with the specific interactions.

Animals↗
PubMed1986

The anticoagulant properties of mast cell product, chondroitin sulphate E.

The anticoagulant potency in vitro of chondroitin sulphate E has been found to be similar to that of the heparinoids. In purified systems chondroitin sulphate E was shown to be principally an activator of heparin cofactor II. Maximum acceleration of heparin cofactor II:thrombin interaction was 185-fold (9.3 X 10(7) M-1 min-1), antithrombin III:thrombin interaction was 11-fold (4.16 X 10(6) M-1 min-1) and antithrombin III:factor Xa was 146-fold (3.86 X 10(6) M-1 min-1). Chondroitin sulphate E was observed to prolong the thrombin clotting time of fibrinogen in the absence of antithrombin III and heparin cofactor II. The effect appeared to be related to interference in thrombin:fibrinogen interaction rather than in fibrin monomer polymerization.

Anticoagulants↗
PubMed1986

Immunological characterization of human vitronectin and its binding to glycosaminoglycans.

The cell-adhesive glycoprotein vitronectin in human plasma was characterized with a monospecific anti-vitronectin antibody. Vitronectin, a mixture of monomeric 75 and 65 kDa polypeptides, was found to have different ratios of amounts of 75 and 65 kDa polypeptides in immunoblots of sera from various healthy human donors. Two states of vitronectin were previously reported; the open state binds to heparin, but the cryptic state does not (Hayashi et al. (1985) J. Biochem. 98, 1135-1138). The anti-vitronectin antibody was suggested to react more strongly with the open state of vitronectin than with the cryptic state. To quantitate all vitronectin regardless of its state, an enzyme-linked immunosorbent assay of vitronectin was developed based on prior boiling of vitronectin-containing samples in 2% (w/v) sodium dodecyl sulfate and 40 mM dithiothreitol to destroy conformational differences. About 12-20% of the vitronectin molecules in plasma were found to bind to heparin-Sepharose under physiological conditions. Vitronectin in plasma bound 30-fold more efficiently to heparin immobilized by amino groups than by carboxyl groups. Its affinity for heparin was higher than for chondroitin sulfate A or C, or dermatan sulfate. Vitronectin was also found to contain covalently-linked small polypeptides of 15 and 13 kDa. These light chains seemed to be disulfide-bonded to the 65 kDa polypeptide, and might be endogenously derived from nicks in the carboxy-terminal portion of the 75 kDa polypeptide in plasma.

Chromatography↗
PubMed1986

Lectin binding sites related with rat ascites hepatoma cell adhesion.

In order to elucidate the correlation between cell surface lectin binding sites and the degree of cell adhesiveness, quantitative lectin binding assays were performed using three types of rat ascites hepatoma cell lines (free cell, mixed cell, and island-forming cell types). The lectin binding site patterns showed no remarkable differences among the intact tumor cell lines, but treatment of the cells with L-1-tosylamide-2-phenylethyl chloromethyl ketone (TPCK)-trypsin or neuraminidase induced remarkable differences in the modulation of the number of lectin binding sites. TPCK-trypsin treatment caused a marked decrease in the number of peanut agglutinin binding sites on the island-forming and mixed cell types, concomitant with disaggregation of the cells, showing that trypsin sensitive binding sites are involved in the cell-cell adhesion. Neuraminidase treatment caused a decrease in wheat germ agglutinin binding sites and an increase in castor bean agglutinin binding sites, and these effects were greater for the free cell type. These results indicated that alpha-sialyl-beta-D-galactosyl residues are more abundant on the cell surface of the free cell type than the other cell types. Therefore, it was suggested that electrostatic repulsion due to negative charges of the cell surface sialic acid contributes to the low cell adhesiveness of the free cell type.

Animals↗
PubMed1986

Preparation of high capacity affinity adsorbents using new hydrazino-carriers and their use for low and high performance affinity chromatography of lectins.

Two kinds of carriers with high concentrations of hydrazino groups were prepared by simple and convenient procedures. Hydrazino-carriers (I) and (II) were obtained on incubation of epoxy-activated carriers with hydrazine hydrate and adipic acid dihydrazide, respectively. Disaccharides were coupled to the hydrazino carriers through reductive amination in the presence of sodium cyanoborohydride. The reaction time was much shorter (24 h) than that in the case of the method involving amino-Sepharose 6B (800 h) [Matsumoto, I., Kitagaki, H., Akai, Y., Ito, Y., & Seno, N. (1981) Anal. Biochem. 116, 103-110]. The glycamyl-Sepharose thus obtained showed high adsorption capacities for lectins. Glycamyl-TSKgel G3000 PW obtained by the same method with TSKgel G3000 PW, which is a hydrophobic vinyl polymer matrix for high performance gel permeation liquid chromatography, could be successfully used for the high performance liquid affinity chromatography of lectins. N-Acetylglutamic acid was coupled to hydrazino-Sepharose 4B (I) in the presence of N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline. The adsorbent obtained was used for the affinity chromatography of Japanese horseshoe crab lectin.

Animals↗
PubMed1985

The development of the dorsal root potential and the responsiveness of primary afferent fibers to gamma-aminobutyric acid in the spinal cord of rat fetuses.

The development of the dorsal root potential (DRP) and the responsiveness of primary afferent fibers to gamma-aminobutyric acid (GABA) were investigated in the isolated spinal cord of rat fetuses. At embryonic day 15.5, stimulation of the lumbar dorsal root was first effective in eliciting the DRP, which was not inhibited by bicuculline. A bicuculline-sensitive component of the DRP appeared at embryonic day 17.5. GABA (10 microM to 1 mM) caused a dose-dependent depolarization of the primary afferent fibers from embryonic day 13.5. The amplitude of the depolarization gradually increased with age until embryonic day 17.5 and was maintained thereafter. If the bicuculline-sensitive DRP solely reflects GABAergic activity, it is suggested that GABAergic activity develops at embryonic day 17.5 and the development of the responsiveness of primary afferent fibers to GABA precedes the functional onset of GABAergic neurons.

Animals↗
PubMed1985

Isolation and characterization of a lectin from the fruit of Clerodendron trichotomum.

An agglutinin of Clerodendron trichotomum fruit (CTA), found to be specific for N-acetyl-D-galactosamine and D-galactose, was isolated and characterized. The fruit extract was decolorized first by passage through a Toyopearl column and a phenyl-Sepharose column. Then the lectin activity was adsorbed on p-aminophenyl N-acetyl-alpha-D-galactosaminide- or p-aminophenyl beta-D-galactoside-Sepharose, and eluted as a sharp peak with 0.2 M lactose. The purified CTA was found to be homogeneous by SDS-polyacrylamide gel electrophoresis, gel chromatography and ultracentrifugal analysis, and was determined to be a glycoprotein homodimer with a molecular weight of 56,000 daltons. Hemagglutination-inhibition assay indicated that CTA is most specific for N-acetyl-D-galactosaminide with a hydrophobic aglycon.

Adsorption↗
PubMed1985

Immobilization of protein ligands on new formyl-spacer-carriers for the preparation of stable and high capacity affinity adsorbents.

New procedures to immobilize high concentrations of protein ligands by reductive amination on two types of formyl-carriers (I & II) having different spacer lengths were investigated in order to prepare stable and high-capacity adsorbents essential for efficient affinity chromatography. Formyl-carrier (I) was prepared by reductive amination with glutaraldehyde of the amino-carrier obtained on amination of an epoxy-activated carrier. Formyl-carrier (II) was prepared by sodium metaperiodate (NaIO4) treatment of a glyceryl-carrier obtained on hydrolysis of an epoxy-activated carrier. Especially high concentrations of protein ligands were immobilized on formyl-Sepharose 4B (I) under very mild conditions (pH 7.0, 4 degrees C). A series of lectins, one of the most useful classes of group-specific ligands, was successfully immobilized by the procedures. Concanavalin A-Sepharose 4B (I) thus obtained exhibited an adsorption capacity five times greater than that of concanavalin A-Sepharose 4B made by Pharmacia Fine Chemicals, and could be repeatedly used over twenty times without a significant reduction in its adsorption capacity.

Adsorption↗
PubMed1985

Dermatan sulfate-reactive lectin from chicken liver.

A lectin highly reactive with dermatan sulfate (DS-lectin) was purified from adult chicken liver by gel filtration on Toyopearl HW-55 and subsequent affinity chromatography on new adsorbents which were prepared by immobilizing heparin or dermatan sulfate via the reducing ends on hydrazino-Toyopearl. The DS-lectin behaved as a single protein on polyacrylamide gel electrophoresis. On excitation at 280 nm, the DS-lectin emitted fluorescence centered at 336 nm, which was attributable to tryptophan residues and could be quenched by the addition of specific saccharides. The affinity constants of the DS-lectin with specific saccharides were calculated from the changes in intensities of fluorescence-difference spectra induced by the saccharides. Dermatan sulfate and protuberic acid, which is composed of L-iduronic acid and D-glucuronic acid (1:2), had the highest affinity constants among the polysaccharides tested. Partially N-desulfated heparin had a higher affinity constant than that of native heparin while dextran sulfate showed no affinity. D-Glucuronic acid and N-acetylneuraminic acid induced weak but significant quenching, but not N-acetylgalactosamine or cellobiose. These results were essentially in good agreement with those of hemagglutination inhibition tests and indicated that DS-lectin has a strong affinity for L-iduronic acid residues and probably carboxyl groups in the saccharides, while sulfate groups on the saccharides interfere with the specific interaction.

Amino Acids↗
PubMed1985

[Clinical study on the penetration of latamoxef into the pulmonary tissue in surgery of the chest].

In the surgery of the chest, we are often experienced pulmonary infections, so it is considered that the grasping antibiotic levels in pulmonary tissue is very important because of the decision of antibiotic dose schedule against pulmonary infections. For this purpose, latamoxef (LMOX) at a dose of 2 g was intravenously administered to 14 cases with pulmonary cancer, 3 cases with pulmonary tuberculosis, 2 cases with pulmonary abscess and 1 case with bronchiectasis, totally 20 cases and the concentrations in serum, pulmonary tissue, bronchia and sputum were measured up to 6 hours and the results obtained were as follows; The average serum concentration of LMOX was 137.2 micrograms/ml at 1/2 hour and decreased gradually, fell to 20.3 micrograms/ml at 6 hours. The average levels of LMOX in normal alveolus of pulmonary tissue and in bronchia were 63% and 48% of the serum level, respectively. The average level of LMOX in inflammatory alveolus of pulmonary tissue was approximately 20% lower than the normal alveolus. The average level of LMOX in sputum increased gradually and appeared 4.6 micrograms/ml at 6 hours. LMOX was shown a good results on the penetration into the pulmonary tissue and its level almost exceeded the minimal inhibitory concentration against clinical isolates from sputum of respiratory tract infections.

Adult↗
PubMed1984

Analysis of polysulfated chondroitin disaccharides by high-performance liquid chromatography.

A high-performance liquid chromatography method for analyzing disaccharides derived from chondroitin sulfate glycosaminoglycans has been developed which employs a Whatman Partisil-10 PAC amino-cyano column and an acetonitrile/methanol/ammonium acetate solvent to resolve disulfated, monosulfated, and unsulfated disaccharides in a chromatographic run of less than 20 min. The single known trisulfated chrondroitin disaccharide can be eluted in an alternate solvent system containing the same mobile phase components in different proportions. Disaccharides were prepared for chromatography from glycosaminoglycans and proteoglycans of known compositions by digestion with chondroitinase ABC, with the exception of king crab cartilage glycosaminoglycan which was incubated sequentially with hyaluronidase and chondroitinase ABC. Disaccharides were extracted from the digestion mixtures in 80% ethanol, dried over nitrogen, resuspended in the HPLC solvent, and chromatographed at a flow rate of 1 ml/min. Unsaturated disaccharides in the column eluate were detected by continuous ultraviolet absorbance monitoring at 232 nm; alternatively, fractions were collected and assayed for uronic acid content or radioactivity. By utilizing the HPLC technique in conjunction with chondroitinase ABC and AC digestion and sulfatase hydrolysis, the epimeric structures of chondroitin sulfates E and H were confirmed. With this technique, rapid and reproducible analyses of chondroitin sulfate disaccharides generated from mouse mast cell proteoglycan and from glycosaminoglycans of squid cranial cartilage, shark skin, hagfish skin, and hagfish notocord were in close agreement with compositions obtained by other techniques.

Animals↗
PubMed1984

The development of responsiveness to substance P and glutamate in the spinal motoneurons of rat fetuses.

The development of responsiveness of motoneurons to substance P (SP) and glutamate was investigated in the isolated spinal cord of the rat fetus at embryonic days 13.5-21.5. The motoneurons at embryonic day 14.5 first responded to SP with a slow depolarization. The responsiveness to SP increased by embryonic days 19.5-21.5. In a low Ca2+ solution, responsiveness was reduced after embryonic day 17.5. Glutamate caused a slow depolarization of motoneurons from embryonic day 13.5. The responsiveness increased until embryonic day 17.5 and decreased thereafter.

Animals↗