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Biomedical subjects

T Osawa

Publications and source records attributed to T Osawa.

At least 397 records · Page 22Linked to original sources
PubMed1981

Identification of a serum glycoprotein that increased in OK-432-treated mice as hemopexin.

LB, the antitumor protein whose level is increased in the serum of OK-432-treated mice, was identified as hemopexin, which is a serum beta-glycoprotein having a heme-binding capability. When incubated with heme, LB showed several absorption maxima in the ultraviolet and visible regions, and the addition of dithionite increased the extinction of the Soret band, which was shifted to a longer wavelength. The ultraviolet circular dichroism spectrum of LB exhibits a positive maximum at 292 nm and a shoulder near 280 nm. The far ultraviolet CD spectrum of LB has a prominent positive maximum at 231 nm. These spectral characteristics are identical with those of rabbit, rat or human hemopexin. In addition, the relative mobility on polyacrylamide gradient gel electrophoresis and the results of amino acid analysis ae in very good agreement with results obtained previously for rabbit and human hemopexins. Since LB has enhancing effects on the tumor cell killing and binding by OK-432-elicited macrophages, these results strongly imply that the increase of hemopexin in serum plays an important role in the host-mediated antitumor activity of certain antitumor drugs.

Amino Acids↗
PubMed1981

Antitumor and toxohormone-neutralizing activities of human ceruloplasmin.

Human ceruloplasmin was found to have a neutralizing effect against the toxohormone activities of the basic protein isolated from Ehrlich carcinoma cells; this basic protein decreases the levels of serum iron and liver catalase activity upon intraperitoneal injection into mice and shows direct cytotoxicity toward normal mouse lymphocytes and macrophages. Furthermore, it was also shown that ceruloplasmin has antitumor activity against sarcoma-180 cells implanted in ICR mice.

Animals↗
PubMed1980

Studies on adenosine 3',5'-monophosphate phosphodiesterase of human erythrocyte membranes.

In this work we show the existence of cyclic AMP phosphodiesterase (EC 3.1.4.17) in human erythrocyte membranes and have clarified some properties of the enzyme. In human erythrocytes, about 23% of the total cyclic AMP phosphodiesterase activity is in a membrane-bound form. Although it could be solubilized with Triton X-100 in 5 mM Tris-HCl buffer (pH 8.0), it was not solubilized by a low or high concentration of salt. The enzyme seems to be localized in the cytoplasmic surface, since it is detected in sealed inside-out vesicles of human erythrocyte membranes, but not in intact human erythrocytes. The optimum pH was found to lie between 7.4 and 8.0, and Mg2+ was found to be necessary for its activity. Ca2+ and calmodulin could not stimulate the activity of this enzyme. Theophylline was a strong inhibitor, but cyclic GMP could not inhibit the enzymic hydrolysis of cyclic [32P]AMP and this membrane-bound enzyme therefore seems to be specific to cyclic AMP.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1980

The carbohydrate moiety of band-3 glycoprotein of human erythrocyte membranes.

Band-3 glycoprotein was purified from human blood-group-A erythrocyte membranes by selective solubilization and gel chromatography on Sepharose 6B in the presence of sodium dodecyl sulphate. The purified glycoprotein was subjected to hydrazinolysis in order to release the carbohydrate moiety. The released oligosaccharides were N-acetylated and applied to a column of DEAE-cellulose. Most of the band-3 oligosaccharides obtained were found to be free of sialic acids. When this neutral fraction was subjected to gel chromatography on a column of Sephadex G-50, two broad peaks were observed indicating that the band-3 glycoprotein was heterogeneous in the size of the oligosaccharide moieties. All fractions from gel chromatography were found to contain galactose, mannose, N-acetylglucosamine and fucose. The higher-molecular-weight (mol.wt. 3000-8000) peak consisted of fucose, mannose, galactose, N-acetylglucosamine and N-acetylgalactosamine in a molar proportion of 1.6:3.0:8.4:10.5:0.2. Most of these oligosaccharides were digested with a mixture of beta-galactosidase and beta-N-acetylhexosaminidase after alpha-L-fucosidase treatment to give a small oligosaccharide with the structure alpha Man2-beta Man-beta GlcNAc-GlcNAc. Methylation studies and limited degradation by nitrous acid deamination showed that the oligosaccharides contained the repeating disaccharide Gal beta 1----4GlcNAc beta 1----3, with branching points at C-6 of some of the galactose residues. These results indicate that a major portion of the band-3 oligosaccharide has a common core structure, with heterogeneity in the numbers of the repeating disaccharides, and contains fucose residues both in the peripheral portion and in the core portion. Haemagglutination tests were also carried out to determine the blood-group specificities of the glycoprotein and the results demonstrated the presence of both blood-group-H and I antigenic activities.

ABO Blood-Group System↗
PubMed1980

Comparative lectin-binding and agglutination properties of the strain-specific, TA-3-St, and the non-strain-specific, TA3-Ha, murine mammary carcinoma ascites sublines. Further studies of receptors in epiglycanin.

The complex carbohydrates at the cell surfaces of two TA3, murine mammary carcinoma ascites sublines (the strain-specific, TA3-St subline and the nonstrain-specific. TA3-Ha line) were compared by binding studies with 125I-labelled concanavalin A (con A), Ricinis communis agglutinin (RCA), and eel-serum agglutinin (ESA). The TA3-Ha cell bound equal amounts of con A, 1.5-fold more RCA, and 4-fold more ESA than the TA3-St cell. Binding-inhibition studies by these lectins and two others [wheat-germ agglutinin (WGA) and potato lectin (STA)] suggest complementary binding-sites between con A and both RCA and ESA. Quantitative agglutination studies with the five lectins, and inhibition determinations by both neuraminidase-treated and untreated epiglycanin revealed that TA3-St, but not TA3-Ha, cells were agglutinated by con A, and that epiglycanin inhibited this agglutination, as well as the agglutination of rabbit erythrocytes by con A. The presence of a con A receptor on epiglycanin was also suggested by the binding of epiglycanin to con A-Sepharose, and its specific elution with methyl alpha-D-mannopyranoside. TA3-St cells were agglutinated at a 10-15-fold lower concentration of either STA or RCA than TA3-Ha cells, but both cells were agglutinated by the same concentration of WGA and ESA. Inhibition by epiglycanin of agglutination of TA3-St cells by either STA or ESA occurred at a concentration lower than that of TA3-Ha cells, but epiglycanin inhibited RCA agglutination of TA3-Hs cells at a concentration lower than that of TA3-St cells. Epiglycanin, but not asialoepiglycanin, inhibited the agglutination of TA3-Ha cells by WGA.

Agglutination↗
PubMed1980

The major sialoglycoprotein of human T-lymphocytes.

Human, peripheral-blood T-lymphocytes and human, T-lymphoblastoid cells of a MOLT 4B cell-line were surface-labeled by lactoperoxidase-catalyzed iodination, periodate and sodium borotritide, and galactose oxidase and sodium borotritide, and analyzed by dodecyl sodium sulfate-polyacrylamide gel-electrophoresis. Both types of cells were found to show a major, cell-surface sialoglycoprotein with an apparent mol. wt. of 95,000. After neuraminidase treatment, this glycoprotein showed a higher mol. wt. of 120,000. The major sialoglycoprotein of both types of cells bound to wheat-germ agglutinin and concanavalin A and, after neuraminidase treatment, to Arachis hypogaea agglutinin. The glycopeptides obtained from these glycoproteins by Pronase digestion gave similar elution-profiles on Sephadex G-50 gel filtration. These results suggest that the major sialoglycoprotein of normal T cells and that of MOLT 4B cells are very similar, if not identical.

Cell Line↗
PubMed1980

Amber mutations in the structural gene for RNA polymerase sigma factor of Escherichia coli.

Amber mutants of Escherichia coli K-12 affected in the structural gene (rpoD) for the sigma subunit of RNA polymerase have been obtained from a strain harboring a temperature-sensitive amber suppressor (supF-Ts6) which is active only at low temperatures. These mutants grow normally at low temperature (30 degrees C) but do not grow at high temperature (42 degrees C) due to the inability to synthesize sigma factor. In one mutant studied in detail (rpoD40), the rate of sigma-factor synthesis at 30 degrees C is about half that of the wild type and is decreased to 10%-15% within 1 h of incubation at 42 degrees C. The synthesis of core polymerase subunits or bulk protein is virtually unaffected at least for 2 h. The defect of the mutant in sigma synthesis and growth at high temperature can be suppressed by any of the amber suppressors tested (supD. supE or supF). RNA-polymerase holoenzymes prepared from the mutant cells carrying each of the suppressors (grown at 42 degrees C) exhibit different thermostabilities attributable to alterations in the sigma factor. The reduced sigma synthesis in the mutant is accompanied by the synthesis of polypeptide tentatively identified as 'amber fragment'. These results as well as the genetic mapping data indicate that the amber mutation (rpoD40) resides within the structural gene for the sigma factor and directly affects sigma synthesis upon inactivation of the suppressor at high temperature.

Chromosome Mapping↗
PubMed1980

Formation of mutagens by sorbic acid-nitrite reaction: effects of reaction conditions on biological activities.

Conditions of the reaction between sorbic acid and sodium nitrite generating mutagenic principles were examined. In the rec-assay and the Ames reversion assay, the maximal mutagenic activity was obtained in a pH range of 3.5-4.2. Mutagenic and growth-inhibitor activities of five C-nitro and C-nitroso compounds were studied. The product Y, 2-methyl-1,4-dinitropyrrole, was the strongest mutagen among them.

Drug Interactions↗
PubMed1980

Amination and subsequent derivatization of epoxy-activated agarose for the preparation of new affinity adsorbents.

We have found a simple procedure to convert epoxy-activated agarose into amino derivatives using ammonia solution. The amino derivatives of agarose were succinylated with succinic anhydride and then activated with N-hydroxysuccinimide according to the method of Cuatrecasas and Parikh. Formula: (See Text). Coupling of the ligand to the activated agarose (shown above) was performed under mild conditions. The adsorbent thus obtained has no charged group in the linkage region between the ligand and agarose, thus reducing the nonspecific adsorption, and the bonds formed (ether and amide bonds) are stable even in an alkaline medium. Lens culinaris hemagglutinin-Sepharose 4B prepared by this method was successfully used for the affinity chromatography of solubilized human red blood cell membrane in detergent solution and was stable when elution was performed with borate buffer, pH 9.8.

Amination↗
PubMed1980

Heterogeneity of Band 3, the major intrinsic protein of human erythrocyte membranes. Studies by crossed immunoelectrophoresis and crossed immuno-affinoelectrophoresis.

The binding of the human erythrocyte membrane major intrinsic protein (Band 3) to concanavalin A (Con A), Ricinus communis agglutinin (RCA), and pokeweed mitogens (PWM) has been studied by crossed immunoelectrophoresis and crossed immuno-affinoelectrophoresis. Although pure by the criterion of polyacrylamide gel electrophoresis, Band 3 was resolved by crossed immunoelectrophoresis into three peaks. A major peak was split on both the anodic and cathodic sides, and consisted of at least two components. A smaller additional peak was also observed. By introducing Con A in the first dimension of the run, the main peak was dissociated into 2 peaks of different heights. Partial binding to Con A was confirmed by introducing Con A Sepharose 4B in an intermediate gel in the second dimension of the electrophoresis. When the first dimension gel of the crossed immunoelectrophoresis contained RCA, or when RCA Sepharose 4B was present in an intermediate gel in the second dimension of the run, the major peak was far smaller because of binding to that lectin. In experiments with a frist dimension gel containing pokeweed mitogens, the major peak was higher but narrower than in the control experiments, presumably due to selective binding of some constitutents of Band 3. Immuno-affinoelectrophoretic methods therefore show that Band 3 is heterogeneous and that its components bind differently to the above lectins.

Anion Exchange Protein 1, Erythrocyte↗
PubMed1980

Alpha-casein-binding proteins of guinea pig macrophage membranes and their possible roles in chemotaxis.

The existence of specific binding proteins for alpha-casein, a well-known potent chemoattractant, on guinea pig peritoneal macrophages was demonstrated. Binding of 3H-alpha-casein to macrophages was found to be specific and reversible. Its association constant and the number of binding sites were 5.0 X 10(-6) M and 7.5 X 10(5) per cell, respectively. The presence of formyl-methionyl-leucyl-phenylalanine (fMet-Leu-Phe), another chemoattractant, at a chemotactically active concentration (10(-11) to 10(-6) M) reduced this binding significantly. Pretreatment of macrophages with proteases such as trypsin and chymotrypsin decreased the number of binding sites for alpha-casein and the chemotactic response to alpha-casein. alpha-Casein binding proteins were isolated from radiolabeled macrophage surface membranes by affinity chromatography on a column of alpha-casein Sepharose 4B. SDS-polyacrylamide gel electrophoresis of the isolated proteins revealed three radioactive bands; two (MW 130,000 and 65,000) were glycoproteins, and the other (MW 100,000) was a protein which contained little carbohydrate. These three proteins were also isolated from the same source by affinity chromatography using a column of fMet-Leu-Phe-AH-Sepharose. This result, together with the observation that fMet-Leu-Phe inhibited the binding of alpha-casein to macrophages, indicated that alpha-casein and the tripeptide have at least partly common binding sites on guinea pig macrophages. Some of the proteins obtained in this work may constitute chemotactic receptors on the macrophage membrane.

Animals↗
PubMed1980

Effect of puromycin on guinea pig lymphotoxin release and lectin-induced cellular cytotoxicity.

In order to confirm the role of guinea pig lymphotoxin (GLT) in lectin-induced cellular cytotoxicity (LICC), the effect of puromycin, a potent enhancer of GLT activity, on the LICC to target L . P3 cells induced by phytohemagglutinin (PHA) was investigated under serum-free conditions. LICC was completely inhibited by puromycin, when it was added at the initiation of LICC culture, because of the inhibition of the release of GLT from the effector lymph node cells. However, LICC was markedly enhanced when puromycin was added several hours after the initiation of LICC culture. The interpretation of these facts is that GLT release can be inhibited by puromycin, but that the GLT already released exerts an enhanced cytotoxic effect on the target cells in the presence of puromycin. Enhancement of the cytotoxicity by the addition of puromycin several hours after the initiation of LICC culture was observed even after the removal of the GLT present in the supernatant, suggesting that the morphologically intact target cells were already affected by GLT in the early stages of LICC culture.

Animals↗
PubMed1980

Ia requirement in T cell response to various mitogenic lectins.

Various lectins, mainly T cell mitogens, were examined to determine the relationship between anti-Ia antiserum-treated T cell responses and the sugar specificities of these lectins. Anti-Ia antiserum and complement treatment diminished nylon wook column-purified T cell responses to most lectins such as concanavalin A, Lens culinaris hemagglutinin, Pisum sativum agglutinin, Witaria floribunda mitogen, Limulus polyphemus agglutinin and pokeweed mitogen, and the diminished response to these lectins could be restored by adding nylon wool-adherent cells, whereas the same treatment rather enhanced their response to Phaseolus vulgaris phytohemagglutinin. We concluded that whether t cell responses to various lectins require Ia-bearing cells more or less is independent of the sugar-binding specificities of these lectins.

Animals↗