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K Onozaki

Publications and source records attributed to K Onozaki.

At least 55 records · Page 3Linked to original sources
PubMed1995

A human erythrocyte-derived growth-promoting factor with a wide target cell spectrum: identification as catalase.

We have reported previously that a factor with a molecular weight of 53,000 under SDS-polyacrylamide gel electrophoresis purified from human erythrocyte extracts promoted the growth of a wide variety of cell types from different species, including T cells, B cells, myeloid leukemia cells, melanoma cells, and mastocytoma cells, as well as normal and transformed fibroblast cells. In the present study, amino acid sequence analysis revealed that this factor has homology with human catalase. The purified factor exhibited catalase activity. Catalases derived from human erythrocytes, bovine liver, Aspergillus niger, and recombinant rat liver catalase are all able to promote the growth of cells. Antibody against human catalase absorbed both the growth-promoting activity and the enzyme activity of the purified factor. In addition, treatment of the factor with an irreversible enzyme inhibitor, aminotriazole, resulted in abrogation of both the growth-promoting activity and enzyme activity. These results indicate that the growth-promoting factor is catalase, and its activity is associated with the decomposition of hydrogen peroxide.

Amino Acid Sequence↗
PubMed1995

Type I and type II interferons upregulate functional type I interleukin-1 receptor in a human fibroblast cell line TIG-1.

The regulation of type I interleukin-1 receptor (IL-1R) expression by type I, interferon (IFN)-alpha A/D, and type II IFN, IFN-gamma, in a human fibroblast cell line TIG-1 was investigated. After 2 h stimulation with human IFN-alpha A/D or IFN-gamma, the levels of type I IL-1R mRNA increased. We previously reported that IL-1 upregulates transcription and cell surface molecules of type I IL-1R in TIG-1 cells through induction of prostaglandin (PG) E2 and cAMP accumulation. However, indomethacin was unable to inhibit the effect of IFNs, indicating that IFNs augment IL-1R expression through a pathway distinct from that of IL-1. The augmentation was also observed in other fibroblast cell lines. Nuclear run-on assays and studies of the stability of mRNA suggested that the increase in IL-1R mRNA was a result of the enhanced transcription of IL-1R gene. Binding studies using 125I-IL-1 alpha revealed that the number of cell surface IL-1R increased with no change in binding affinity by treatment with these IFNs. Pretreatment of the cells with IFNs enhanced IL-1-induced IL-6 production, indicating that IFNs upregulate functional IL-1R. IL-1 and IFNs are produced by the same cell types, as well as by the adjacent different cell types, and are concomitantly present in lesions of immune and inflammatory reactions. These results therefore suggest that IFNs exhibit synergistic effects with IL-1 through upregulation of IL-1R. Augmented production of IL-6 may also contribute to the reactions.

Cell Line↗
PubMed1995

Glycosylated human recombinant interleukin-1 alpha, neo interleukin-1 alpha, with D-mannose dimer exhibits selective activities in vivo.

To investigate the effect of carbohydrate-introduction on IL-1 activity, especially in vivo, and to develop IL-1 with less deleterious effects, recombinant human IL-1 alpha was coupled with mannose dimer, alpha-D-Man-1-6-D-Man [Man2 alpha(1-6)] by an acyl azide method. Previous studies demonstrated that the glycosylated IL-1 exhibited reduced activities compared with original IL-1 in all the experiments performed in vitro. In this study, we investigated the in vivo activities of Man2 alpha(1-6)-conjugated IL-1 alpha. The glycosylated IL-1 alpha exhibited very low pyrogenic activity and alpha 1-acid glycoprotein induction compared with untreated IL-1 alpha. Untreated IL-1 alpha increased the serum level of IL-6, but the glycosylated IL-1 alpha did not. However, the glycosylated IL-1 alpha possessed the same potency as untreated IL-1 alpha in reduction of serum levels of glucose and triglyceride and in recovery of peripheral white blood cells in 5-fluorouracil-treated mice. Therefore, glycosylation of IL-1 appeared to be useful for the development of neoIL-1 with selective activity in vivo.

Animals↗
PubMed1995

Transfection of human melanoma cells with type I interleukin-1 (IL-1) receptor cDNA rendered them IL-1-responsive and revealed the importance of ODC activity down-regulation in IL-1-induced growth inhibition.

A human type I IL-1 receptor expression plasmid has been constructed and used to transfect human melanoma cells (A375-5), which have been shown to be unresponsive to the antiproliferative effect of IL-1. Five stable transfectant cell lines have been established, of which three are sensitive and the other two resistant to the anti-proliferative effect of IL-1. All the transfectant cell lines, but not progenitor A375-5 cells, expressed functional type I IL-1 receptors and could produce IL-6 in response to IL-1, suggesting that the unresponsiveness of A375-5 melanoma cells is exactly due to an IL-1 receptor deficiency. The three IL-1-sensitive stable transfectant cell lines expressed much more type I IL-1 receptor than the IL-1-sensitive A375-6 cells, thus they are expected to be useful for investigating the signal transduction pathway of IL-1-induced growth inhibition in melanoma cells. The two resistant cell lines produced comparable amounts of IL-6 in response to IL-1, as sensitive cell lines did, indicating that IL-6 induction does not play a major role in IL-1-induced growth inhibition in these melanoma cells. The possibility of an IL-6 receptor and/or IL-6 receptor signaling deficiency was ruled out as the IL-1-sensitive and -resistant transfectants responded similarly to a high dose of exogenous human recombinant IL-6. Examination of the ornithine decarboxylase (ODC) activity of recombinant human IL-1 alpha treated cells showed that all the sensitive but none of the resistant cell lines could down-regulate their ODC activity. These results suggest that IL-1-induced ODC activity down-regulation is an important step in the signal transduction pathway of IL-1-induced growth inhibition of melanoma cells.

Cell Division↗
PubMed1995

Differential effects of prostaglandin E1 and prostaglandin E2 on growth and differentiation of murine myeloid leukemic cell line, M1.

Effects of prostaglandins (PGs) of the E series on growth and differentiation of murine myeloid leukemic cell line M1 were studied. PGE1, but not PGE2, inhibited the growth of M1 cells. PGE2 neither inhibited nor augmented the antiproliferative effect of PGE1. PGE1 augmented the differentiation of M1 cells into macrophage-like cells induced by interleukin 6. PGE2, however, did not exhibit any effect on the differentiation. PGE1 caused a marked increased in intracellular cAMP level in M1 cells, whereas PGE2 had no effect. These results indicate that M1 cells are able to respond only to PGE1. Radiolabeled PGE1 binding experiments, however, revealed that there was no specific binding in M1 cells, suggesting that the cells express low numbers of receptors or very low affinity receptors specific for PGE1. Stable agonists of PGI2, iloprost, cicaprost or carbacyclin, also potently inhibited the growth of M1 cells. These findings suggest that PGE1 as well as PGI2 may play a role in the differentiation of monocyte-macrophage lineage cells.

Alprostadil↗
PubMed1994

Purification and characterization of a human erythrocyte-derived growth factor with a wide target cell spectrum.

A cell extract from human erythrocytes promoted the growth of a wide variety of cell types, namely human and mouse myeloid cells, human and mouse T cells, human B cells, human melanoma cells, mouse transformed fibroblast cells, mouse mastocytoma cells, human lung fibroblast cells, and mouse bone marrow fibroblast/stroma-like cells. The growth-promoting activity was acid- and heat-labile and sensitive to proteases, indicating the proteinaceous nature of the molecule. The activity was also lost upon exposure to 2-mercaptoethanol. The novel growth-promoting factor, termed basic growth factor because of its fundamental effect and a wide target cell spectrum, was purified by sequential anion-exchange, hydrophobic, gel filtration, hydroxylapatite, and reverse-phase high performance liquid chromatographies. The purified factor has an apparent molecular mass of 53 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing and nonreducing conditions. The factor migrated at 270 kDa on native gradient polyacrylamide gel electrophoresis. Therefore, the factor consists of a homopolymer of a single polypeptide chain. The purified factor promoted the growth of the same cell types as the cell extract, except for bone marrow cells.

Amino Acids↗
PubMed1994

Resistance to the anti-proliferative effect of IL-1 on human melanoma cell line is associated with endogenous production of IL-1 and IL-6.

A human melanoma cell line (A375-6) became resistant to the anti-proliferative effect of human IL-1 after a long period of culture. Two stable resistant sub-clones were obtained, and the mechanism of the IL-1 resistance was investigated. Resistant cells, but not sensitive cells, appeared to produce constitutively IL-1 activity. The activity was neutralized by anti-IL-1 alpha antibody but not by anti-IL-1 beta antibody. Resistant cells expressed IL-1 alpha but not IL-1 beta mRNA. Therefore, the resistant cells appeared to produce IL-1 alpha mRNA for IL-1 receptor antagonist (IL-1Ra) was not detected in resistant cells, indicating that the resistance is not attributable to IL-1Ra. These resistant cells were also resistant to the anti-proliferative effect of human IL-6, but not to that of human TNF. Resistant cells appeared to produce constitutively IL-6 more than sensitive cells, and IL-6 production both in sensitive and in resistant cells was augmented by exogenous IL-1. Furthermore, constitutive production of IL-6 in resistant cells was inhibited by IL-1Ra. Type I IL-1 receptor (IL-1R) mRNA was expressed equally in resistant and sensitive cells. These data indicate that the resistance is not the result of loss of functional IL-1R and that IL-1 induces IL-6 in an autocrine manner. It is, therefore, conceivable that endogenous IL-1 and IL-6 contribute to IL-1 resistance.

Cell Division↗
PubMed1994

Binding of Legionella pneumophila to macrophages increases cellular cytokine mRNA.

Infection of macrophages with Legionella pneumophila induces formation of interleukin 1 beta (IL-1 beta), but the molecular basis of this is not understood. Binding of bacteria to macrophage surfaces is the first step in an infection process. Therefore, we examined whether this step was sufficient to increase the cellular level of mRNAs for IL-1 beta and other cytokines. To assess the effect of binding of L. pneumophila on the steady-state levels of cytokine mRNAs, cultures of thioglycolate-elicited macrophages from L. pneumophila-susceptible A/J mice were treated with cytochalasin D and infected with L. pneumophila and the total RNA was extracted for analysis by reverse transcription-PCR with primers for IL-1 alpha, IL-1 beta, IL-6, tumor necrosis factor alpha, granulocyte macrophage colony-stimulating factor, and beta interferon (IFN-beta). L. pneumophila treatment increased the cellular steady-state mRNA levels of all cytokines except IFN-beta. To determine the specificity of this effect, macrophage cultures were treated with cytochalasin D and either bacterial lipopolysaccharide, bovine serum albumin-sensitized latex, Salmonella typhimurium, or Escherichia coli. Lipopolysaccharide treatment increased all mRNAs, bovine serum albumin-sensitized latex had no significant effect, and treatment with S. typhimurium or E. coli increased all mRNAs except that of IFN-beta. These results suggested that the binding of gram-negative bacteria to the macrophage surface was sufficient to induce a unique pattern of cytokine mRNAs. Additional studies that examined the characteristics of the bacterial ligands involved indicated involvement of both heat-labile and heat-stable surface ligands.

Animals↗
PubMed1994

Down regulation of Ia expression in macrophages following incubation with mycobacteria.

Macrophages are known to release cytokines in response to various kinds of stimulators. In the present study, peritoneal macrophages from C3H/He or C3H/HeJ mice were incubated in vitro with heat-killed M. lepraemurium, M. intracellulare or M. gordonare for 3 days followed by harvest culture supernatant to analyze cytokine activities. It, therefore, seems that macrophages phagocytizing these mycobacteria, released interleukin-1 (IL-1) and tumor necrosis factor (TNF) in culture media. The amount of release was dose dependent on mycobacteria employed. In addition, macrophages, as already have reported elsewhere, treated with IFN for 2 to 3 days showed enhanced expression of surface Ia; although the expression was inhibited if the cells phagocytized mycobacteria. Similarly, the reduced expression of Ia was observed in peritoneal macrophages from MRL/lpr mice after 3 day-culture with mycobacteria in vitro. More importantly, in the presence of the supernatant obtained from macrophages incubated with mycobacteria, IFN gamma-treated normal macrophages exhibited suppressed expression of Ia. These results demonstrate that cytokine release and reduced expression of surface Ia in macrophages are simultaneous phenomena after phagocytosis of mycobacteria. Suppression of Ia may be in part induced by Ia suppressive factor(s) released from mycobacterium-phagocytized macrophages.

Animals↗
PubMed1994

Meeting report of the International Symposium of Molecular Cell Biology of Macrophages '94.

The International Symposium of Molecular Cell Biology of Macrophages '94 was held on February 10-11, 1994 at Kanazawa University, Kanazawa, Japan. Fifteen speakers including 4 speakers from Europe and the United States were invited to this symposium. In spite of unusual heavy snow and chilly weather, 150 macrophage scientists gathered and had hot discussions. In the meeting reception, a special recognition speech entitled "Dawn of Cytokines--Discovery of a Blastogenic Factor in the Early 1960's" was given by a discoverer of blastogenic factor, Dr. Shinpei Kasakura. This symposium was initiated in 1991 with the intention of bringing together experts of macrophage investigation once a year to discuss progress in the field of molecular cell biology of macrophages.

Animals↗
PubMed1994

Interleukin-1 down-regulates type I interleukin 1 receptor mRNA expression in a human fibroblast cell line TIG-1 in the absence of prostaglandin E2 synthesis.

We have shown that interleukin-1 (IL-1) up-regulates transcription of its own receptor and number of cell surface IL-1 receptor (IL-1R) type I molecule through induction of prostaglandin E2 (PGE2) synthesis and subsequent intracellular cAMP accumulation in a human lung fibroblast cell line (TIG-1). In this study, the effect of IL-1 on IL-1R mRNA expression was investigated in the absence of PGE2 synthesis. In the presence of indomethacin, an inhibitor of cyclooxygenase, IL-1 inhibited the expression of IL-1R mRNA within 4 h after treatment, and the inhibition sustained at least for 24 h. IL-1 beta as well as IL-1 alpha at higher than 1 U/ml exhibited the inhibitory effect. The inhibitory effect of IL-1 was inhibited by cycloheximide suggesting that de novo protein synthesis is required. IL-1 appeared to destabilize IL-1R mRNA within 30 min after treatment of the cells. Furthermore, this effect of IL-1 was not observed in a synthetic medium and was dependent on serum concentrations indicating that a serum component(s) is involved. These results indicate that IL-1 regulates IL-1R mRNA expression in both a positive and negative manner, and that the negative effect represents a negative feedback mechanism.

Cell Line↗
PubMed1994

Development of glycosylated human interleukin-1 alpha, neoglyco IL-1 alpha, coupled with D-mannose dimer: synthesis and biological activities in vitro.

Interleukin 1 (IL-1) is a nonglycosylated cytokine with pleiotropic effects on various cell types. In order to investigate the effect of carbohydrate introduction on IL-1 activity and to develop IL-1 with less deleterious effects recombinant human IL-1 alpha was chemically coupled with mannose dimers, alpha-D-Man1-6-D-Man[Man2(alpha 1,6)] and alpha-D-Man1-4-D-Man[Man2(alpha 1,4)]. About 5 molecules of mannose dimers were introduced per molecule of IL-1. Anti-IL-1 alpha antibody reacted only weakly with the glycosylated IL-1s. Conversely, antibody against the mannose dimer reacted with only glycosylated IL-1. The effect of glycosylation on IL-1 activity was evaluated by measuring a variety of IL-1 activities in vitro, including proliferative effect on T cells, antiproliferative effect on melanoma cells, stimulatory effect on IL-6 synthesis by melanoma cells, and stimulatory effect on prostaglandin E2 synthesis by fibroblast cells. Glycosylated IL-1s exhibited reduced activities, which were 10-fold to more than several hundred-fold lower than those of the original IL-1 alpha depending upon different aspects of activities addressed. Man2(alpha 1,6)-introduced IL-1 exhibited lower activity than Man2(alpha 1,4)-introduced IL-1. The competitive binding of 125I-IL-1 alpha to mouse T cells with unlabeled IL-1s suggests that the reduced activity of glycosylated IL-1s is due, at least partially, to the decrease of their receptor binding abilities.

Animals↗
PubMed1994

Novel growth promoting activity with a wide target cell spectrum is present in extracts of various cell types.

Cell extracts from a variety of cell lines, myeloid, T, B, mastocytoma, fibroblast, melanoma and breast carcinoma of human, mouse and guinea pig promoted the growth of a wide variety of cell types, namely human myeloid cells HL-60, human B cells Daudi, human melanoma cells A375-C6, mouse transformed-fibroblast cells L929, human myelomonocytic cells THP-1. Among them, the activities in extracts from U937, A375-C6 and Daudi were characterized because these extracts exhibited much more potent activity. These growth promoting activities were acid-labile, sensitive to 2-mercaptoethanol and to heat treatment at 50C or 70C for 5 min. The activities were also sensitive to proteases indicating the proteinous nature of these active entities. The molecular weight of activities from A375 and Daudi cells were estimated to be 100,000-150,000 daltons by gel filtration high performance liquid chromatography, while that from U937 cells was 60,000-70,000 daltons. The isoelectric point of these activities were 5.5-6.5.

Animals↗
PubMed1993

Human myelomonocytic cell line THP-1 produces a novel growth-promoting factor with a wide target cell spectrum.

Conditioned medium from a human myelomonocytic cell line THP-1 promoted the growth of a wide variety of cell types, i.e., human and mouse myeloid cells (HL-60, U937, K562, and M1), mouse T-cells (EL-4), human B cells (Daudi and Raji), mouse mastocytoma cells (IC-2), human melanoma cells (A375-C6), mouse transformed fibroblast cells (L929), human lung fibroblast cells (TIG-1), and mouse bone marrow fibroblast/stromal-like cells. The growth-promoting activity was acid-labile. The activity was resistant to 50 degrees C for 5 min but completely lost in 5 min at 70 degrees C. The activity was resistant to treatment with trypsin but sensitive to chymotrypsin alpha, Pronase E, and proteinase K, indicating the proteinous nature of this activity. The activity was lost by dithiothreitol and 2-mercaptoethanol. Molecular weight (M(r) 50,000-70,000) was estimated by gel filtration-high performance liquid chromatography. After the sequential anion exchange, hydrophobic, and hydroxylapatite high performance liquid chromatography, the partially purified factor exhibited the same target cell spectrum as the conditioned medium.

Animals↗
PubMed1993

Regulation by dietary essential fatty acid balance of tumor necrosis factor production in mouse macrophages.

Increasing the dietary alpha-linolenate (18:3n - 3)/linoleate (18:2n - 6) ratio results in an increase in lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNF) production in mouse resident and casein-induced peritoneal macrophages [3]. We found that prostaglandin E2 (PGE2) production is inversely related to TNF production and that indomethacin abolishes the effect of changing the essential fatty acid balance in resident macrophages. The resident macrophages enriched in n - 3 did not produce a significant amount of PGE3. Accordingly, the decreased production of PGE2 appears to be a major negative regulatory factor for enhancement of TNF production in the n - 3 enriched resident macrophages. In casein-induced macrophages the situation is more complex. Indomethacin decreased PGE2 production and increased TNF production; however, the differences in TNF production between the n - 6 enriched and n - 3 enriched macrophages were not completely abolished by indomethacin treatment. Lysosomal acid phosphatase activity, a marker of activation/maturation stages, was elevated in the n - 3 enriched compared to the n - 6 enriched casein-induced macrophages but was similar in the resident macrophages of the two dietary groups. Expression of CD14, which is a receptor for LPS, was not different in casein-induced macrophages of the two dietary groups. Thus, the differences in production of TNF between the n - 3 and n - 6 enriched resident macrophages can be accounted for mostly by a difference in the production of a negative feedback effector, PGE2. However, a significant portion of the TNF production in casein-induced macrophages is regulated by a factor(s) other than PGE2 and LPS receptor; advanced activation/maturation stages induced by the diet high in alpha-linolenate may underlie the enhanced TNF production in casein-induced macrophages.

5'-Nucleotidase↗
PubMed1993

Purification and substrate specificity of beta-xylosidase from sycamore cell (Acer pseudoplatanus L.): application for structural analysis of xylose-containing N-linked oligosaccharides.

A beta-xylosidase was purified 51-fold from culture medium of sycamore (Acer pseudoplatanus L.) cells using p-nitrophenyl beta-D-xylopyranoside as a substrate. This enzyme can remove a xylose residue from asparagine-linked oligosaccharides, derivatized with 2-aminopyridine. A pentasaccharide, Xy1 beta 2Man beta 4GlcNAc beta 4(Fuc-alpha 3)GlcNAc was the favorite substrate in N-linked oligosaccharides, but a xylose residue in Xy1 beta 2(Man-alpha 3)Man beta sequence could not be removed by the enzyme. We also propose an efficient method for detection of xylose residue in N-linked oligosaccharides by a combination of the two-dimensional sugar mapping technique and the xylosidase digestion.

Carbohydrate Sequence↗