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Biomedical subjects

T Nishihara

Publications and source records attributed to T Nishihara.

At least 217 records · Page 12Linked to original sources
PubMed1988

Serological properties and immunobiological activities of lipopolysaccharides from black-pigmented and related oral Bacteroides species.

Lipopolysaccharides (LPS) from five species of oral Bacteroides, B. gingivalis strains 381 and ATCC 33277, B. oralis ATCC 33269, B. loescheii ATCC 15930, B. intermedius ATCC 25611 and B. corporis ATCC 33547, were extracted from whole cells by the phenol/water procedure, and subsequently purified by treatment with nuclease and ultracentrifugation. The LPS were composed of hexoses, glucosamine, fatty acids and phosphorus. Heptose and 2-keto-3-deoxyoctonate were not detected. The LPS preparations from B. gingivalis strains 381 and ATCC 33277 presented very similar SDS-polyacrylamide gel electrophoresis patterns when stained with ammoniacal silver. They produced a fused precipitin band against an antiserum to B. gingivalis 381 LPS in immunodiffusion tests. Antisera raised against the LPS from B. loescheii and B. intermedius reacted with the LPS prepared from all the oral Bacteroides strains except those of B. gingivalis. All the LPS preparations were mitogenic for spleen cells of BALB/c (nu/nu) mice, but not for thymus cells from C3H/HeN mice. The LPS induced marked mitogenic responses and polyclonal B cell activation for spleen cells of not only C3H/HeN (LPS responder) mice, but also C3H/HeJ (LPS nonresponder) mice. The mitogenic responses were not suppressed significantly upon addition of polymyxin B to the reaction mixture. These LPS also enhanced interleukin-1 production by murine peritoneal macrophages and mouse cell line J744. 1 macrophages. Hydrolysis of B. gingivalis ATCC 33277 LPS in 1 m-HCl at 100 degrees C for 1 h yielded lipid and polysaccharide. The lipid portion was largely composed of fatty acids and glucosamine, and was mitogenic for spleen cells from C3H/HeJ as well as C3H/HeN mice, while the polysaccharide portion induced no significant mitogenic responses under similar experimental conditions.

Animals↗
PubMed1988

Immunobiological properties of lipopolysaccharides isolated from Fusobacterium nucleatum and F. necrophorum.

Lipopolysaccharides (LPSs) were isolated from Fusobacterium nucleatum ATCC 10953 and F. necrophorum ATCC 25286 by the hot phenol/water procedure. F. nucleatum LPS was composed of 16% (w/w) carbohydrate, 10% (w/w) hexosamine and 40% (w/w) fatty acid, while F. necrophorum LPS was composed of 26% (w/w) carbohydrate, 12% (w/w) hexosamine and 28% (w/w) fatty acid. These LPS preparations induced mitogenic responses in spleen cells of BALB/c, BALB/c (nu/nu) and C3H/HeN mice, and these responses were suppressed by the addition of polymyxin B. The preparations also induced the polyclonal responses of C3H/HeN spleen cells. In addition, enhanced glucose utilization and interleukin-1 production by murine peritoneal macrophages were demonstrated. Neither spleen cells nor macrophages from the 'LPS-nonresponsive' C3H/HeJ mouse were activated by LPSs from the Fusobacterium species.

Animals↗
PubMed1988

Immunological detection of 22K protein in sporulating cells of Bacillus megaterium ATCC 12872.

The synthesis and deposition of 22,000-dalton (22K) spore coat protein were examined immunochemically on the sporulating cells of Bacillus megaterium ATCC 12872 using the antibody to purified 22K spore coat protein. This antibody cross-reacted with 44K and 25K proteins in immunoblot analysis of dormant spore coat proteins. Immunoblot analysis on the sporulating cells showed that 22K protein was detected from t8 in forespore coat protein fractions. Sandwich enzyme immunoassay revealed that 22K protein in the spore coat protein fraction appeared at t6 and reached a plateau at t9, and 22K protein in the mother cell cytoplasmic fraction was detected at only t7 and t8 at a very low level.

Bacillus megaterium↗
PubMed1988

Isolation and characterization of outermost layer deficient mutant spores of Bacillus megaterium.

Outermost layer deficient mutant spores of Bacillus megaterium ATCC 12872 were isolated by Urografin density gradient centrifugation after mutagenesis with ethyl methanesulfonate. Although the composition of the cortex peptidoglycan was the same as that of the parent spores, three major proteins (48, 36, and 22 K daltons) were missing, suggesting that these proteins are components of the outermost layer. All mutant spores were also found to have very hydrophobic surface by 'salt aggregation test,' which would facilitate selection of such mutants.

Bacillus megaterium↗
PubMed1988

Immunoelectron microscopic studies on spore coat proteins of Bacillus megaterium.

An immunochemical staining technique for the spore coat proteins of Bacillus megaterium ATCC 12872 was developed using colloidal gold as a second antibody. For reducing the non-specific immunogold binding and increasing the specific binding, the affinity-purified IgG was used as a first antibody. In sporulating cells at t10, gold particles were found not only in the spore coat but also in the mother cell cytoplasm, suggesting that some coat proteins were synthesized in the cytoplasm. Use of the specific affinity-purified antibody to 48K-protein demonstrated that this protein was one of the components of the outer coat.

Bacillus megaterium↗
PubMed1988

Production of an interleukin-1 inhibitor by cell line P388D1 murine macrophages stimulated with Haemophilus actinomycetemcomitans lipopolysaccharide.

Murine macrophages of the P388D1 cell line stimulated with lipopolysaccharide (LPS) from Haemophilus actinomycetemcomitans Y4 released an interleukin-1 (IL-1) inhibitor, as well as IL-1. Maximal IL-1 activity in culture supernatants was detected after 24 h of culture. On the other hand, IL-1 inhibitor activity reached a maximum level after 72 h of culture. An IL-1 inhibitor was partially purified from the culture supernatant of P388D1 cells stimulated with Y4 LPS for 72 h by ammonium sulfate precipitation, followed by Sephacryl S-200 gel chromatography. A 160-kilodalton peak inhibitory to IL-1 and a 14-kilodalton peak showing IL-1 activity were separated by Sephacryl S-200 column chromatography. The partially purified IL-1 inhibitor significantly suppressed the proliferation of C3H/HeJ murine thymocytes that had been induced with murine and human IL-1 in the presence of a submitogenic dose of concanavalin A. The IL-1 inhibitor more strongly suppressed human recombinant IL-1 beta than human recombinant IL-1 alpha. This inhibitory activity of the partially purified preparation was unaffected by the presence of trypsin inhibitor and the protease inhibitor aprotinin. The IL-1 inhibitor did not exhibit either IL-2 or IL-2 inhibitor activity. The inhibitor suppressed C3H/HeJ thymocyte proliferation induced by IL-1 in the presence of a saturated concentration of IL-2 instead of a suboptimal concentration of concanavalin A. These results indicate that prolonged culture of Y4 LPS-stimulated murine macrophages releases a specific inhibitor of IL-1.

Animals↗
PubMed1988

Suppression of murine macrophage interleukin-1 release by the polysaccharide portion of Haemophilus actinomycetemcomitans lipopolysaccharide.

Lipopolysaccharide (LPS) was extracted from whole cells of Haemophilus actinomycetemcomitans Y4 by the hot phenol-water procedure. LPS was cleaved into its lipid A and polysaccharide moieties by hydrolysis in 1% acetic acid. The major component sugars of the polysaccharide were glucose, heptose, rhamnose, galactose, and fucose. LPS and lipid A from H. actinomycetemcomitans induced the release of interleukin-1 (IL-1) by LPS-responsive C3H/HeN murine peritoneal macrophages and cell line macrophages (P388D1 and J744.1), but not by LPS-nonresponsive C3H/HeJ peritoneal macrophages. The polysaccharide was unable to induce the release of IL-1. It suppressed the IL-1 release from LPS- and lipid A-stimulated macrophages, but not the production of cell-associated and intracellular IL-1. The addition of rhamnose, a sugar component of the polysaccharide, abrogated the inhibitory effect of the polysaccharide on IL-1 release. These results suggest the participation of a lectinlike molecule in IL-1 release.

Cell Line↗
PubMed1988

Purification, characterization, and primary structure of Escherichia coli protease VII with specificity for paired basic residues: identity of protease VII and OmpT.

Escherichia coli cells were found to contain a novel outer membrane-associated protease, designated protease VII (K. Sugimura and N. Higashi, J. Bacteriol. 170:3650-3654, 1988). This enzyme was purified to homogeneity and exhibited an apparent molecular weight of 36,000 on sodium dodecyl sulfate gels and 180,000 on a TSK G-3000SW column in the presence of Triton X-100. It was capable of cleaving several peptides at the center of paired basic residues but not at single basic residues, implying that it is distinct from trypsinlike proteases. Protease VII was most active at pH 6.0 and was sensitive to a serine protease inhibitor, diisopropylfluorophosphate, and to the bivalent cations Zn2+, Cu2+, and Fe2+. The nucleotide sequence of a protease VII gene-carrying DNA fragment, which had been cloned by complementation analysis (K. Sugimura, Biochem. Biophys. Res. Commun. 153:753-759, 1988) was determined. It carried two putative promoter regions and a putative Shine-Dalgarno sequence in addition to the complete structural gene, which encoded pre-protease VII of 317 amino acid residues, with the N-terminal 20 residues being a signal peptide. By comparing their amino acid sequences, protease VII and OmpT, which specifically cleaves ferric enterobactin receptor protein, were found to be identical.

Amino Acid Sequence↗
PubMed1988

Effects of tumor necrosis factor on cell growth and expression of transferrin receptors in human fibroblasts.

Human recombinant tumor necrosis factor (TNF) stimulated the growth of confluent human fibroblasts (FS-4) in the presence of fetal calf serum. Epidermal growth factor (EGF) similarly stimulated cellular growth; however other mitogenic factors such as insulin, fibroblast growth factor, 12-O-tetradecanoyl-phorbol-12-acetate and Ca2+ ionophore A23187 did not. The growth-stimulating action of TNF was not synergistic with the activity of EGF in the presence of serum. TNF induced a rapid increase in the binding of transferrin to the cell surface, followed by a return to the basal level within 5 min. A similar increase in transferrin binding was observed in FS-4 cells exposed to EGF. In contrast, insulin caused a prolonged stimulation of transferrin binding. These results suggest that TNF and EGF generate similar or identical intracellular signals for cellular growth and the regulation of transferrin receptor expression.

Cell Division↗
PubMed1988

Effects of nitrogen dioxide on Mycoplasma pulmonis infection and humoral immune responses in mice.

The effects of continuous exposure to nitrogen dioxide (NO2) on the pathologic and immunologic responses of ddY mice to the infection with Mycoplasma pulmonis were investigated. The organisms grew well in the trachea as early as 7 days after infection but barely grew in the lung even after 28 days, causing slight pneumonic lesions in only a few of the infected mice exposed to 1 and 5 ppm NO2. When mice were exposed to 10 ppm NO2 at or after the infection, however, mycoplasmal growth in the lung, but not in the trachea, was greatly enhanced, and pneumonic lesions were evident in the lung of almost all the mice examined. The serum antibody titers to M. pulmonis increased with time after infection regardless of the concentration of NO2 exposed or the mycoplasmal number in the respiratory tract in the infected mice. The in vitro immune responses of the spleen cells of the infected mice were significantly depressed by exposure to 10 ppm NO2 in not only mitogenic response to LPS and ConA but also antibody production to SRBC, whereas uninfected healthy mice were apparently not modulated except for a slight decrease in Con A response.

Animals↗
PubMed1987

Site-directed cleavage of RNA.

Using complementary chimeric oligonucleotides containing deoxyribonucleotides and 2'-O-methylribonucleotides (1), enzymatically synthesized RNA (90 mer) were cleaved at a single site with Escherichia coli RNaseH, either at a hairpin loop or at a stem region. Especially, site-specific cleavage occurred in even the target region being enclosed within a stable, base-paired stem. The method is proved to be generally applicable to RNA containing secondary structures.

Base Sequence↗
PubMed1987

Induction of activated natural killer cells from murine spleen cells primed in vivo and subsequently challenged in vitro with the streptococcal preparation OK432.

The present study shows that natural killer cell-mediated cytotoxicity of BALB/c mouse spleen cells to syngeneic tumor cells was augmented by in vivo priming or in vitro stimulation with the streptococcal preparation OK432. The augmentation of spleen cell cytotoxicity to syngeneic tumor cells by in vivo priming alone with OK432 was lower than that obtained by in vitro stimulation alone with OK432. When the murine spleen cells primed in vivo with OK432 were rechallenged in vitro with OK432 at various intervals, the natural cytotoxicity was more strongly enhanced than that seen with in vitro stimulation alone. The cell surface phenotype of killer cells activated with OK432 was Thy 1+ and asialo GM1+, suggesting the activated natural killer cell. Next, mice were transplanted with syngeneic colon adenocarcinoma cells, and primed in vivo with OK432. These spleen cells were subsequently challenged in vitro with OK432. These spleen cells displayed a strong cytotoxic activity not only to the transplanted adenocarcinoma cells but also to other syngeneic tumor cells.

Adenocarcinoma↗
PubMed1987

Regulatory action of immune B cells on delayed-type hypersensitivity in mice.

Suppressor cells in delayed-type hypersensitivity (DTH) to soluble protein antigens were induced in vitro from BALB/c spleen cells. Transfer of these cells into syngeneic recipients resulted in suppression of the hosts' DTH responses in an antigen-specific manner. These suppressor cells were characterized as B cells by their adherence to nylon-wool columns, resistance to treatment of anti-Thy 1, -Ly 1, and -Ly 2 antibodies plus complement, adherence to anti-mouse immunoglobulin-coated dishes, and nonadherence to uncoated plastic dishes. In addition to being radiation sensitive, these suppressor B cells showed the capability of binding to the primed antigen. Thus, it was demonstrated that our in vitro-induced suppressor cells were antigen-specific B cells. When these suppressor B cells were transferred into the recipients, serum titers of specific antibodies were elevated and effector phase suppressor T cells were induced in the recipients. These results suggest that suppressor B cells exert their suppressor activity through the idiotype-anti-idiotype network.

Animals↗
PubMed1987

Comparative studies of the biological activities of human tumor necrosis factor and its derivatives.

Biological activities of human tumor necrosis factor (TNF) and its derivatives were compared. In cytotoxicity assay with L929 cells, one derivative, designated as TNF(Asn), showed significantly lower activity than any other TNF examined. In binding assay, this derivative was also shown to have lower affinity for TNF receptors on L929 cells, suggesting that the cytotoxic activity of TNFs on L929 cells correlates with their affinity for receptors. We also found that the cytotoxic activity of TNF on A673 cells and its inhibitory effect on lipoprotein lipase were parallel with the cytotoxic activity on L929 cells, but the growth-enhancing activity on FS-4 cells and the cytotoxic activity on endothelial cells were not. It was also shown that TNF(Asn) had lower affinity than any other TNF for receptors on these target cells tested. These results suggested that there might be at least two types of cellular responses to TNF; one might correlate with the receptor-binding affinity of TNFs and the other not.

Animals↗
PubMed1987

Tumor necrosis factor-induced permeability increase of negatively charged phospholipid vesicles.

The effect of human tumor necrosis factor (TNF) on the permeability properties of liposomes containing phosphatidylserine at pH 5-6, as demonstrated by the calcein efflux. However, it did not induce any permeability change in such liposomes at neutral pH. The TNF-induced calcein efflux was also observed when an other acidic lipid was used as a component of the liposomes, i.e., phosphatidic acid or dicetyl phosphate. On the other hand, liposomes composed of neutral phospholipids such as phosphatidylcholine, phosphatidylethanolamine, and sphingomyelin showed little increases in permeability when incubated with TNF above pH 5.0. The TNF-induced permeability change was inhibited by the addition of polyaspartic acid, while it was not affected by the presence of 0.5 mM calcium ions. These data suggest that the negative charges on the liposomal surface trigger the interaction between TNF and liposomes. However, when the pH of the reaction mixture was decreased to 4.5, TNF-induced calcein efflux was observed even from neutral liposomes. When TNF was incubated with 8-anilinonaphthalene-1-sulfonic acid, the fluorescence intensity of this fluorophore increased with a decrease in the pH of the solution from 7 to 5, and a drastic increase in fluorescence was observed at pH 4.5. These data suggest that the hydrophobic region of TNF is also important for liposomal damage. Furthermore, the potencies of TNF and its derivative as to the induction of the permeability change paralleled their cytotoxic effects on mouse L929 cells, suggesting that the effect of TNF on liposomal membranes is related to its biological action.

Anilino Naphthalenesulfonates↗