Biomedical subjects

T Toida

Publications and source records attributed to T Toida.

At least 73 records · Page 4Linked to original sources

PubMed1995

Analysis of antitumor properties of effector cells stimulated with a cell wall preparation (WPG) of Bifidobacterium infantis.

Intestinal Bifidobacterium species are thought to be beneficial in animal and human intestines. We studied the mechanisms of Bifidobacteria in antitumor activity using a cell wall preparation (WPG) of B. infantis (Cancer Res., 45, 1300, (1985)). WPG enhanced the in vitro antitumor activities of mouse peritoneal exudate cells elicited with proteose-peptone (P-PEC) and thioglycollate broth (TG-PEC), determined by cytostatic ([3H]thymidine uptake inhibition) and cytolytic ([3H]uridine release) assays. Tumor necrosis factor-alpha (TNF-alpha) and reactive nitrogen intermediates (RNI) play a role in such augmented cytotoxicity, because anti-TNF-alpha antibody almost completely blocked the increased cytolytic activity of P-PEC in the presence of WPG. Moreover, WPG induced RNI in the supernatant of TG-PEC in a dose-dependent manner. The mRNA expression of several cytokines (IL-1 beta, IL-6, IL-10, IFN-alpha and TNF-alpha) was induced in BALB/c mouse peritoneal cells 3 h after an intraperitoneal injection of WPG (3 h WPG-PEC). However, this expression disappeared from 24 h WPG-PEC, except for that of IFN-alpha. IFN-gamma was not induced. Kinetic studies of the tumor neutralizing activities of the WPG-PECs by means of the in vivo Winn assay revealed that the activity emerged at 1.5 h, became maximal at 3 h and disappeared at 24h. These results indicated that Bifidobacterial WPG is a Biological Response Modifier (BRM) with characteristics similar to those of other bacterial BRMs.

Animals↗

PubMed1995

Determination of a depolymerized holothurian glycosaminoglycan in plasma after intravenous administration by postcolumn HPLC.

Depolymerized holothurian glycosaminoglycan (DHG) produced artificially from sea cucumber has an anticoagulant and antithrombotic activity. In the present study, we attempted to determine the plasma level of DHG using a chemical procedure. A general method for determination of chondroitin sulfates by forming unsaturated disaccharides with chondroitinase digestion was difficult to apply to DHG, because it was resistant to any chondroitinase digestion. We therefore developed a highly sensitive postcolumn HPLC method for determination of intact DHG. DHG was applied to Asahipak NH2P-50, an amino-bonded column, eluted by alkaline solution and then detected with arginine under a strong alkaline condition as a postcolumn reagent. The limit of detection for DHG was 10 ng. The present method was applicable to the determination of DHG in plasma after intravenous administration.

Animals↗

PubMed1994

Time-course experiment involving glycosaminoglycan synthesis by normal human dermal fibroblasts cultured in the presence of dibutyryl cyclic AMP and retinoic acid.

Glycosaminoglycan (GAG) synthesis by cultured normal human dermal fibroblasts was examined. Hyaluronic acid (HA) synthesis reached a maximum on day 3 (0.3 microgram/ml medium) and then decreased to a low level (0.15 microgram/ml medium). The amounts of dermatan sulfate (DS) and chondroitin sulfate (ChS) synthesized by the cells increased with increasing cell numbers during the initial stage to attain constant levels (0.092 microgram DS/ml, 0.026 microgram ChS/ml medium) after the cells reached confluence. We also tested the effects of dibutyryl cyclic AMP (dbcAMP) and retinoic acid (RA) on GAG synthesis by the cells. The synthesis of HA and ChS by cells was stimulated, when the cells were cultured in medium containing dbcAMP (0.1 mM), whereas DS synthesis was scarcely affected. However, addition of RA (5 mM) suppressed GAG synthesis by the cells.

Bucladesine↗

PubMed1994

Analytical studies on hyaluronic acid synthesis by normal human epidermal keratinocytes cultured in a serum-free medium.

Hyaluronic acid (HA) synthesized by normal human epidermal keratinocytes cultured in a serum-free medium was monitored by a highly sensitive HPLC method, which was established by us for the simultaneous determination of HA, chondroitin sulfate (ChS) and dermatan sulfate (DS) as their unsaturated disaccharides. The major glycosaminoglycan (GAG) in the medium of the keratinocytes was HA, and the ability of the cells to synthesize HA increased relatively with an increase in cell numbers during the logarithmic phase and reached a maximum level after the cells became confluent. HA synthesis by the keratinocytes was inhibited by the addition of calcium chloride to the culture medium, and was strongly stimulated by the addition of retinoic acid (RA), respectively. It was shown that the ability of the cells to synthesize HA exists in the spinous cell stage. Furthermore, we found that HA synthesis by the cells was slightly increased by the addition of dibutyryl cyclic AMP (dbcAMP). Our results indicate that the measurement of time-course levels of HA in the culture medium is useful for the screening of active substances for proliferation and differentiation of the keratinocytes.

Bucladesine↗

PubMed1994

Adjuvant activity of the cell wall of Bifidobacterium infantis for in vivo immune responses in mice.

We examined the adjuvant activity of the Bifidobacterial Cell Wall preparation (WPG) for in vivo immune responses in mice. We studied three classical immune responses, which are thought to be T-cell mediated responses, to evaluate the adjuvant activity of WPG. The delayed type hypersensitivity (DTH) responses of sheep blood red cell (SRBC)-sensitized mice were significantly augmented by WPG, although the enhancement varied with the timing, route and dosage of injection. The adjuvant activity of WPG was also confirmed by using a glutaraldehyde treated- and Concanavalin A associated- tumor vaccine (G-Con A tumor vaccine) system. BALB/c mice sensitized with G-Con A tumor vaccine and WPG improved synergistically in survival time and cure rate compared with those given G-Con A vaccine alone. Spleen cells of Meth A tumor-bearing mice induced antitumor neutralizing activity with the growth of tumor but the activity declined and disappeared at the late stage of tumor growth (over 28 days after tumor transplantation). On the other hand, antitumor neutralizing immunity was prolonged for as long as 33 days in mice inoculated with Meth A tumor and WPG. The requirement of a T-cell subpopulation in the spleen cells of tumor plus WPG treated mice was confirmed using anti-Thy 1.2 antiserum + complement to deplete them. The adjuvant activities of the Bifidobacterial cell wall demonstrated by the in vivo immune responses predict that Bifidobacteria may play a role as an immunomodulator in human and animal intestines.

Adjuvants, Immunologic↗

PubMed1993

Separation and characterization of dermatan sulfate in normal human urine.

Dermatan sulfate excreted in normal human urine was isolated and characterized by TLC and cellulose acetate strip electrophoresis after cetylpyridinium chloride precipitation and pronase digestion. In these separation methods, dermatan sulfate and chondroitin sulfate were extracted and then monitored by sensitive HPLC methods with post column fluorometric derivatization coupled with chondroitinase ABC, ACII and B digestion. From the results, we demonstrated that human urinary dermatan sulfate contains iduronic acid as its major uronic acid (80-90% of total uronic acid), and is composed mainly of repeated mono-sulfated disaccharide units [Di-4S (structure shown in Fig. 1), 89%] and small numbers of di-sulfated disaccharide units (Di-diSB, 7% and Di-diSE, 1%).

Chondroitin Sulfates↗

PubMed1993

Structural analysis of a low-sulfated chondroitin sulfate chain in human urinary trypsin inhibitor.

The low-sulfated chondroitin 4-sulfate(LSC) chain from human urinary trypsin inhibitor was purified and the structure was characterized. After hyaluronidase SD digestion of LSC, an oligosaccharide which contains the linkage region could be obtained. The structure of oligosaccharide was analyzed by HPLC and 500 MHz 1H-NMR spectroscopy. The analytical results revealed that 4-O-sulfo GalNAc residues were located in the neighborhood of the linkage region.

Carbohydrate Conformation↗

PubMed1992

Chemiluminescence high-performance liquid chromatography for the determination of hyaluronic acid, chondroitin sulphate and dermatan sulphate.

A sensitive chemiluminescence high-performance liquid chromatographic method has been developed for the determination of hyaluronic acid, chondroitin sulphate and dermatan sulphate as their unsaturated disaccharide-dansylhydrazine derivatives involving an effective sample clean-up system. The dansylhydrazones of the unsaturated disaccharides derived from the hyaluronic acid, chondroitin sulphate and dermatan sulphate by chondroitinase ABC and/or chondroitinase ACII, were separated by reversed-phase chromatography using a mixture of 0.1 M sodium acetate buffer (pH 6.0) and 80% acetonitrile on a column (250 mm x 4.0 mm I.D.) packed with amide-80 silica beads (5 microns diameter). For post-column elution in the chemiluminescence system, 1 mM bis[2-(3,6,9-trioxadecanyloxycarbonyl)-4-nitrophenyl]oxalate and 3mM hydrogen peroxide in acetonitrile were used. The detection limit of each glycosaminoglycan was 100 fmol. The method was applicable to the determination of the levels of hyaluronic acid, chondroitin sulphate and dermatan sulphate in rat peritoneal mast cells.

Animals↗

PubMed1992

Structural analysis of the N-linked oligosaccharides from human urinary trypsin inhibitor.

The N-linked oligosaccharides from human urinary trypsin inhibitor were purified and their structures were investigated by compositional analysis, the two-dimensional sugar map method and 500 MHz 1H-NMR. The results revealed that they were composed of disialosyl, monosialosyl and asialosyl oligosaccharides, which have the common biantennary core structure; Gal1-4GlcNAc1-2Man1-3(Gal1-4GlcNAc1-2Man1-6)M an1-4GlcNAc1-4GlcNAc.

Carbohydrate Conformation↗

PubMed1991

Microdetermination of hyaluronic acid in human urine by high performance liquid chromatography.

A high performance liquid chromatographic (HPLC) system is described for determination of the unsaturated disaccharide (delta Di-HA) derived from hyaluronic acid (HA) in human urine by digestion with hyaluronidase SD. The effects of eluents on the separation of delta Di-HA and delta Di-0S, which is derived from the reaction of chondroitin with the enzyme, have been studied. The established chromatographic conditions were as follows--column: a stainless steel tube (4 mm i.d. x 250 mm) packed with TSKgel NH2-60; eluent: a mixture of acetonitrile and 0.1 M Tris-HCl buffer containing 0.1 M boric acid and 10 mM sodium sulphate, pH 7.0 (64:36, v/v). The strong fluorescence of unsaturated disaccharide after the reaction with 2-cyanoacetamide in alkaline medium was used for post-column detection. The calibration curve for delta Di-HA was linear in the range 5 pmol-5nmol with a practical detection limit of 2 pmol. The assay coefficients of variation (n = 5) at 200 pmol for delta Di-HA and delta Di-0S were 1.7 and 1.5%, respectively. This HPLC system has been applied to the determination of HA in human urine.

Adult↗

PubMed1991

Crystal structure of Streptomyces erythraeus trypsin at 2.7 A resolution.

The crystal structure of Streptomyces erythraeus trypsin (abbreviated as SET) has been determined in order to clarify the precise structure of the vicinity of the active site of serine protease and to understand its structure-function relationship. Crystals of SET were prepared at its active pH range (pH 5-10) without any inhibitors which might have affected the circumstances around the active sites. The structure model of SET was made based on the electron density map obtained by the multiple isomorphous replacement method at 3.5 A resolution, and refined by the restrained least-squares method. The current model yields a crystallographic R-factor of 0.272 for 4,968 reflections between 8 and 2.7 A resolution. Though the sequence homology among SET, Streptomyces griseus trypsin and bovine trypsin, 32-37%, is not so high, their overall structures are similar to each other. Comparison of the three molecular structures shows that: 1) the folding of the main chains of the three proteins is essentially the same though there are significant differences on the molecular surface; 2) the spatial arrangements of the catalytic triads in the three proteins are similar to each other; 3) in SET and S. griseus trypsin a short stretch of 3(10)-helix is found through Ala56 to Thr59; His57 in this segment is one important amino acid residue involved in the active sites.

Amino Acid Sequence↗

PubMed1991

[Metabolic study on chondroitin sulfates in rabbits].

The metabolic studies on three types of chondroitin sulfates in rabbits were performed. The analyses of chondroitin sulfates gave the following results concerning their metabolic behavior. 1) Desulfation of chondroitin sulfates on GalNAc C6 position was observed more clearly after the administration of low molecular weight chondroitin sulfate (molecular weight, 6000 and 16,000) than after that of high molecular weight one (molecular weight, 50,000). 2) The disappearance velocities of the administered low molecular weight chondroitin sulfates in the blood and those of excretion into the urine were faster than those of high molecular weight one. 3) Intact chondroitin sulfates were excreted into the urine after the administration of low molecular weight chondroitin sulfates, while chondroitin sulfates having more than 30000 molecular weight were not detected in the collected urine after the administration of high molecular weight one.

Animals↗

PubMed1990

Isolation and characterization of major urinary amino acid O-glycosides and a dipeptide O-glycoside from a new lysosomal storage disorder (Kanzaki disease). Excessive excretion of serine- and threonine-linked glycan in the patient urine.

Four major sialo compounds, termed GP-M1, GP-D1, GP-D2, and GP-D3 have been isolated from the urine of a novel glycoprotein storage disorder patient with angiokeratoma corporis diffusum which was discovered by Kanzaki et al. (Kanzaki, T., Yokota, M., Mizuno, N., Matsumoto, Y., and Hirabayashi, Y. (1989) Lancet April 22, 875-877). Based on the results of fast atom bombardment mass spectrometry, methylation analysis, and proton nuclear magnetic resonance spectroscopy, their chemical structures were concluded to be: (formula; see text) The yields of GP-M1, GP-D1, GP-D2, and GP-D3 were approximately 15, 6, 50, and 5 mg/liter of urine, respectively. The most major compound GP-D2, was further purified into single molecular species, threonine and serine type, by reversed phase high performance liquid chromatography. NMR analysis of the two purified compounds with single molecular species showed that the chemical shifts of anomeric protons of GalNAc were significantly different between threonine- and serine-linked GalNAc. Neither mannose-containing glycopeptides nor glycosphingolipids were excreted in the patient urine. From these results, this disease is thought to be caused by the deficiency of a lysosomal enzyme(s) acting on O-linked glycan chains.

Carbohydrate Metabolism, Inborn Errors↗

PubMed1990

Cloning and sequence analysis of the arginine deiminase gene from Mycoplasma arginini.

Arginine deiminase from Mycoplasma arginini was purified. The purified enzyme has a molecular weight of 46,000 daltons as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Its specific activity (20 units/mg protein) and amino acid composition showed a strong similarity to that of the Mycoplasma arthritidis arginine deiminase. The amino acid sequences of the N-terminal region and three internal peptides generated by enzymatic cleavage of the purified protein were determined. Using a synthetic oligonucleotide mixture complementary to part of the determined N-terminal amino acid sequence, the gene coding for arginine deiminase was isolated from a phage library. A nucleotide sequence of 2189 bp encoding the gene was determined. An open reading frame (ORF) contained the amino acid sequences corresponding to the determined N-terminal region and the three internal peptides of arginine deiminase. Thus it was concluded that this ORF encoded the arginine deiminase, a 385 amino acid polypeptide (mol.wt. 43,900 daltons). The three tryptophan residues in the sequenced peptides align with UGA codons in the nucleotide sequence, indicating that the nonsense codon UGA is used as a tryptophan codon in M. arginini.

Amino Acid Sequence↗

PubMed1990

Surface-active novel glycolipid and linked 3-hydroxy fatty acids produced by Serratia rubidaea.

A Serratia rubidaea isolate with wetting activity when grown at 30 but not 37 degrees C was examined for the production of specific lipids. Two novel lipids (rubiwettins R1 and RG1) were isolated and shown to be able to lower the surface tension of saline to 26 mN/m. These lipids were located in extracellular vesicles found in a 30 degrees C culture of S. rubidaea. Chemical structures of these biosurfactants were determined by degradation product analyses, infrared spectroscopy, mass spectrometry, and proton nuclear magnetic resonance spectroscopy. Rubiwettin R1 was proposed to be a mixture of 3-(3'-hydroxytetradecanoyloxy)decanoate, 3-(3'-hydroxyhexadecenoyloxy)decanoate, and minor molecular isomers. The structure of rubiwettin RG1 was proposed to be beta-D-glucopyranosyl 3-(3'-hydroxytetradecanoyloxy)decanoate. The importance of such surface-active exolipids in bacterial occupancy on surfaces was suggested.

Chromatography, Thin Layer↗

PubMed1990

Metabolic pathways of sodium bisulfite injected intravenously in rabbits.

The metabolites of sodium bisulfite in rabbits through intravenous injection were examined according to the high performance liquid chromatography method for determination of sulfur-containing substances. Most of the administered sulfite was oxidized to sulfate, and a small part was converted to thiosulfate, S-sulfoalbumin, S-sulfoglutathione and S-sulfocysteine. Furthermore, it was found that S-sulfocysteine administered intravenously to a rabbit was partially changed to inorganic sulfate and thiosulfate. These metabolites produced from sulfite in rabbits indicated the presence of the many and complicated metabolic pathways of sulfite in vivo.

Animals↗

PubMed1989

Mono-sulfated globopentaosylceramide from human kidney.

A novel sulfated glycosphingolipid that belongs to the "globo-series" was isolated from human kidney. This lipid was purified from a pooled kidney preparation by chloroform/methanol extraction, mild alkaline treatment, DEAE-Sephadex and silicic acid column chromatographies, and preparative thin layer chromatography. The structure and the properties were studied by infrared spectroscopy, two-dimensional proton magnetic resonance spectroscopy, negative secondary ion mass spectrometry, solvolysis, compositional and methylation analyses, monoclonal antibodies, and sulfatide-binding proteins. From the results of the above analyses, the structure of this glycolipid was proposed to be HSO3-3Gal beta 1-3GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4Glc beta 1-1ceramide. The proton resonance at 3.93 ppm of the H-3 of the sulfated nonreducing terminal galactose of this lipid was downfield-shifted (delta 0.48 ppm), as compared with H-3 of the internal beta-galactose because of the electronegativity of the sulfate ester. This sulfated lipid reacted with a monoclonal anti-SSEA-3 (MC-631) (Kannagi, R., Cochran, N. A., Ishigami, F., Hakomori, S., Andrews, P. W., Knowles, B. B., and Solter, D. (1983) EMBO J. 2, 2355-2361), whose epitope is R-3GalNAc beta 1-3Gal alpha 1-4Gal beta 1-R', on thin layer chromatograms and solid-phase radioimmunoassay. This lipid also bound to the 125I-labeled sulfatide-binding protein, thrombospondin. The yield of this sulfated glycolipid was 0.19 nmol/g of tissue, which was about 0.09 and 0.5 mol % of galactosyl and lactosyl sulfatides in human kidney.

Chromatography, Ion Exchange↗

PubMed1989

A sulfated glucosylceramide from rat kidney.

A novel sulfated glycosphingolipid containing a sulfated glucosyl residue was isolated from rat kidney and purified to homogeneity by column chromatographies with DEAE-Sephadex and silica beads. By compositional analyses, permethylation studies, one- and two-dimensional proton magnetic resonance spectroscopy, infrared spectroscopy, negative secondary ion mass spectrometry, solvolysis, and immunostaining on thin layer chromatogram, the structure of this glycolipid was proposed to be HSO3-3Glc beta 1-1Cer (where Cer is ceramide). The ceramide portion consisted of 4-D-hydroxysphinganine as the sole long chain base, and the fatty acid consisted of predominantly tetracosanoic acid, deduced from both composition analysis and negative secondary ion mass spectrometry. The yield of glucosyl sulfatide was about 5 nmol/g of tissue, being about three times as much as that of lactosylceramide sulfate.

Animals↗