Biomedical subjects

S Tsuboi

Publications and source records attributed to S Tsuboi.

At least 19 recordsLinked to original sources

PubMed1995

Effect of stopping fluoride administration on the distribution profiles of fluoride in three different kinds of rat bones.

The aim of this work was to explore the reduction of fluoride concentrations in the skeleton after stopping experimental fluoride administration. Fluoride was administered to the rats at varying doses (0, 50, 100 ppm in drinking water) and for different lengths of time (4, 13, 25 weeks). A series of fluoride concentrations across the full thickness of humerus, parietal bone, and vertebra arch in rats were measured by means of an abrasive micro-sampling technique. The distribution profiles of fluoride from periosteal to endosteal surfaces, which were apparently related to the histological structure of these bones, were U shaped in the humerus, V shaped in the parietal bone, and W shaped in the vertebra arch. The average fluoride concentrations in the bones increased significantly with each increasing dose and length of fluoride administration. The relative increments were similar between the different regions or the different bones. After stopping fluoride administration, on the other hand, the relative reduction of the average fluoride concentrations in the bones were 30-100%. They were greatly related to the length after stopping fluoride administration and the dose and length of fluoride administration, but also dependent upon the type of bone and the region examined.

Animals↗

PubMed1995

Changes in levels of glutathione and related compounds and activities of glutathione-related enzymes during rat liver regeneration.

The levels of glutathione and glutathione disulfide increased during the regeneration process of rat liver, reaching a maximum (about twice the control value) on day 2 and reverting to the normal level within 5 days. During this regeneration process, changes in the hepatic level of cysteine, glycine and glutamate, the substrates for glutathione synthesis, were determined. The cysteine level in liver increased, reaching a maximum on day 2 and returned to the normal level after 5 days. The levels of glycine and glutamate did not change. The enzyme activities of cystathionine-beta synthase and gamma-cystathionase for cysteine synthesis, and of gamma-glutamylcysteine synthetase, which is a limiting enzyme for glutathione synthesis, were clearly increased in regenerating liver. The increase of glutathione level could be clearly accounted for by the elevation of these enzyme activities.

Animals↗

PubMed1995

[Desensitization immunotherapy on patients with mite-positive bronchial asthma using purified mite feces antigen fractions].

At present, we are performing desensitization immunotherapy on patients with mite-positive bronchial asthma using purified mite feces antigen fractions. We obtained the following results in 13 patients 4 to 12 months after the start of treatment. 1) The fraction showing the maximum reaction in the skin test was HM1-2 (molecular weight: 150-155 kD) in two patients, HM2 (30-40 kD) in nine patients and HM3 (10-20 kD) in two patients, and immunotherapy was performed using these fractions. None of the patients showed the maximum skin reactions or was treated with HM4 (less than 10 kD). 2) In nine patients with perennial asthma, the effects of treatment were excellent in four and moderate in three, while two cases remained unchanged. In four patients with seasonal asthma, the treatment was effective in all cases. 3) In patients in whom immunotherapy was effective, specific IgG antibody increased after the treatment, but it did not increase in the unchanged cases. Among the subclasses of specific IgG antibody increased by the treatment, no clear changes were seen in IgG4 antibody and no conclusion has been reached at present. 4) None of the patients had definite adverse reactions such as the occurrence of asthma attacks or anaphylaxis. These results suggested that purified mite feces antigens are safe and effective. We plan to continue this study with more patients in the future.

Adolescent↗

PubMed1994

Expression of distinct fucosylated oligosaccharides and carbohydrate-mediated adhesion efficiency directed by two different alpha-1,3-fucosyltransferases. Comparison of E- and L-selectin-mediated adhesion.

Among five different human alpha 1 --> 3 fucosyltransferases cloned, fucosyltransferases III (Fuc-TIII) and IV (Fuc-TIV) differ significantly from each other. Fuc-TIII transfers a fucose to both sialylated and nonsialylated N-acetyllactosamine, but Fuc-TIV apparently transfers a fucose only to neutral N-acetyllactosamine. In this study, Chinese hamster ovary (CHO) cells were stably transfected with Fuc-TIII or Fuc-TIV, and the resultant cell lines, CHO-FTIII and CHO-FTIV, were compared for the carbohydrate structures and for their binding to E-selectin or L-selectin. CHO-FTIII and CHO-FTIV cells were labeled metabolically with [3H]galactose, and glycopeptides obtained from these cells were fractionated by serial lectin affinity chromatography. The fractionated glycopeptides were then subjected to various combinations of exoglycosidase treatment or endo-beta-galactosidase digestion. The results obtained can be summarized as follows. CHO-FTIII cells express sialyl Lewisx, Lewisx, and VIM-2 structures, whereas CHO-FTIV cells express only an Lex structure with a small amount of VIM-2 structure. When CHO-FTIII and CHO-FTIV cells were tested for adhesion to E-selectin expressed by tumor necrosis factor-activated endothelial cells and to an E-selectin chimeric protein, only CHO-FTIII cells were found to adhere well to E-selectin. Moreover, both CHO-FTIII and CHO-FTIV cells failed to adhere to an L-selectin chimeric protein. These results clearly indicate that FT-III and FT-IV direct distinctly different fucosylated oligosaccharides. This difference in oligosaccharide structures results in an entirely different efficiency in adhesion to E-selectin. The results also demonstrate that expression of sialyl Lex itself is not sufficient for L-selectin binding.

Animals↗

PubMed1994

Differential E-selectin-dependent adhesion efficiency in sublines of a human colon cancer exhibiting distinct metastatic potentials.

Previously we have shown that high metastatic colonic carcinoma cells express relatively more lamp molecules and sialyl Le(x) structures on the cell surface than their corresponding low metastatic counterparts (Saitoh, O., Wang, W.-L., Lotan, R., and Fukuda, M. (1992) J. Biol. Chem. 267, 5700-5711). In the present study, we extended these findings by testing whether these high and low metastatic colonic carcinoma cells differ in their adhesion efficiency to E-selectin-expressing cells. First, it was found that the high metastatic cells, as compared to their low metastatic counterparts, bind more efficiently to activated human endothelial cells that express E-selectin. This was also true when the adhesion was tested for Chinese hamster ovary cells stably expressing E-selectin. In addition, it was found that the high metastatic cells also adhere more efficiently to mouse endothelioma cells after activation with interleukin-1 beta. It was also shown that the adhesion can be inhibited by soluble lamp-1 or soluble leukosialin that contain sialy Le(x) termini. The inhibition was not, however, observed when these soluble glycoproteins lack sialyl Le(x) structures. The results indicate that the efficiency of the E-selectin-mediated binding of colonic carcinoma cells to human and mouse endothelial cells correlates with the metastatic potential of the cells and suggest that this adhesive event may be one of the critical factors for the metastatic spread of tumor cells. Soluble forms of leukosialin or lamp-1 may be useful as therapeutic agents for the inhibition of E-selectin-mediated binding to tumor cells.

Animals↗

PubMed1994

Fluoride profiles in different surfaces of human permanent molar enamels from a naturally fluoridated and a non-fluoridated area.

The surfaces, sites and depths of fluoride profiles of enamel from a naturally fluoridated area (West Hartlepool, WHP, 1.9 parts/10(6) F in drinking water) and a non-fluoridated area (Leeds, LDS, < 0.1 parts/10(6) F in drinking water) were examined in detail. Using a regression curve, y = ax-b, comparisons of fluoride concentrations were made at depths of 1, 3, 5, 10, 20 and 30 microns at occlusal, middle and cervical sites of buccal, lingual, mesial and distal molar enamel surfaces. In the outermost enamels (< 10 mu in depth), fluoride concentrations tended to be lower in the occlusal sites than in the cervical sites. In interior enamels (> 20 microns in depth), the fluoride concentrations of molars from WHP were statistically higher than those from LDS, while few significant differences were found in the outermost layers. In the mesio- and disto-occlusal sites of approximal surfaces, there were some significant differences in the exponential regression coefficients of fluoride profiles between the two groups of molars.

Dental Enamel↗

PubMed1994

Amino acids and peptides. XXXVII. Synthesis of stereoisomeric nonapeptides corresponding to sequence 41-49 of eglin c and examination of their inhibitory activity against human leukocyte cathepsin G and alpha-chymotrypsin.

A nonapeptide, H-Ser-Pro-Val-Thr-Leu-Asp-Leu-Arg-Tyr-OH, corresponding to sequence 41-49 of eglin c inhibited leukocyte cathepsin G and alpha-chymotrypsin with Ki values of 2.2 x 10(-5) and 7.2 x 10(-6) M, respectively, although eglin c itself inhibited leukocyte elastase, cathepsin G and alpha-chymotrypsin with Ki values of 6.0 x 10(-9), 5.5 x 10(-9) and 2.5 x 10(-9) M, respectively. The inhibitory activity of the nonapeptide decreased following incubation with cathepsin G due to the cleavage of the Leu45-Asp46 peptide bond. Therefore, Leu45 and/or Asp46 were replaced with D-amino acids and the inhibitory activities of the resultant nonapeptides were examined. Their inhibitory activities against cathepsin G and alpha-chymotrypsin were much weaker than those of the all-L-type nonapeptide, suggesting that the amino acids at the active site, Leu45 and Asp46 are required to be in the L-configuration for potent activity.

Amino Acid Sequence↗

PubMed1994

Differences in protein structure and similarities in catalytic function of two L-stereoselective carbonyl reductases from bakers' yeast.

We purified and studied two L-stereoselective carbonyl reductases from bakers' yeast (Saccharomyces cerevisiae). One catalyzed exclusively the enantioselective reduction of carbonyl compounds such as beta-keto esters and the other acted on alpha-acetoxy ketones and beta-keto esters. The enzymes had identical molecular weights and catalyzed the L-stereoselective reduction of various carbonyl compounds with similar substrate specificity, but they were different proteins coded by different genes.

Alcohol Oxidoreductases↗

PubMed1994

Low-molecular-size allergens, LM-1s, in feces extract of Dermatophagoides farinae which elicit histamine release from washed blood cells of patients with bronchial asthma.

A significant activity to elicit histamine release was found in an ultrafilterable (Mr-cutoff < 10 kD) fraction of a mite feces extract from a spent mite medium. The activity was divided into two fractions (LM1 and LM2) on an Ultrogel AcA 54 column when monitored by histamine release assay using washed peripheral blood cells from mite-allergic patients. The larger-molecular-size antigen, LM1 was further separated into three allergenic fractions by consecutive chromatography on Sephadex G-50, GM-DEAE, and TSKgel ODS-120T. All the purified allergens, LM1s (LM1a, LM1b, and LM1c), which produced positive reactions in skin tests on allergic patients, were glycoproteins (molecular weight: 8 kD on SDS-PAGE; 12.5 kD on Gel filtration) with different carbohydrate contents and pI values ranging from 4 to 5 on an IEF plate. LM1s were cross-reactive with anti-Der f II but not with anti-Der f I. The reaction of LM1c to anti-Der f II serum was completely inhibited with the other antigens, while the reactions of LM1a and LM1b to the serum were partially inhibited with LM1c or the other antigens, respectively.

Allergens↗

PubMed1994

A low-molecular-size LM2 allergen in mite, Dermatophagoides farinae, extracts containing feces which elicits conjunctival reactions in patients with bronchial asthma.

A low-molecular-size antigen, LM2, which possesses a significant activity to elicit histamine release, was isolated from an ultrafilterable (Mr-cutoff < 10k) fraction of a mite extract containing feces by consecutive chromatography on Ultrogel AcA 54 and Sephadex G-25. The isolated antigen was still heterogeneous glycoproteins, giving a smeary band around pI 3-5 on an Ampholine PAG plate, with molecular weights of 6-8k and 4k on SDS-PAGE and Sephadex G-50 gel filtration, respectively. Chemical treatments of the antigen suggested that the epitope responsible for the elicitation of histamine release resided in the protein moiety. The antigen had activities to provoke conjunctival congestion and histamine release in mite-allergic patients, but not immunogenicity by itself. The antigen competitively inhibited the reactions of HM1, HM2, and HM3 fractions to corresponding antisera, but did not cross-react with anti-Der f I or anti-Der f II sera.

Allergens↗

PubMed1994

[Refractive changes following cataract extraction with intraocular lens implantation].

Refractive changes after six cataract surgery procedures with intraocular lens implantation were assessed; namely, (1) 11 mm incision with running suture closure (n = 25 eyes), (2) 6.5 mm incision with running suture closure (n = 23). (3) 6.5 mm incision with horizontal suture closure (n = 21), (4) 6.5 mm incision without suture closure (n = 40), (5) 5.5 mm incision without suture closure (n = 46), and (6) 3.2 mm incision without suture closure (n = 46). The axis-based keratometric cylinder, absolute value of the induced cylinder vector, and spherical equivalent of the refractive power were analyzed up to 6 months after surgery. Refractive changes did not reach a stable state at 3 months postoperatively for the 11 mm and 6.5 mm running suture closure groups, but they stabilized at 3 months postoperatively for the 6.5 mm horizontal suture and sutureless closure groups, at 1 month postoperatively for the 5.5 mm sutureless closure group, and at 2 weeks postoperatively for the 3.2 mm sutureless closure group. It is concluded that the timing of refractive stabilization differs significantly among procedures, and smaller incisions with wound closure methods which do not exert a vertical force cause fewer postoperative refractive changes.

Aged↗

PubMed1993

Immortalization of epithelial-like cells from human liver tissue with SV40 T-antigen gene.

The cells derived from the human embryo liver tissue were transfected with a plasmid pSV3neo containing both the large and small T-antigen gene of the early region of simian virus 40 (SV40), and two cell strains, OUMS-21 and -22, were obtained. OUMS-22 cells, to date, have reached over 100 population doublings through a culture crisis and are considered to have become an immortal cell line. However, OUMS-21 cells failed to become an immortal cell line. Both OUMS-21 and -22 cells were SV40 T-antigen-positive, epithelial-like, and immunoreactive against an anti-keratin 18 monoclonal antibody but against neither an anti-vimentin nor an anti-von Willebrandt factor VIII monoclonal antibody. The staining pattern of cytokeratin in these cells was similar to that in the differentiated human hepatoblastoma and hepatocellular carcinoma cell lines but not to that in the human cholangiocellular carcinoma cell lines. OUMS-21 and -22 cells expressed neither alpha-fetoprotein nor albumin mRNAs. These cells showed no tyrosine aminotransferase activity. However, both OUMS-21 and -22 cells were sensitive to cytotoxicity of aflatoxin B1, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole, and benzo[a]pyrene, whereas human embryo lung fibroblasts were insensitive to the cytotoxicity of these carcinogens. These findings suggest that OUMS-21 and -22 cells may arise from undifferentiated liver stem cells or from hepatocytes that lost their ability to express the liver-specific functions prior to immortalization. Both OUMS-21 and -22 cells expressed glutathione S-transferase pi (GST-pi) mRNA. The expression of GST-pi mRNA highly increased in OUMS-22 cells with their immortalization. Karyotypic analysis showed that numerical and structural aberrations of the chromosomes were profound, but neither specific events nor marker chromosomes were found in OUMS-21 and -22 cells. Both OUMS-21 and -22 cells could grow in soft agar, but they were not tumorigenic when transplanted into nude mice.

Aflatoxin B1↗

PubMed1992

Crystal structure of papain-succinyl-Gln-Val-Val-Ala-Ala-p-nitroanilide complex at 1.7-A resolution: noncovalent binding mode of a common sequence of endogenous thiol protease inhibitors.

Succinyl-Gln-Val-Val-Ala-Ala-p-nitroanilide corresponding to a common sequence of endogenous thiol protease inhibitors is a noncompetitive reversible inhibitor of papain. In order to elucidate the binding mode of the inhibitor at the atomic level, its complex with papain was crystallized at ca. pH 7.0 using the hanging drop method, and the crystal structure was analyzed at 1.7-A resolution. The crystal has space group P2(1)2(1)2(1), with a = 43.09, b = 102.32, c = 49.69 A, and Z = 4. A total of 47,215 observed reflections were collected on the imaging plates using the same single crystal, and 19,833 unique reflections with Fo > sigma (Fo) were used for structure determination and refinement. The papain structure was determined by use of the atomic coordinates of papain previously reported, and then refined by the X-PLOR program. The inhibitor molecule was located on a difference Fourier map and fitted into the electron density with the aid of computer graphics. The complex structure was finally refined to R = 19.6% including 118 solvent molecules. The X-ray analysis of the complex crystal shows that the inhibitor is located at the R-domain side, not in the center of the binding site created by the R- and L-domains of papain. Such a binding mode of the inhibitor explains well the biological behavior that the inhibitor exhibits against papain. Comparison with the structure of papain-stefin B complex indicates that the structure of the Gln-Val-Val-Ala-Gly sequence itself is not necessarily the essential requisite for inhibitory activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1992

Sensitive determination of cystathionine and assays for cystathionine beta- and gamma-lyase, as well as cystathionine beta-synthase, using high-performance liquid chromatography.

Cystathionine was cleaved into 2-ketobutyric acid, cysteine and ammonia by cystathionase. 2-Ketobutyric acid was converted into 3-ethyl-2-hydroxy-6,7-dimethoxyquinoxaline (EHDQ) by reaction with 1,2-diamino-4,5-dimethoxybenzene. When EHDQ was measured in a mobile phase of pH 2.1 using high-performance liquid chromatography with ultraviolet detection, 250 pmol of L-cystathionine in 250 microliters of the reaction mixture could be determined. Because EHDQ has a strong fluorescence in a mobile phase of pH 6.5 at 447 nm, on excitation at 365 nm, as little as 2.5 pmol of cystathionine in 250 microliters of the reaction mixture could be determined by high-performance liquid chromatography with fluorimetric detection. Cystathionase activity was assayed on the basis of the same principle by determining cystathionine in as little as 63 ng of rat liver by fluorimetric detection. Cystathionine beta-synthase activity was measured by the same method by determining cystathionine formed in only 113 ng of wet weight of rat liver. Using these methods, both cystathionine beta- and gamma-lyase activities in Saccharomyces cerevisiae were determined, because quinoxaline derivatives from pyruvate and 2-ketobutyrate could be measured simultaneously by high-performance liquid chromatography.

Animals↗

PubMed1992

Effect of continuous circular capsulorhexis and intraocular lens fixation on the blood-aqueous barrier.

Permeability across the blood-aqueous barrier to fluorescein was estimated fluorophotometrically in pseudophakic eyes for which a continuous circular capsulorhexis was performed. The permeability index was significantly higher in the in-the-bag fixations than in the out-of-the-bag fixations at 3 and 6 months after surgery, when only bilateral cases were enrolled. Damage to the barrier in eyes with the in-the-bag fixations was attributable to the broad attachment of optics to the anterior capsule, because a positive linear regression was obtained between the permeability index and the doughnut-shaped contact area in eyes with the in-the-bag fixations. The permeability index in eyes with the in-the-bag fixations was as low as that in eyes with the out-of-the-bag fixations when the contact area was small. These results indicate an unfavorable effect of the in-the-bag fixation with broad contact of the optics with the anterior capsule and, thus, suggest either an in-the-bag fixation with a large capsulorhexis or an out-of-the-bag fixation.

Adult↗

PubMed1992

Effects of barbiturates with or without liver-tumor-promoting activity on survival and DNA synthesis of suckling and adult rat hepatocytes in serum-free primary culture.

The effects of four barbiturates with or without liver-tumor-promoting activity were examined on survival and deoxyribonucleic acid (DNA) synthesis of suckling and adult rat hepatocytes in serum-free primary culture: Of the four barbiturates, two promoters, phenobarbital and barbital, enhanced DNA synthesis of suckling rat hepatocytes at low concentrations of 0.5-2 mM or 0.5 mM, but suppressed it at high concentrations of 3 mM or 1.5-4 mM. DNA synthesis of adult rat hepatocytes was, however, only suppressed by phenobarbital within the dose range tested of 1-3 mM. On the other hand, two remaining non-promoters, barbituric acid and amobarbital, did not increase but only suppressed DNA synthesis of suckling rat hepatocytes within the dose ranges of 0.5-4 mM and 0.05-0.5 mM respectively. Phenobarbital and amobarbital were effective for supporting survival and maintaining morphological features of suckling and adult rat hepatocytes at the relatively high concentrations of 3-4 mM and 0.5-0.75 mM respectively. However, barbital and barbituric acid were ineffective for maintenance of hepatocytes. The results show that the ability to support survival of primary cultured hepatocytes is not a common property of liver-tumor-promoter barbiturates but is a common property of some barbiturates with high lipophilicity, and that the maintenance of hepatocytes by phenobarbital or amobarbital is not due to a counterbalance of stimulated proliferation and death of the cells.

Aging↗

PubMed1992

Increase of S-(1,2-dicarboxyethyl)glutathione in regenerating rat liver.

Changes in the level of S-(1,2-dicarboxyethyl)glutathione (DCE-GS) with time were determined during regeneration of rat liver after partial hepatectomy. The DCE-GS level increased in regenerating rat liver, reaching a maximum level (4.7-fold) on day 2 and reverted to the normal level in 1 week. During the liver regeneration process, the activity of DCE-GS-synthesizing enzyme in the liver retained its control level, while the level of glutathione--a substrate of the enzyme in the liver--fluctuated in parallel with that of DCE-GS.

Animals↗

PubMed1992

Human hepatocyte growth factor stimulates the growth of HUH-6 clone 5 human hepatoblastoma cells.

The effects of human hepatocyte growth factor (hHGF), a potent mitogen for rat and human hepatocytes in primary culture, on proliferation of human hepatoma and hepatoblastoma cells were examined. Out of five cell lines; HLE, HuH-6 clone 5, HuH-7, PLC/PRF/5, and Hep G2, only HuH-6 Clone 5 cells were stimulated by recombinant hHGF. Both native and recombinant hHGFs caused dose-dependent increases in cell number and DNA synthesis of cells. This stimulation was strongly inhibited by anti-hHGF monoclonal antibody.

Animals↗