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Biomedical subjects

Y Seyama

Publications and source records attributed to Y Seyama.

At least 127 records · Page 7Linked to original sources
PubMed1990

Composition of long-chain bases in sphingomyelin of the guinea pig Harderian gland.

Sphingomyelin from the guinea pig Harderian gland was isolated and characterized. The purified sphingomyelin gave a broad spot on thin-layer chromatography. The fatty acid composition of the whole sphingomyelin was 71% nonhydroxy acids and 29% 2-hydroxy acids. Methyl-branched fatty acids were only 2% of the total acids. The long-chain bases were composed of straight-chain sphingenines (50%) and sphinganines (6%). Methyl-branched long-chain bases were 44% of the bases. The sphingomyelin was further separated into four fractions (I, II, III, IV) by high-performance liquid chromatography. The ratio of fractions I, II, III, and IV was approximately 2:5:2:1, respectively. The fatty acids of fractions I and II consisted of nonhydroxy acids and those of fractions III and IV were 2-hydroxy acids. The long-chain bases of fractions I and III were sphinganines including 10-, 9-, and 8-methylsphinganines and anteiso-sphinganines. These methyl-branched bases occupied about 70% of the total sphinganines. The long-chain bases of fractions II and IV consisted of sphingenines. The methyl-branched unsaturated bases were only 30% of the total sphingenines, all in the anteiso-form. Thus, the sphingomyelin obtained from guinea pig Harderian gland had complex compositions of fatty acids and long-chain bases, and half the number of long-chain bases had methyl branches. The methyl-branched fatty acids were only a minor component. These characteristics are similar to those of cerebrosides isolated from the same source.

Animals↗
PubMed1990

[Effects of the anti-ulcer agents on the amine contents and regulating enzyme activity of gastric mucosa during the healing process of acetic acid induced gastric ulcer in rats].

In order to elucidate the action of an H2 blocker (cimetidine) and gastric mucosal protection agents (sucralfate and sofalcone) on the relapse and recurrence of gastric ulcer, the effects of cimetidine, sucralfate and sofalcone on the contents of histamine and serotonin and histidine decarboxylase (HDC) activity in the gastric mucosa were examined in the ulcer region and the intact region at the 10th day after the operation to produce acetic acid-induced gastric ulcer in rats. The following results were obtained: 1) HDC activity in the gastric mucosa of rats treated with cimetidine (100 mg/kg twice daily) tended to increase in the intact region, and it was significantly increased in the ulcer region. 2) Increased HDC activity due to cimetidine treatment was observed at the 10th day after interruption of cimetidine administration. 3) The HDC activity in the gastric mucosa was not changed by the treatment with sucralfate (500 mg/kg/day) and sofalcone (200 mg/kg/day). The results suggest that the increased HDC activity in the gastric mucosa might participate in the relapse and recurrence of gastric ulcer after discontinuation of cimetidine administration.

Acetates↗
PubMed1990

[Study of the microwave fixation using the immunohistochemical method].

We evaluated microwave fixation of formalin immersed and microwave irradiated kidney and liver tissues by keratin immunostain using the paraffin embedded section. In the microwave irradiated tissues, the formalin fixed areas and the alcohol fixed areas were clearly detected because; 1) Alcohol fixed tissues are easily digested by pepsinization but formalin fixed tissues are not, and 2) Formalin fixed tissues revealed intense keratin staining after pepsinization, whereas intense keratin staining were noted in alcohol fixed tissues without preliminary pepsinization. The microwave irradiation fixed completely the tissues in about 20 sec. A thin, formalin-fixed layer was observed in the periphery of the tissues. A thin, alcohol-fixed layer was observed beneath the peripheral formalin fixed layer in some specimens. This area was thought to have been fixed during the dehydration of the paraffin blocks. However with microwave fixation, most of the central areas differed from those of the formalin or alcohol fixed areas. They showed sharp contrast on hematoxylin-eosin staining, conspicuous cellular membrane and only slight cellular shrinkage and swelling. Unlike the formalin and alcohol fixed tissues, the intensity of keratin staining was independent of preliminary pepsinization and keratin staining was observed. Therefore, the central areas were thought to be fixed by the microwave irradiation. This fixation method seems to be useful for immunostaining.

Alcohols↗
PubMed1990

Biosynthesis and functions of leukotriene C4.

1. The LTC4 synthase activity is rich in the microsomal fraction of the guinea pig spleen and lung. The enzyme was partially purified from the guinea pig lung and separated from the microsomal glutathione S-transferase (GST), by column chromatograpy. The enzyme has a specific activity of 40 nmol/min.mg, and acts preferentially on 5, 6-LTA4. Various types of cytosolic GSTs utilize all types of LTA4 isomers (5,6-, 11,12- and 14,15-LTA4) almost to the same extent, and methyl ester forms are better substrates for GST. 2. Two different types of GSTs (Yn1n1 and P) were purified from rat brain cytosol, to homogeneity. Because both types have a high LTC4 synthase activity, they may participate in the LTC4 production in the rat brain. 3. LTC4, produced in the guinea pig atrium, stimulates pertussis toxin (IAP)-sensitive muscarinic K+ channel (IK.ACh). The negative chronotropic action of alpha 1-adrenergic agonist might relate to the production of arachidonate lipoxygenase metabolites. These results together with the findings in Aplysia sensory neurons, suggest a novel mode of eicosanoid actions.

Animals↗
PubMed1990

Prostaglandin E2 evokes intracellular calcium rise in mouse osteoblastic cell line, MC3T3E-1.

The effect of prostaglandin (PG) E2 on the intracellular free calcium concentration ([Ca2+]i) in mouse osteoblastic clone, MC3T3E-1 cells, was investigated with the fluorescent probe fura-2. In the growing phase of MC3T3E-1, PGE2 evoked a transient rise of [Ca2+]i in a dose-dependent manner in the range from 0.28 microM to 5.6 microM of PGE2. The elevation of [Ca2+]i induced by PGE2 was not affected by the presence of EGTA. Moreover, PGE2 (5.6 microM) evoked an acute accumulation of intracellular inositol-1,4,5-trisphosphate (InsP3). The absolute content of InsP3 increased from 5.8 pmole/10(6) cells to 19.2 pmole/10(6) cells within 30 sec after the PGE2 treatment. The degree of [Ca2+]i elevation induced by PGE2 was dependent on the day after subculturing. At day 3, the response to PGE2 was the most pronounced during the 8-day experimental periods. These data suggest that PGE2 evoked a [Ca2+]i rise via a phospholipase C-InsP3 pathway in osteoblasts, particularly in the growing phase.

Animals↗
PubMed1989

Purification and characterization of a growth factor from guinea pig harderian gland.

A polypeptide growth factor, Harderian gland-derived growth factor (HGDGF), has been purified approximately 43,000-fold from guinea pig Harderian gland by column chromatography on TSK gel DEAE-5PW, blue-Sepharose CL-6B, and Superose 12. The yield was approximately 10%. The Superose 12 fraction was further purified by Aquapore BU-300 reversed-phase chromatography to apparent homogeneity. HGDGF was eluted from TSK gel DEAE-5PW at 0.20-0.35 M NaCl, with a linear gradient of 0.15-0.80 M NaCl and at 2.20 M NaCl from blue-Sepharose CL-6B. The activity of HGDGF toward human embryonic cells (TIG-3) was quantitated, [3H]thymidine incorporation for 48 h being stimulated in a linear and dose-dependent manner. Purified HGDGF has a molecular weight of approximately 13,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and molecular sieve column chromatography. HGDGF is labile to treatment with SH reagents or acetic acid. Both trypsin digestion and boiling decrease the activity of HGDGF. Its pI is 5.1. HGDGF stimulates the multiplication of TIG-3 cells but has no effect on human endothelial cells K2T1 or A2T2 which require fibroblast growth factor for growth. HGDGF appears to differ from other growth factors, suggesting that it is a previously undescribed growth factor.

Animals↗
PubMed1989

Enzymic conversion of 11,12-leukotriene A4 to 11,12-dihydroxy-5,14-cis-7,9-trans-eicosatetraenoic acid. Purification of an epoxide hydrolase from the guinea pig liver cytosol.

(11S,12S)-Epoxy-5,14-cis-7,9-trans-eicosatetraenoic acid (11,12-leukotriene A4) was nonenzymically converted to seven compounds: two diastereomers of (12S)-hydroxyeicosatetraeno-delta-lactones (major products), two diastereomers of (5,12S)-dihydroxyeicosatetraenoic acid and three stereoisomers of (11,12S)-dihydroxyeicosatetraenoic acid. Among these compounds, (11R,12S)-dihydroxy-5,14-cis-7,9-trans-eicosatetraenoic acid proved to be the only enzymic product. This hydrolysis activity was present in the cytosol fractions of various tissues of guinea pig such as liver, adrenal gland, small intestine, and brain. We purified the epoxide hydrolase to an apparent homogeneity from the guinea pig liver. The enzyme had a molecular weight of 60,000 as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and an isoelectric point of 7.3. The partial amino acid sequence was different from that of the microsomal enzyme. Km and Vmax values for 11,12-leukotriene A4 were 18 microM and 2.4 mumol/min/mg protein, respectively. These results indicate that 11,12-dihydroxyeicosatetraenoic acid is enzymically synthesized from 11,12-leukotriene A4 by the action of the cytosolic epoxide hydrolase in vitro.

Animals↗
PubMed1989

Leukotriene A4 hydrolase from guinea pig lung: the presence of two catalytically active forms.

Leukotriene A4 hydrolase was purified to apparent homogeneity from the guinea pig lung. The molecular weight was determined to be 70 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme exhibited two active forms with different pI values (5.7 and 5.4) depending on the presence or absence of SH-reducing reagents during purification procedures. No significant differences were observed between both forms of the enzyme as regards the catalytic properties. The N-terminal 20 amino acid sequence (PEVVDTXSLASPATVXRTKH) showed a 90% identity to the human enzyme with a constitutive substitution of Ile-3 and Ser-14 (human) by Val-3 and Thr-14 (guinea pig), respectively.

Amino Acids↗
PubMed1989

Purification of human lung angiotensin-converting enzyme by high-performance liquid chromatography: properties and N-terminal amino acid sequence.

Angiotensin-converting enzyme from the human lung was purified to apparent homogeneity, using high-performance liquid chromatography following trypsin treatment of the detergent-extract. A 1,750-fold purification was achieved with a 26% yield. The specific activity of the enzyme was 105 units per mg protein with the substrate hippuryl-L-histidyl-L-leucine (HHL) at 37 degrees C, and the Km value for HHL was 1.9 mM. The molecular weight was estimated to be 170,000 by sodium dodecyl sulfate gel electrophoresis, and the isoelectric point was about 4.8, by chromatofocusing. The N-terminal amino acid sequence was (NH2)-X-X-Pro-Gly-Leu-Glu-Pro-Gly-X-Phe-Ser-Ala-Arg-Glu-Ala-Gly-Ala. This is highly homologous to the corresponding sequences of the enzymes from bovine and rabbit lung and from pig, bovine, and mouse kidney, but significantly different from that of the human kidney enzyme.

Amino Acid Sequence↗
PubMed1989

[Changes in amine contents and regulating enzyme activity of gastric mucosa during the healing process of acetic acid induced ulcer in rats].

Changes in the contents of various amines and histidine decarboxylase (HDC) activity in the gastric mucosa during the healing process of acetic acid induced gastric ulcer in rats were sequentially examined in the ulcer region and intact region at 2, 10, 40, 80, 180 and 365 days after the operation. The following results were obtained: 1) Histamine (HA) content in the ulcer region was decreased as compared with the intact region at 2 and 10 days and returned to the control level in 40 days. After 180 days, the contents in the ulcer region and intact region were also increased as compared with that of the normal control region. 2) Changes in serotonin (5-HT) content as well as HA content were observed. 3) Norepinephrine content in the ulcer region was decreased as compared with the intact region at 2, 10, 80 and 180 days. 4) HDC activity in the ulcer region was decreased as compared with the intact region at 2, 10 and 40 days, and a lower level was maintained still at 180 days. 5) In the relapse and recurrence of gastric ulcer at 365 days, HA and 5-HT contents in the intact region and ulcer region were not different from those in healing rats, but the contents of these amines were higher at 180 days. The results suggest that the change of HA, 5-HT contents and HDC activity in the gastric mucosa may be one of the factors involved in the relapse and recurrence of chronic ulcers.

Acetates↗
PubMed1989

[Comparative effects of traditional Chinese medicines (dai-saiko-to, hatimi-zio-gan and byakko-ka-ninzin-to) on experimental diabetes and hyperlipidemia].

In order to clarify the anti-hyperglycemic and anti-hyperlipidemic actions of traditional Chinese medicines, experiments were carried out with experimentally diabetic rats induced by cyproheptadine treatment. The following results were obtained: 1) Dai-saiko-to decreased the blood glucose level at 30, 60 and 120 min after glucose loading in the tolerance test; and the drug tended to increase serum insulin level and increased the ratio (glucose/insulin) at 120 min after the glucose loading. A significant decrease in serum total cholesterol was observed in the high fat diet-treated rats. 2) Hatimi-zio-gan also decreased the blood glucose level at 30 and 120 min after glucose loading. The drug did not lower the serum insulin, but rather increased the glucagon level at 120 min after the loading. The serum lipid level was not reduced by the drug. 3) Byakko-ka-ninzin-to also increased serum glucagon at 120 min, but no change of glucose and lipid in the sera under the above experimental conditions was observed. Such experimental results suggest that Dai-saiko-to might exert an effect that can improve the pathological conditions of diabetes mellitus.

Animals↗
PubMed1989

Molecular cloning and expression of human leukotriene A4 hydrolase cDNA.

We have isolated a near full-length cDNA encoding human leukotriene A4 (LTA4) hydrolase from a human spleen cDNA library. The mature form of the enzyme consists of 610 amino acid residues and its molecular weight is calculated to be 69,153. The cDNA was inserted into an expression vector (pUC 9) and expressed in Escherichia coli. The fusion protein (EX85) possessed the LTA4 hydrolase activity with kinetic properties similar to the native enzyme.

Amino Acid Sequence↗
PubMed1989

11,12-leukotriene A4: synthesis and metabolism.

11(S),12(S)-oxido-5Z,7E,9E,14Z-eicosatetraenoic acid (11,12-LTA4) was chemically synthesized from 12-hydroperoxy-eicosatetraenoic acid (12-HPETE). 11,12-LTA4 methyl ester was nonenzymically hydrolyzed to at least four products. These products were identified to be epimers of 5,12(S)-dihydroxy-eicosatetraenoic acid methyl ester (5,12(S)-diHETE-Me) and epimers of 11,12-diHETE-Me. In addition to the above nonenzymic products, 11,12-LTA4 was converted to 11,12-LTC4 by the rat liver glutathione S-transferase, and to an isomer of 11,12-diHETE by homogenates of the guinea pig adrenal gland and liver.

Adrenal Glands↗
PubMed1988

Synthesis and structural identification of four dihydroxy acids and 11,12-leukotriene C4 derived from 11,12-leukotriene A4.

Using a partially purified 12-lipoxygenase from porcine leukocytes, (5Z,8Z,10E,14Z)-12-hydroperoxy-5,8,10,14-icosate traenoic acid was synthesized from arachidonic acid with a yield of over 35%. The absolute configuration of C-12 was determined as S by chiral-phase column chromatography. It was chemically converted to at least three epoxides with the conjugated triene structure. Two were identified by proton NMR and mass spectrometry to be (5Z,7E,9E,14Z)-(11S,12S)-11,12-oxido-5,7,9,14-ic osatetraenoic acid (11,12-leukotriene A4) and (5Z,7Z,9E,14Z)-(11S,12S)-11,12-oxido-5,7,9,14-ic osatetraenoic acid (7-cis-11,12-leukotriene A4). 11,12-Leukotriene A4 underwent acid hydrolysis to yield two diastereomers of (6E,8E,10E,14Z)-(12S)-5,12-dihydroxy-6,8,10,14-i cosatetraenoic acid and two isomers of (14Z)-(12S)-11,12-dihydroxy-5,7,9,14-icosatetraenoic acid. Upon incubation with rat liver glutathione S-transferase, 11,12-leukotriene A4 was converted to 11,12-leukotriene C4, a spasmogenic compound.

Animals↗