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Biomedical subjects

H Kuroda

Publications and source records attributed to H Kuroda.

At least 253 records · Page 14Linked to original sources
PubMed1989

Partial purification, and some properties and reactivities of cetraxate benzyl ester hydrochloride-hydrolyzing enzyme.

Debenzylating enzyme from Aspergillus niger enzyme (commercial crude cellulase) catalyzes the hydrolysis of cetraxate benzyl ester hydrochloride (2), a precursor of the antiulcer agent (1). The enzyme was highly purified by three kinds of chromatographies (hydrophobic, ion exchange, gel filtration) with a recovery of 36%. The content of the debenzylating enzyme was about 0.1% in the crude cellulase, but the enzyme showed no cellulase activity. The purified enzyme was inactivated by Hg2+, and diisopropyl phosphorofluoridate (DFP). It was a monomer with a molecular weight of about 35,000, and its isoelectric point was estimated to be 5.3. It showed a debenzylating activity for the phenylpropionic acid benzyl ester moiety of various benzyl ester derivatives, and the benzyl ester of phenylalanine or that of tyrosine was also well hydrolyzed.

Aspergillus niger↗
PubMed1989

[The synthesis of 4'-(2-carboxyethyl)phenyl trans-4-aminomethyl cyclohexane carboxylate hydrochloride (cetraxate hydrochloride) by means of enzymatic debenzylation].

Cetraxate hydrochloride (1) (antiulcer agent) has been industrially produced by the chemical protective method of p-hydroxy propionic acid derivatives. Screening of enzymes which quantitatively hydrolyzed cetraxate benzyl ester hydrochloride (2) into 1 was undertaken to establish a novel enzymatic method of production of 1. It was found that the enzyme activity for debenzylation of 2 is contained in cellulase enzymes originated from Aspergillus sp. Lower alkyl groups or phenyl groups of p-hydroxy propionic acid derivatives are likewise selectively hydrolyzed by the cellulase enzyme. This enzymatic synthetic method is very useful for the industrial preparation of 1.

Anti-Ulcer Agents↗
PubMed1989

[Ultrastructural study of interlobular bile ductal disorganization in autoimmune liver diseases].

Ultrastructures of interlobular bile ductules were examined in 7 cases of lupoid hepatitis and related disease (LH) and 8 cases of primary biliary cirrhosis (PBC). Mononuclear cell, especially lymphocyte infiltrations into bile ducts cross basement membrane were common findings found in 57.1% of LH and in 50% of PBC patients, thus, statistically, the occurrence of lymphocyte infiltration was similar for both groups. Cell contact between bile duct epithelia and infiltrated cells differed in LH and PBC. In LH, 86.7% of the cells contacted at small point, but 92.3% PBC cells had broad contact with each other. Stratification of bile duct epithelia and other visible changes in PBC epithelial cells was statistically more extensive compared with LH. Dilatation of intercellular space was often observed in basal region of LH. Destruction and degeneration of LH and PBC bile duct epithelial cells was mainly observed in basal and luminal regions, respectively. Rupture and thickening of basement membrane was seen in PBC, but rarely in LH. The average diameter of interlobular bile ducts were larger and oval shaped in patients with PBC compared to smaller circular ducts observed in LH patients. These results not only revealed that several changes in interlobular bile ducts occur but that similar changes with LH and PBC. This suggests that both distinctions and similarities exist between LH and PBC as autoimmune hepatic disease.

Adult↗
PubMed1989

Suppression of the activities of T-lymphocyte-related enzymes in spleen by administration of an immunosuppressant, 15-deoxyspergualin.

We investigated the effects of both (-) and (+)-enantiomers of 15-deoxyspergualin (DSG), an immunosuppressant, on enzyme networks in spleens of ICR mice. Of the 14 hydrolytic enzymes tested, the activity of dipeptidyl aminopeptidase IV (DAP-IV) was found to be significantly suppressed by the administration of (-)-DSG but not by that of (+)-DSG. In contrast, the activity of N-acetyl-beta-D-glucosaminidase (GlcNAc-ase) was suppressed by the administration of both enantiomers of DSG. Judging from the previous reports of ours and others which suggested specific relations of these enzymes to T-lymphocytes, the above findings may warrant further studies of these enzymes in relation to immunologic functions of the body.

Acetylglucosaminidase↗
PubMed1988

[Effects of phorbol esters and various growth factors on prostaglandin E2 synthesis by cultured porcine thyroid cells].

The effects of phorbol esters and various growth factors on prostaglandin E2 (PGE2) synthesis by cultured porcine thyroid cells were examined. Phorbol 12-myristate, 13-acetate (PMA), phorbol 12, 13-dibutylate, epidermal growth factor (EGF) and fetal bovine serum (FBS) stimulated PGE2 production in a dose-related fashion. PMA stimulated PGE2 production over fifty-fold with a dose of 10(-7) M compared with controls during 4h incubation. EGF (10(-7) M) also stimulated it about ten-fold. The ED50 values of PMA and EGF were around 1 X 10(-9) M and 5 X 10(-10) M respectively. FBS also clearly stimulated it about ten-fold with a concentration of 20%. Thyroid stimulating hormone (TSH) and inactive phorbol, however, did not stimulate PGE2 production. The release of radioactivity from [3H]-arachidonic acid (A.A.) prelabeled cells was also stimulated by PMA, EGF and FBS. These results indicate that PMA, EGF and FBS are potent stimulators of PGE2 production, associated with the activity to stimulate A.A. release in thyroid cells. Secondly, in order to elucidate some mechanism(s) of the stimulation by these factors of PGE2 production, effects of various agents such as EGTA, Ca2+-ionophore (A-23187), cycloheximide, hydrocortisone and para-bromophenacylbromide (p-BPB) were examined on PGE2 production and/or [3H]-A.A. release by thyroid cells. Chelation of Ca2+ by EGTA, treatment of cells with hydrocortisone, an inducer of lipocortin which may inhibit phospholipase A2 (PLA2) activity, or inhibition of PLA2 activity by p-BPB clearly inhibited PGE2 production and/or [3H]-A.A. release induced by PMA, EGF and FBS. Cycloheximide also blocked the PMA-or EGF-stimulated [3H]-A.A. release and PGE2 synthesis. On the other hand, Ca2+-ionophore (A-23187) potentiated PMA- or EGF-stimulation. Furthermore, the Ca2+-ionophore alone-induced stimulation was clearly inhibited by the treatment with hydrocortisone or p-BPB. These data indicate that an increase of Ca2+ in cytosol is important to PLA2 activation, and also that PLA2 may be activated by PMA, EGF and FBS in thyroid cells. The inhibition of PMA- or EGF-induced [3H]-A.A. release and PGE2 synthesis by cycloheximide suggests that both factors-induced stimulation may be sensitive to regulation by short-lived protein(s). Finally, other growth factors such as insulin (0.1-10 micrograms/ml), insulin like growth factor I (IGF-1) (10(-9)-10(-7) M) and interleukin 1 alpha (1-100 ng/ml) also stimulated PGE2 production by thyroid cells in the presence or absence of EGF (10(-10)-10(-9) M).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Reticular meshwork of the spleen in rats studied by electron microscopy.

The reticular meshwork of the rat spleen, which consists of both fibrous and cellular reticula, was investigated by transmission electron microscopy. The fibrous reticulum of the splenic pulp is composed of reticular fibers and basement membranes of the sinuses. These reticular fibers and basement membranes are continuous with each other. The reticular fibers are enfolded by reticular cells and are composed of two basic elements: 1) peripheral basal laminae of the reticular cells, and 2) central connective tissue spaces in which microfibrils, collagenous fibrils, elastic fibers, and unmyelinated adrenergic nerve fibers are present. The basement membranes of the sinuses are sandwiched between reticular cells and sinus endothelial cells and are composed of lamina-densalike material, microfibrils, collagenous fibrils, and elastic fibers. The presence of these connective tissue fibrous components indicates that there are connective tissue spaces in these basement membranes. The basement membrane is divided into three parts: the basal lamina of the reticular cell, the connective tissue space, and the basal lamina of the sinus endothelial cell. When the connective tissue space is very small or absent, the two basal laminae may fuse to form a single, thick basement membrane of the splenic sinus wall. The fibrous reticulum having these structures is responsible for support (collagenous fibrils) and rebounding (elastic fibers). The cells of the cellular reticulum--reticular cells and their cytoplasmic processes, which possess abundant contractile microfilaments, dense bodies, hemidesmosomes, basal laminae, and a well-developed, rough-surfaced endoplasmic reticulum, and Golgi complexes, which are characteristic of both fibroblasts and smooth muscle cells--are considered to be myofibroblasts. They may play roles in splenic contraction and in fibrogenesis of the fibrous reticulum. The contractile ability may be influenced by the unmyelinated adrenergic nerve fibers that pass through the reticular fibers. The three-dimensional reticular meshwork of the spleen consists of sustentacular fibrous reticulum and contractile myofibroblastic cellular reticulum. This meshwork not only supports the organ but also contributes to a contractile mechanism in circulation regulation, in collaboration with major contractile elements in the capsulo-trabecular system.

Animals↗
PubMed1988

Pericanalicular microfilaments of hepatocytes in patients with familial non-hemolytic hyperbilirubinemia.

We observed pericanalicular webs (PCW) of liver cells in cases with familial non-hemolytic hyperbilirubinemia using electron microscopy. The area and width of PCW were determined by morphometric methods as a way of quantitating this feature. The mean PCW width was 0.175 +/- 0.003 micron (mean +/- SE) in Dubin-Johnson syndrome and 0.184 +/- 0.005 micron in Rotor's syndrome. In both of these syndromes PCW width was significantly larger than that in Gilbert's syndrome (0.124 +/- 0.003 micron) (p less than 0.01). The mean PCW area was 0.585 +/- 0.017 micron 2 in Dubin-Johnson syndrome and 0.582 +/- 0.030 micron 2 in Rotor's syndrome. Values in these two syndromes were significantly larger than that in Gilbert's syndrome (0.382 +/- 0.014 micron 2) (p less than 0.01). Widths and areas of PCW in these three syndromes were not significantly different between central, intermediate, and peripheral zones of the hepatic lobules. There was a positive correlation between serum direct bilirubin levels and widths or areas of PCW in these syndromes. These results suggested that disturbances of bile flow caused by the dysfunction of pericanalicular microfilaments are partly involved in the pathogenesis of Dubin-Johnson syndrome and Rotor's syndrome.

Actin Cytoskeleton↗
PubMed1988

Infertility due to growth arrest of ovarian follicles in Sl/Slt mice.

Sl, Sld, and Slt are mutant alleles at the steel locus. All Sl/Sld and most Sl/Slt female mice are infertile, but the cause of the infertility is different. Germ cells are absent in Sl/Sld ovaries but present in Sl/Slt ovaries. The infertility of Sl/Slt female mice was attributed to the growth arrest of ovarian follicles, and the mechanism was analyzed by producing aggregation chimeras between Sl/Slt and +/+ embryos. Sl/Slt oocytes were ovulated and fertilized in Sl/Slt----+/+ chimeras. We investigated the origin of granulosa cells in the growing follicles and that of granulosa-derived luteal cells in the chimeras by using the electrophoretic pattern of phosphoglycerate kinase-1 and the histochemical activity of beta-glucuronidase as markers. Granulosa cells of Sl/Slt genotype developed and constituted pregnant corpora lutea in Sl/Slt----+/+ chimeras. Therefore, the growth arrest of Sl/Slt ovarian follicles may not be due to an intrinsic defect in granulosa cells but may instead be due to an intrinsic defect in ovarian stromal cells. This suggests that normal stromal cells are essential for the development of ovarian follicles.

Alleles↗
PubMed1988

Temperature dependency of calcium-induced reperfusion injury in the isolated rat heart.

The temperature dependence of Ca-induced reperfusion injury was studied in an isolated rat heart preparation. Hearts were subjected to 90 minutes of hypothermic arrest (20 degrees C) followed by 15 minutes of reperfusion at 20, 28, or 37 degrees C with a reperfusate containing various concentrations of Ca (0.1-2.55 mM). When reperfusion was started at 37 degrees C, the Ca concentration in the reperfusate significantly affected both postischemic functional recovery and creatine kinase leakage. Bell-shaped dose-response curves were observed. The optimal Ca concentration for 37 degrees C reperfusion was between 0.3 and 0.7 mM. When reperfusion was started at 20 degrees C, neither functional recovery nor creatine kinase leakage was dependent on the Ca concentration in the reperfusate. At 28 degrees C, functional recovery was not dependent on the Ca concentration, however, creatine kinase leakage was. These results indicate that Ca-induced reperfusion injury depends on the temperature of the reperfusate and that the boundary temperature of the reperfusate at which Ca-induced reperfusion injury becomes manifest seems to be near 28 degrees C.

Animals↗
PubMed1988

Fast and easy detection of mouse sex chimeras using electrophoretic polymorphism of phosphoglycerate kinase-1, an X chromosome-linked enzyme.

About half of the chimeras produced by aggregation of two mouse embryos are sex chimeras composed of both XX and XY cells. We developed a fast and easy method to identify sex chimeras by using electrophoretic bimorphism of an X-linked enzyme, phosphoglycerate kinase-1 (PGK-1), as a marker. When embryos resulting from the crossing of a Pgk-1b/Pgk-1b female and a Pgk-1a/Y male are aggregated, the genotype of sex chimeras is Pgk-1b/Pgk-1a----Pgk-1b/Y. Most of these were identifiable from the PGK-1 electrophoretic pattern of blood cells (i.e., AB type) and the appearance of genitalia (male type or apparently abnormal). Genotypes of functional sperm in the testes of the male-type sex chimeras were also identifiable from the PGK-1 electrophoretic pattern of progenies. Examination of gonads of the sex chimeras revealed that a considerable proportion was hermaphorditic. With this method, reasonable numbers of male-type sex chimeras and hermaphrodites may be selected and used as material for investigating sexual differentiation.

Animals↗
PubMed1988

Synthesis and disulfide structure determination of porcine endothelin: an endothelium-derived vasoconstricting peptide.

All disulfide analogs (types A, B and C) of porcine or human endothelin, a 21-amino acid peptide having two intramolecular disulfide bonds, were synthesized and their retention times on HPLC were compared with that of natural endothelin. One of the analogs (type A) having disulfide bonds between positions 1 and 15 and between 3 and 11 was found to be identical with natural endothelin. Random oxidation of fully reduced endothelin formed a mixture of type A and B in a ratio of 3:1, with almost none of type C, which has disulfide bonds between positions 1 and 3 and between 11 and 15. Type A endothelin was also synthesized by the segment condensation procedure in solution applying our maximum protection strategy. This product was found to have full vasoconstricting activity in rat pulmonary artery ring preparations; the potency was as high as that of the natural product.

Amino Acid Sequence↗
PubMed1988

Studies of Sl/Sld in equilibrium with +/+ mouse aggregation chimaeras. I. Different distribution patterns between melanocytes and mast cells in the skin.

In spite of their different origin, both melanocytes and mast cells are deficient in the skin of mutant mice of the Sl/Sld genotype. Since the neural crest and the liver of Sl/Sld embryos contain normal precursors of melanocytes and mast cells, respectively, the deficiency is attributed to a defect in tissue environment necessary for migration and/or differentiation of precursor cells. We investigated whether the tissue environment used for differentiation of melanocytes and mast cells was identical by producing aggregation chimaeras from Sl/Sld and +/+ embryos. Chimaeric mice with apparent pigmented and nonpigmented stripes were obtained. In the nonpigmented stripes of these Sl/Sld in equilibrium with +/+ chimaeras, melanocytes were not detectable in hair follicles but were detectable in the dermis. In contrast, melanocytes were detectable neither in hair follicles nor in the dermis of nonchimaeric Sl/Sld mice. Concentrations of mast cells were comparable in the pigmented and nonpigmented stripes of Sl/Sld in equilibrium with +/+ chimaeras, but the average concentration of mast cells significantly varied in the chimaeras (from 8% to 74% of the value observed in control +/+ mice). The present result suggests that mesodermal cells that support the migration and differentiation of both melanocyte precursors and mast-cell precursors mix homogeneously in the dermis and that ectodermal cells that influence the invasion of differentiating melanocytes into hair follicles make discrete patches.

Animals↗
PubMed1988

Studies of Sl/Sld in equilibrium with +/+ mouse aggregation chimaeras. II. Effect of the steel locus on spermatogenesis.

Mutant mice of Sl/Sld genotype are deficient in melanocytes, erythrocytes, mast cells and germ cells. Deficiency of melanocytes, erythrocytes and mast cells is not attributable to an intrinsic defect in their precursor cells but to a defect in the tissue environment that is necessary for migration, proliferation and/or differentiation. We investigated the mechanism of germ cell deficiency in male Sl/Sld mice by producing aggregation chimaeras from Sl/Sld and +/+ embryos. Chimaeric mice with apparent white stripes were obtained. Two of four such chimaeras were fertile and the phenotypes of resulting progenies showed that some Sl/Sld germ cells had differentiated into functioning sperms in the testis of the chimaeras. In cross sections of the testes of chimaeras, both differentiated and nondifferentiated tubules were observed. However, the proportions of type A spermatogonia to Sertoli cells in both types of tubules were comparable to the values observed in differentiated tubules of normal +/+ mice. We reconstructed the whole length of four tubules from serial sections. Differentiated and nondifferentiated segments alternated in a single tubule. The shortest differentiated segment contained about 180 Sertoli cells and the shortest nondifferentiated segment about 150 Sertoli cells. These results suggest that Sertoli cells of either Sl/Sld or +/+ genotype make discrete patches and that differentiation of type A spermatogonia does not occur in patches of Sl/Sld Sertoli cells.

Animals↗