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Biomedical subjects

E Miyamoto

Publications and source records attributed to E Miyamoto.

At least 163 records · Page 9Linked to original sources
PubMed1990

Effect of dimethadione derived from repeated oral administration of trimethadione on pancreatic secretion in dogs.

The effect of the weak organic acid of dimethadione (DMO) on secretin-stimulated pancreatic secretion was studied with repeated oral administration of trimethadione (TMO), the precursor of DMO, to dogs at a dose of 10 to 160mg/kg/day for a period of 14 days. The bicarbonate concentration in pancreatic juice at a steady state decreased significantly, reflecting a close correlation with the dose of TMO and DMO concentrations in plasma and pancreatic juice. The maximal decrement from the control of cases of no TMO administration was 18.8 mEq/l (12.1% of the control level). The chloride concentration in pancreatic juice showed a reciprocal relation to the bicarbonate concentration. The sum of both anion concentration was constant, irrespective of the dose of TMO. The average carbon dioxide tension of pancreatic juice in all doses of TMO was lower than that of the control, but differences were not statistically significant. The pH, flow rate, sodium and potassium concentrations in pancreatic juice at a steady state did not differ significantly in relation to the dose of TMO. These findings suggest that repeated oral administration of TMO cause a significant decrease in bicarbonate concentration in pancreatic juice, resulting probably from the buffer action of bicarbonate on protons provided from the undissociated form of DMO.

Administration, Oral↗
PubMed1990

Ca2+/calmodulin-dependent protein kinase II: localization in the interphase nucleus and the mitotic apparatus of mammalian cells.

Indirect immunofluorescence was used to determine the distribution of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) in rat embryo fibroblast 3Y1 cells, rat C6 glioma cells, and human epidermoid carcinoma KB cells. During interphase at growing phase, CaM kinase II was localized diffusely in the cytoplasm and in the nucleus. In the nucleus, the enzyme was localized within the whole nuclear matrix in which the enzyme was specially concentrated in nucleoli. During mitosis, CaM kinase II was found to be a dynamic component of the mitotic apparatus, particularly present at microtubule-organizing centers. In metaphase and anaphase, CaM kinase II was observed at centrosomes and between the spindle poles. During telophase, CaM kinase II was condensed as a bright fluorescent dot at the midzone of the intercellular bridge between two daughter cells, while tubulin was found at each side of the midbody. Colchicine, a microtubule inhibitor, disorganized the tubulin- and CaM kinase II specific fluorescent structure of mitotic 3Y1 cells. In cold-treated cells, CaM kinase II was localized predominantly at centrosomes. The localization of CaM kinase II in the cell nucleus and the mitotic apparatus suggests that the enzyme may play a role in the cell cycle progression of mammalian cells.

Animals↗
PubMed1990

Dephosphorylation of tau factor by protein phosphatase 2A in synaptosomal cytosol fractions, and inhibition by aluminum.

When the synaptosomal cytosol fraction from rat brain was chromatographed on a DEAE-cellulose column and assayed for protein phosphatases for tau factor and histone H1, two peaks of activities, termed peak 1 (major) and peak 2 (minor), were separated. Each peak was in a single form 2 (minor), were separated. Each peak was in a single form on Sephacryl S-300 column chromatography. Both peaks 1 and 2 dephosphorylated tau factor phosphorylated by Ca2+/calmodulin-dependent protein kinase II and the catalytic subunit of cyclic AMP-dependent protein kinase. The Km values were in the range of 0.42-0.84 microM for tau factor. There were no differences in kinetic properties of dephosphorylation between the substrates phosphorylated by the two kinases. The phosphatase activities did not depend on Ca2+, Mn2+ and Mg2+. Immunoprecipitation and immunoblotting analysis using polyclonal antibodies to the catalytic subunit of brain protein phosphatase 2A revealed that both protein phosphatases are the holoenzymic forms of protein phosphatase 2A. Aluminum chloride inhibited the activities of both peaks 1 and 2 with IC50 values of 40-60 microM. These results suggest that dephosphorylation of tau factor in presynaptic nerve terminals is controlled mainly by protein phosphatase 2A and that the neurotoxic effect of aluminum seems to be related mostly to inhibition of dephosphorylation of tau factor.

Adenosine Triphosphate↗
PubMed1990

Phosphorylation of P0 glycoprotein in peripheral nerve myelin.

The P0 protein in mammalian PNS myelin is known to undergo several posttranslational modifications, such as glycosylation, acylation, sulfation, and phosphorylation. Phosphorylation of purified P0 protein in vitro was studied comparatively using three enzymes, i.e., calcium/phospholipid-dependent protein kinase (protein kinase C), calcium/calmodulin-dependent protein kinase II (CaM kinase II), and the catalytic subunit of cyclic AMP-dependent protein kinase (A kinase). The phosphorylation of P0 protein by CaM kinase II was the greatest, followed by that by protein kinase C; phosphorylation by A kinase, however, was much lower. In order to identify phosphorylation sites, P0 protein was phosphorylated with [32P]ATP and each kinase and then digested with lysylendopeptidase. The resulting phosphopeptides were isolated by HPLC. Subsequent amino acid sequence analysis and comparison with the known sequence of P0 protein revealed that Ser181 and Ser204 were strongly phosphorylated by both protein kinase C and CaM kinase II. In addition, Ser214 was also phosphorylated by protein kinase C, but not by CaM kinase II. Because all of these sites are located in the cytoplasmic domain of P0 protein, phosphorylation may be important for maintenance of the major dense line of PNS myelin.

Amino Acids↗
PubMed1990

Regulation of Ca2+/calmodulin-dependent protein kinase II by brain gangliosides.

Purified rat brain Ca2+/calmodulin-dependent protein kinase II (CaM-kinase II) is stimulated by brain gangliosides to a level of about 30% the activity obtained in the presence of Ca2+/calmodulin (CaM). Of the various gangliosides tested, GT1b was the most potent, giving half-maximal activation at 25 microM. Gangliosides GD1a and GM1 also gave activation, but asialo-GM1 was without effect. Activation was rapid and did not require calcium. The same gangliosides also stimulated the autophosphorylation of CaM-kinase II on serine residues, but did not produce the Ca2+-independent form of the kinase. Ganglioside stimulation of CaM-kinase II was also present in rat brain synaptic membrane fractions. Higher concentrations (125-250 microM) of GT1b, GD1a, and GM1 also inhibited CaM-kinase II activity. This inhibition appears to be substrate-directed, as the extent of inhibition is very dependent on the substrate used. The molecular mechanism of the stimulatory effect of gangliosides was further investigated using a synthetic peptide (CaMK 281-309), which contains the CaM-binding, inhibitory, and autophosphorylation domains of CaM-kinase II. Using purified brain CaM-kinase II in which these regulatory domains were removed by limited proteolysis. CaMK 281-309 strongly inhibited kinase activity (IC50 = 0.2 microM). GT1b completely reversed this inhibition, but did not stimulate phosphorylation of the peptide on threonine-286. These results demonstrate that GT1b can partially mimic the effects of Ca2+/CaM on native CaM-kinase II and on peptide CaMK 281-309.

Animals↗
PubMed1990

Antiplaque activity of some antimicrobial agents using a simple in vitro method.

Some antimicrobial compounds were tested for their antiplaque activity by a simple method involving measurement of the weight of Streptococcus sobrinus plaque on a glass surface. Chlorhexidine, cetylpyridinium chloride, and n-tetradecylamine reduced the weight of plaque produced by S. sobrinus in vitro; some N-palmityl and palmitoyl derivatives of polymethylenediamines were also found to possess antiplaque activity. The results suggest that an antiplaque effect does not necessarily depend on high bactericidal activity.

Amines↗
PubMed1990

[A study on the role and mechanism of intracellular Ca2+ in the central nervous system].

When the stimuli by nerve impulses, neurotransmitters, hormones, peptides and growth factors are administered to the neurons, one of the responses of the nerve cells is the enhancement of Ca2+ influx and/or the release of Ca2+ from the intracellular storage site. Ca2+ may be related to several types of neuronal functions such as biosynthesis of neurotransmitters, stimulus-secretion coupling of neurotransmitters and hormones, microtubule assembly-disassembly cycle and many metabolic reactions. Although the precise molecular mechanism mediating the actions of Ca2+ in the brain remains to be elucidated, accumulating evidence suggests that the actions of Ca2+ are mediated through Ca2(+)-binding proteins. The role of troponin C, a Ca2(+)-binding protein, was extensively studied in the skeletal muscle first. Subsequently calmodulin, a ubiquitous Ca2(+)-binding protein, was found to be widely distributed in many tissues and to be involved in a variety of Ca2(+)-mediated cellular processes. In an attempt to elucidate Ca2+ actions in the central nervous system, we have been studying Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) and calcineurin (Ca2+/calmodulin-dependent protein phosphatase). These enzymes have many common substrates and, therefore, may be involved in the neuronal functions via phosphorylation and dephosphorylation of specific proteins.

Animals↗
PubMed1989

Dephosphorylation of cofilin accompanies heat shock-induced nuclear accumulation of cofilin.

Cofilin is a widely distributed 21-kDa actin-modulating protein that forms intranuclear actin/cofilin rods in cultured fibroblastic cells exposed to heat shock or 10% dimethyl sulfoxide. In this study, cofilin was shown to be phosphorylated on a serine residue in cultured rat fibroblastic 3Y1 cells. Two-dimensional gel electrophoresis revealed that about 50% of the cofilin was phosphorylated in 3Y1 cells at 37 degrees C. Exposure of the cells to heat shock at 43 degrees C induced dephosphorylation of cofilin. The dephosphorylation of cofilin was detected about 30 min after the temperature shift and was completed within 120 min. Moreover, treatment of cells with 10% dimethyl sulfoxide also caused the dephosphorylation of cofilin. However, incubation of the cells with an isotonic NaCl solution, which induced cytoplasmic actin/cofilin rods, did not induce dephosphorylation of cofilin. Other cellular stress agents such as 6% ethanol or 50 microM sodium arsenite, which caused some heat shock responses in cells, did not induce dephosphorylation of cofilin. Thus, cofilin dephosphorylation was closely correlated with its nuclear accumulation. Incubation of the enucleated 3Y1 cells at 43 degrees C still induced dephosphorylation of cofilin, suggesting that the dephosphorylation occurred mostly in the cytoplasm in intact cells. It is likely that cofilin is dephosphorylated in the cytoplasm prior to its nuclear accumulation.

Actin Depolymerizing Factors↗
PubMed1989

Characterization of polyclonal antibodies to brain protein phosphatase 2A and immunohistochemical localization of the enzyme in rat brain.

Polyclonal antibodies to the catalytic subunit of protein phosphatase 2A (PrP-2A) from bovine brain were prepared by immunizing rabbits and then purified by antigen-affinity column chromatography. The purified antibodies recognized only PrP-2A among proteins examined, including calcineurin and PrP-1. The antibodies cross-reacted only with a protein in the crude homogenate from rat brain, which comigrated with the catalytic subunit of PrP-2A on SDS-PAGE. The antibodies completely inhibited the activity of PrP-2A, and immunoprecipitated the purified enzyme. Immunoblot analysis demonstrates that, among the subcellular fractions from rat brain, the cytosol fraction and the synaptosomal cytosol fraction show high immunoreactivities, and that any of examined regions of rat brain shows immunoreactivity more or less, in which the caudatoputamen was highest. Immunohistochemical studies demonstrate that the enzyme is distributed widely in various regions of rat brain and that the immunoreactivity is localized mainly in neurons. In general, immunostaining of neurons was strong in neurites as well as somata. It was noted that intracerebellar nuclei were strongly stained in both neuronal somata and dendrites. Amygdaloid complex, thalamus, neocortex, hippocampal formation, and caudatoputamen were moderately stained.

Animals↗
PubMed1989

Testicular oximetry: a new method for the assessment of tissue perfusion and viability following torsion and detorsion.

This study evaluated the use of a polarographic surface PO2 electrode to assess testicular perfusion and viability following torsion and detorsion. Adult male Sprague-Dawley rats were divided into groups and subjected to unilateral testicular torsion and detorsion of varying degrees and durations. Rats subjected to sham torsion or 720 degrees torsion did not show significant decreases in testicular PO2 after 15 minutes, whereas those subjected to 1,080 degrees torsion or spermatic cord ligation uniformly decreased their testicular PO2 to 0 mm Hg within 10 minutes. Testicular PO2 values were similar in rats subjected to 60 minutes of 720 degrees torsion followed by detorsion and those undergoing 15 minutes of 1,080 degrees torsion and detorsion. Rats subjected to breathing 100% oxygen uniformly increased their testicular PO2 to an average of more than twice room-air values. However, rats subjected to 1,080 degrees torsion for 6 hours followed by detorsion did not increase their testicular PO2 when subjected to breathing 100% oxygen, whereas those subjected to 720 degrees torsion for 6 hours followed by detorsion did increase their testicular PO2 when subjected to breathing 100% oxygen. The latter rats did not show microscopic changes associated with acute testicular infarction, whereas the former did.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗
PubMed1989

Phosphorylation of P400 protein by cyclic AMP-dependent protein kinase and Ca2+/calmodulin-dependent protein kinase II.

Purified P400 protein was phosphorylated by both purified Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) and the catalytic subunit of cyclic AMP-dependent protein kinase (A-kinase). Because P400 protein was suggested to function as an integral membrane protein, we investigated the phosphorylation of P400 protein using crude mitochondrial and microsomal fractions (P2/P3 fraction). Incubation of the P2/P3 fraction from mouse cerebellum with cyclic AMP or the catalytic subunit of A-kinase stimulated the phosphorylation of P400 protein. The phosphorylation of P400 protein was not observed in the P2/P3 fraction from mouse forebrain. Cyclic AMP and A-kinase enhanced the phosphorylation of several proteins, including P400 protein, suggesting that P400 protein is one of the best substrates for A-kinase in the P2/P3 fraction. Although endogenous and exogenous CaM kinase II stimulated the phosphorylation of some proteins in the P2/P3 fraction, the phosphorylation of P400 protein was weak. Immunoprecipitation with the monoclonal antibody to P400 protein confirmed that the P400 protein itself was definitely phosphorylated by the catalytic subunit of A-kinase and CaM kinase II. A-kinase phosphorylated only the seryl residue in P400 protein. Immunoblot analysis of the cells in primary culture of mouse cerebellum confirmed the expression of P400 protein, which migrated at the same position on sodium dodecyl sulfate-polyacrylamide gel electrophoresis as that in the P2/P3 fraction. Incubation of the cultured cerebellar cells with [32P]orthophosphate resulted in the labeling of P400 protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗