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H Kanoh

Publications and source records attributed to H Kanoh.

At least 55 records · Page 3Linked to original sources
PubMed1996

Simultaneous determination of alpha-tocopherol and alpha-tocopherolquinone by high-performance liquid chromatography and coulometric detection in the redox mode.

A simple, selective and highly sensitive assay method for the simultaneous determination of alpha-tocopherol and alpha-tocopherolquinone in plasma or erythrocyte membrane by high-performance liquid chromatography (HPLC) with a series of multiple coulometric working electrodes (CWE) was investigated. For good separation of alpha-tocopherol and alpha-tocopherolquinone, an MC MEDICAL C18 reversed-phase column and a mobile phase consisting of 96% methanol [methanol-HPLC-grade distilled water (96:4, v/v)] with 40 mM sodium perchlorate were used. Also, selective, highly sensitive and simultaneous detection of these substances was performed in redox mode using a series of four CWE. In this detection mode, the first, second and third CWE were set at -0.45 V for pre-reaction and to prevent interference, the fourth CWE was used as an electrode for actual measurement with its potential set at +0.40 V against a palladium reference electrode. The detection limits were 50-100 pg. Excellent chromatograms of alpha-tocopherol and alpha-tocopherolquinone were obtained within 8 min. The usefulness of reversed-phase HPLC with the redox detection mode was confirmed by application to the determination of the concentrations of alpha-tocopherol and alpha-tocopherolquinone in a crude ethanol-hexane extract of rat plasma or erythrocyte membrane. These findings suggest that reversed-phase HPLC with the redox detection mode using a series of four CWE is applicable to study the preventive effect of alpha-tocopherol on lipid peroxidation.

Animals↗
PubMed1996

Membrane-associated phospholipase D activity in neural cell line PC12.

Stimulation of PC12 cells with carbachol (Cch) or phorbol 12-myristate 13-acetate (PMA) induced [3H]phosphatidylbutanol (PBut) production. Depletion of extracellular Ca2+ abolished the Cch-mediated phospholipase D (PLD) activation, indicating the requirement of Ca2+ influx. Two different types of PLD activity, oleate-dependent and GTP gamma S-dependent, were examined by using exogenous [3H]phosphatidylcholine substrate. PLD activity of the membrane fraction of PC12 cells was highly dependent on oleate and independent of GTP gamma S. This profile is in sharp contrast to that observed in HL60 cells showing the profound GTP gamma S-induced activation of PLD. The oleate-dependent PLD activity of PC12 membrane was inhibited by high concentrations of Ca2+ and Mg2+. These results indicate that Ca2+ may not directly activate PLD but through some Ca(2+)-dependent mechanism(s) in Cch-stimulated cells.

Animals↗
PubMed1996

Molecular properties of enzymes involved in diacylglycerol and phosphatidate metabolism.

Diacylglycerol and phosphatidate serve as key lipid intermediates of glycerolipid biosynthesis and also as the second messengers liberated in signal-stimulated cells. The molecular properties of enzymes involved in the metabolic processing of the two lipids are largely unknown. We have revealed by cDNA cloning the presence of a novel enzyme family of diacylglycerol kinase (DGK) having unique structural characteristics. Furthermore, we recently identified the plasma membrane-bound phosphatidate phosphatase (PAP) as a 35 kDa glycoprotein. Detailed studies on the two enzymes would contribute to understanding the molecular mechanisms underlying the control of both glycerolipid biosynthesis and signal transduction.

Animals↗
PubMed1996

Prevention of bone loss by bisphosphonate YM175 in ovariectomized dogs with dietary calcium restriction.

We have evaluated the effects of YM175 (disodium dihydrogen (cycloheptylamino) methylenebisphosphonate monohydrate), a novel bisphosphonate, on bone mineral densities (BMD) at the lumbar spine and forelimb in ovariectomized beagles with dietary calcium restriction. Groups 1 and 2 were given a sham operation and Groups 3-6 were ovariectomized. One month later (month 0), a low calcium diet was given to Groups 2-6. Groups 4-6 were orally treated with YM175 at doses of 0.01, 0.1 and 1.0 mg/kg, respectively, for 18 months. Changes in BMD at the lumbar spine and left forelimb were determined serially by dual energy X-ray absorptiometry. Calcium restriction decreased lumbar BMD by 19% at month 2 and by up to 30% at month 17 compared to its baseline value, but ovariectomy itself had a minimal effect on bone mass in dogs with restricted calcium intake. YM175 (1 mg/kg) prevented the bone loss at month 2 and YM175 at 0.1 mg/kg or more inhibited the BMD reduction at month 17. The magnitude of BMD reduction of the forelimb was less remarkable as compared to that of the lumbar spine. Urinary hydroxyproline excretion and plasma osteocalcin levels were increased by calcium restriction, indicating a high turnover of bone. YM175 reduced hydroxyproline excretion but not osteocalcin levels. These results indicate that YM175 prevents bone loss induced by calcium restriction and ovariectomy through partially normalizing high bone turnover.

Animals↗
PubMed1995

Chaperone function of calreticulin when expressed in the endoplasmic reticulum as the membrane-anchored and soluble forms.

A unique type of chaperone that requires glucose trimming of the target proteins has been shown to be important for their maturation in the endoplasmic reticulum (ER). Calnexin, an ER membrane chaperone, is the first example of such a class. Here, we focus on calreticulin, a major ER luminal protein, which shares with calnexin two sets of characteristic sequence repeat. We evaluated the chaperone function of calreticulin by expressing it on the ER luminal membrane surface. We constructed a membrane-anchored calreticulin chimera by fusing truncated calreticulin to the membrane-anchoring tagged segment of calnexin. When expressed in HepG2 cells, the calreticulin chimera transiently interacted with a set of nascent secretory proteins in a castanospermine-sensitive manner. The spectrum of proteins recognized by the membrane-anchored calreticulin was remarkably similar to that observed with calnexin. Next, we tested if such a chaperone function of calreticulin is expressed at its physiological location. Luminally expressed calreticulin preferentially bound to nascent transferrin and released it upon chase. Association with other calnexin ligands was observed, however, at low efficiencies. Interactions were abrogated by castanospermine treatment. We conclude that calreticulin per se is another chaperone with apparently the same characteristics as calnexin and selectively interacts with nascent transferrin in the lumen, suggesting that calreticulin may cover the diversity of maturations.

Amino Acid Sequence↗
PubMed1995

Effects of the bisphosphonate YM175 on bone mineral density, strength, structure, and turnover in ovariectomized beagles on concomitant dietary calcium restriction.

We evaluated the effect of YM175 on bone in ovariectomized beagles fed a calcium-restricted diet for 18 months. Groups 1 (n = 6) and 2 (n = 6) underwent sham operation, and groups 3-6 were ovariectomized at the age of 21 months. Group 1 was fed standard dog chow (calcium: 1.4%), and groups 2-6 were given a low calcium diet (0.14%). Groups 3 (n = 7), 4 (n = 7), 5 (n = 7), and 6 (n = 7) were given YM175 orally at doses of 0, 0.01, 0.1, and 1.0 mg/kg/day, respectively. At the end of the experimental period, bone mineral density values measured in the lumbar (L2) vertebra, the femoral neck, and the midfemur, were reduced in groups 2 and 3. Bone strength, measured by compression tests on L2 body and L3 cancellous core specimens and by bending tests on the femoral neck, also decreased. YM175 treatment dose-dependently increased the values of these parameters. However, the reduction in torsional stiffness in the midfemur was not completely prevented. In the L4 body, trabecular thickness decreased in group 2 and trabecular separation increased in group 3. YM175 treatment prevented these changes, and the values in group 6 were maintained at the same levels as those in group 1. Bone formation rates were increased in groups 2 and 3. YM175 treatment decreased these indices, but the reduction was incomplete even with the highest dose (group 6). These results demonstrate that, in our model, YM175 maintained the mass, structure, and mechanical properties of cancellous bone. Increased bone turnover was not completely prevented by the doses employed, but the balance between net resorption and formation in one remodeling cycle would have been equilibrated by YM175.

Administration, Oral↗
PubMed1995

Acute and subacute inhalation toxicity of diborane in male ICR mice.

To clarify the toxicity of diborane, we conducted acute (15 ppm for 1, 2, 4 or 8 h) and subacute (5 ppm for 2 or 4 weeks) inhalation studies on ICR mice. The concentration resulting in a 50% kill after 4 h exposure was 31.5 ppm. Body weight gain was suppressed and the lung weight was increased in diborane-exposed mice in both acute and subacute studies. In the acute study, diffuse pan bronchiolitis-like lesions developed in the lung in various degrees depending on exposure time, which can be pathologically characterized as infiltration of inflammatory cells into the terminal bronchioles and surrounding alveoli, pulmonary congestion and bleeding and/or edema. In the subacute study, we observed lymphoid hyperplasia in the perivascular and peribronchial areas, and infiltration of macrophage and plasma cells into the alveoli. In the mice exposed for 4 weeks, the lesions were more severe than in those exposed for 2 weeks, consisting of hyperplasia and desquamation of Clara cells. In the nasal cavity, we saw mucous exudate and inflammatory cells, suggesting irritation caused by diborane. The histopathological findings, except for the respiratory organs, did not reveal any exposure-related changes. No significant changes were seen in hematological and serum biochemical examinations either. In conclusion, the target organ of diborane inhalation is the respiratory organs, particularly the lung. Further inhalation experiments are essential to investigate the safety exposure levels of diborane.

Administration, Inhalation↗
PubMed1995

Activation of phosphoinositide-specific phospholipase C by transferrin in porcine cerebral arterial smooth muscle cells.

The effect of transferrin on phosphoinositide metabolism was investigated in smooth muscle cells isolated from the porcine basilar artery. Ferric iron-bound transferrin induced a rapid increase in the level of inositol phosphates, the metabolic products of phosphoinositides through the phospholipase C pathway. Neither transferrin free of ferric iron nor ferric iron alone caused the activation of phospholipase C. This study suggests that ferric iron-bound transferrin is capable of eliciting receptor-mediated signal transduction in porcine cerebral arterial smooth muscle cells, which could result in the contraction of smooth muscle cells. Transferrin may be involved with the cerebral arterial narrowing in pathological conditions such as subarachnoid hemorrhage.

Animals↗
PubMed1995

Localisation of the gene encoding diacylglycerol kinase 3 (DAGK3) to human chromosome 3q27-28 and mouse chromosome 16.

The gene encoding a 90 kDa diacylglycerol kinase protein, DAGK3, that is predominately expressed in the retina, was localised by fluorescence in situ hybridisation to human chromosome 3q27-28. This was subsequently confirmed by mapping of its mouse homologue to chromosome 16, a region syntenic to this part of human chromosome 3. No retinopathies have so far been assigned to this region.

Animals↗
PubMed1995

[A case of Binswanger's disease in which an acute infarcted lesion was detected by diffusion-weighted magnetic resonance imaging].

Diffusion-weighted magnetic resonance imaging (DWI) was carried out on a patient with Binswanger's disease suffering from acute cerebral infarction. Though an acute infarcted lesion was demonstrated as a high signal area on the T2-weighted image, it was impossible to determine whether it was acute or chronic because of extensive deep white matter lesions (periventricular hyperintensity and white matter hyperintensity lesions). However, only the acute infarcted lesion was detected on DWI which showed it as a high signal area, suggesting reduced molecular diffusion of water. The apparent diffusion coefficient (ADC), a physiological parameter that characterizes the self-diffusion on water in tissue, was lower in the acute lesion and higher in the chronic lesion. DWI can differentiate acute from chronic infarcts, which is not possible by conventional CT and MRI.

Acute Disease↗
PubMed1994

Preparation of high-molecular-weight DNA: application to mycobacterial cells.

A method for isolating high-molecular-weight DNA from bacteria is described. A special feature of the method is the treatment of whole bacterial cells with an organic solvent (chloroform-methanol (2:1, v/v) or ethanol-ether (1:1, v/v)) prior to DNA extraction from the cells. The DNA preparations obtained from organic solvent-pretreated bacterial cells such as Mycobacterium smegmatis, M. phlei, and Escherichia coli contained highly polymerized DNA, as revealed by pulse-field gel electrophoresis. The size and yield of the DNA obtained from E. coli pretreated with the organic solvent were quite similar to that of the DNA obtained from protoplasts. The results strongly suggest that the organic solvent pretreatment is effective for extracting very large DNA from bacterial cells and especially from bacteria whose protoplasts cannot be easily formed.

DNA, Bacterial↗
PubMed1994

Molecular cloning of a diacylglycerol kinase isozyme predominantly expressed in human retina with a truncated and inactive enzyme expression in most other human cells.

In order to clone novel diacylglycerol kinase (DGK) isozymes, we first obtained a DGK-related cDNA fragment by polymerase chain reaction using the human hepatoma cell line HepG2 mRNA and degenerated primers. The amplified fragment was subsequently used as a probe for screening the cDNA library from HepG2 cells. We obtained a cDNA clone coding for a novel DGK isozyme (designated DGK gamma) comprised of 791 amino acid residues. The amino acid sequence of DGK gamma was 52 and 62% identical to those of previously sequenced porcine 80-kDa and rat 90-kDa enzymes, respectively. DGK gamma, although initially cloned from the HepG2 cDNA libraries, was unexpectedly expressed in the human retina abundantly and to a much lesser extent in the brain. Other human tissues, including the liver and HepG2 cells, contained extremely low levels of DGK gamma mRNA. Furthermore, HepG2 cells and most of the human tissues except for the retina and brain expressed a truncated DGK gamma with an internal deletion of 25 amino acid residues (Ile451-Gly475). When transfected into COS-7 cells, the nontruncated cDNA gave phosphatidylserine-dependent DGK activity with no apparent specificity with regard to the acyl compositions of diacylglycerol. In contrast the truncated cDNA failed to give DGK activity in spite of the expression of its mRNA and enzyme protein in COS cells, thus demonstrating that the truncated DGK gamma is catalytically inactive. The sequence comparison of the three cloned DGKs revealed the presence of four highly conserved regions including the two sets each of EF-hand and zinc finger structures. Although the implication of the catalytically inactive form of DGK gamma remains unknown, this work further demonstrates the occurrence of multiple animal DGK isozymes with a conserved basic structure but with markedly different expression patterns depending on the cell types.

Amino Acid Sequence↗
PubMed1994

Activation of phospholipase D in Chinese hamster ovary cells expressing platelet-activating factor receptor.

Platelet-activating factor (PAF) activated phospholipase D (PLD) in WT-H cells, CHO cells stably expressing cloned guinea-pig PAF receptor. The PLD activation was found to be dependent on extracellular Ca2+, protein kinase C (PKC), and a currently unidentified protein tyrosine kinase (PTK). PTK inhibitors ST-638 and genistein inhibited PLD activation induced by PAF as well as phorbol myristate acetate, indicating that PTK acts downstream of PKC. Furthermore, activation of MAP (mitogen-activated protein) kinases, as assessed by their phosphorylation, was also dependent on Ca2+, PKC, and PTK. The correlation between PLD activity and MAP kinase activation, together with the previously observed MAP kinase activation associated with arachidonic acid release by cPLA2 [Honda et al. (1994) J. Biol. Chem. 269, 2307-2315], led us to examine the involvement of MAP kinase in PLD activation. The results indicate that PLD and MAP kinases are activated through the common pathway consisting of Ca2+, PKC, and the unidentified PTK, which act in parallel, but not in a linear sequence.

Animals↗
PubMed1994

Identification of the gene encoding the major NAD(P)H-flavin oxidoreductase of the bioluminescent bacterium Vibrio fischeri ATCC 7744.

The gene encoding the major NAD(P)H-flavin oxidoreductase (flavin reductase) of the luminous bacterium Vibrio fischeri ATCC 7744 was isolated by using synthetic oligonucleotide probes corresponding to the N-terminal amino acid sequence of the enzyme. Nucleotide sequence analysis suggested that the major flavin reductase of V. fischeri consisted of 218 amino acids and had a calculated molecular weight of 24,562. Cloned flavin reductase expressed in Escherichia coli was purified virtually to homogeneity, and its basic biochemical properties were examined. As in the major flavin reductase in crude extracts of V. fischeri, cloned flavin reductase showed broad substrate specificity and served well as a catalyst to supply reduced flavin mononucleotide (FMNH2) to the bioluminescence reaction. The major flavin reductase of V. fischeri not only showed significant similarity in amino acid sequence to oxygen-insensitive NAD(P)H nitroreductases of Salmonella typhimurium, Enterobacter cloacae, and E. coli but also was associated with a low level of nitroreductase activity. The major flavin reductase of V. fischeri and the nitroreductases of members of the family Enterobacteriaceae would thus appear closely related in evolution and form a novel protein family.

Amino Acid Sequence↗
PubMed1993

Isolation and characterization of the human diacylglycerol kinase gene.

The 80 kDa diacylglycerol kinase (DGK) is abundantly expressed in oligodendrocytes and lymphocytes but not to a detectable extent in other cells such as neurons and hepatocytes. As an initial attempt to delineate the mechanism of the transcriptional control of the DGK gene, we have cloned from a human genomic library a 22 kb genomic fragment. The genomic clone consists of the 5'-flanking region and 17 exons coding for approx. 53% of the total exons of human DGK, including those encoding EF-hand and zinc-finger regions. The translation initiation site is located in the second exon. S1 nuclease mapping and primer extension analysis of the human DGK mRNA identified a major transcription initiation site (position +1) at 264 bp upstream from the initiator ATG. In the 5'-flanking sequence we detected a single GC box at -35 but no canonical TATA and CAAT sequences. However, the sequence starting from the cap site (AGTTCCTGCCA) is very similar to the initiator element that specifies the transcription initiation site of some housekeeping genes. In addition, the 5'-upstream region contains several putative cis-elements. Jurkat and HepG2 cells were transfected with various 5'-deletion mutants of the upstream region fused to the structural gene of chloramphenicol acetyltransferase (CAT). The CAT assay revealed that among constructs containing up to 3.4 kb of the 5'-flanking region, a fragment of 263 bp from the transcription initiation site contains a basic promoter that is active in both types of cells. Moreover, the region between -263 and -850 contains a negative element that is active in HepG2 but not in Jurkat cells. This negative element may, at least in part, be responsible for the cell type-specific expression of the DGK gene.

Animals↗
PubMed1993

Phospholipase D activity of human amnion cells stimulated with phorbol ester and bradykinin.

We investigated the activity of phospholipase D (PLD) in human amnion cells labeled with [3H]oleate. The PLD activity was detected as signal-induced synthesis of phosphatidic acid (PA) and in the presence of ethanol, phosphatidylethanol (PEt). The PLD was shown to be activated by phorbol, 12-myristate, 13-acetate (PMA), calcium ionophore A23187, oxytocin, bombesin and bradykinin, but not by platelet-activating factor (PAF) and epidermal growth factor (EGF). The amniotic PLD thus appeared to be activated by a variety of agonists but with a certain specificity to stimulators. We examined the mode of the PLD activation using PMA (20 nM) and bradykinin (1 microM) as model stimulators. PMA and bradykinin elicited a rapid and sustained response with the peaks of PA-labeling attained at 5 and < 1 min after stimulation, respectively. In both cases, there was a concomitant rise of diacylglycerol (DG), and the PA accumulation was suppressed by ethanol at the expense of labeling of PEt. The PA synthesis caused by the two stimulators was similarly inhibited by staurosporine and by a chronic treatment with PMA (100 nM for 24 h), suggesting that the activation of PLD is linked to the action of protein kinase C. With the cells labeled with radioactive choline and ethanolamine, we found that the amniotic PLD hydrolyzed almost equally phosphatidylcholine and phosphatidylethanolamine. Although bradykinin and PMA stimulated cellular PLD to a comparable extent, prostaglandin (PG)E2 release was not stimulated by bradykinin in contrast to the marked effect by PMA. Further work is thus needed to clarify the significance of the novel PLD signaling pathway in the function of amnion cells.

Amnion↗
PubMed1993

Requirement of adenosine 5'-triphosphate and Ca2+ for guanosine 5'-triphosphate-binding protein-mediated phospholipase D activation in rat pheochromocytoma PC12 cells.

In digitonin-permeabilized PC12 cells labeled with [3H]palmitic acid, formation of [3H]phosphatidylethanol ([3H]PEt), a marker of phospholipase D (PLD) activity, was increased with increasing concentrations of Ca2+ in the presence of ATP, Mg2+ and ethanol. Guanosine-5'-O-(3-thio-triphosphate) (GTP gamma S) significantly enhanced Ca(2+)-stimulated [3H]PEt formation. The effect of GTP gamma S was abolished when any one of Ca2+, ATP or Mg2+ was excluded. ATP could not be replaced by other nucleotides except for ADP. Thus, the GTP-binding protein-mediated PLD activation absolutely required both Ca2+ and Mg-ATP.

Adenosine Triphosphate↗