Biomedical subjects

K Kataoka

Publications and source records attributed to K Kataoka.

At least 145 records · Page 8Linked to original sources

PubMed1998

Differential regulation of microglial activation by propentofylline via cAMP signaling.

A pathological microglial activation is believed to contribute to progressive neuronal damage in neurodegenerative diseases by the release of potentially toxic agents and by triggering reactive astrocytic changes. Using cultured microglia from neonatal rat brains, we investigated the mode of propentofylline action in strengthening cAMP-dependent intracellular signaling. We compared this action with the effects of dibutyryl-cAMP, a cell-permeable cAMP analog. Propentofylline inhibited lipopolysaccharide (LPS)-induced release of both tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta in a dose-dependent manner within the therapeutic low micromolar range. However, LPS-induced release of IL-6 and NO were not affected by propentofylline. All these differential effects of propentofylline on LPS-induced microglial release were mimicked by the addition of dibutyryl-cAMP. Microglial proliferation and phorbol myristate acetate (PMA)-induced O2- release were also dose-dependently inhibited by propentofylline as well as dibutyryl-cAMP. These results suggest that propentofylline, probably via reinforcement of cAMP intracellular signaling, alters the profile of the newly adopted immune properties in a way that it inhibits potentially neurotoxic functions while maintaining beneficial functions. This differential regulation of microglial activation may explain the neuroprotective mechanism exerted by propentofylline.

Animals↗

PubMed1998

Incorporation of water-insoluble anticancer drug into polymeric micelles and control of their particle size.

A water-insoluble anticancer drug, KRN 5500 (KRN), was incorporated into polymeric micelles forming from poly(ethylene glycol-poly(amino acid) block copolymers by physical entrapment utilizing hydrophobic interactions between this drug and the poly(amino acid) chain block of the block copolymers. Three block copolymers were examined for this incorporation; poly(ethylene glycol)-poly(beta-benzyl l-aspartate) (PEG-PBLA) and its two derivatives obtained by partial hydrolysis at the beta-benzyl l-aspartate (BLA) units (PEG-P(Asp, BLA)) and by partial cetyl ester substitution at the BLA units (PEG-P(C16, BLA)), respectively. Among these block copolymers, considerable effects of the cetyl esterification were seen on KRN yield and particle size. Considerable differences in the KRN incorporation yield and particle size were also observed between DMF and DMS used as solvent to dissolve KRN and the block copolymers. Sonication was turned out to be an effective method to obtain a polymer micelles fraction in high efficiency, and sonication was considered to work for separating intermicellar associates into dispersed micelles. A KRN incorporation procedure by dialysis using PEG-P(C16, BLA) and DMSO (as solvent) followed by sonication brought about polymeric micelles of 71 nm in weight-average diameter. This shows successful incorporation of a water-insoluble drug into polymeric micelles by optimizing block copolymer structure and incorporation conditions.

Antibiotics, Antineoplastic↗

PubMed1998

Functional change of NMDA receptors related to enhancement of susceptibility to neurotoxicity in the developing pontine nucleus.

The developing neurons have been reported to be extremely susceptible to toxicity of NMDA during a restricted developmental period. Pontosubicular neuronal necrosis is a typical type of perinatal human brain lesion and often coexists with other forms of cerebral hypoxic and ischemic injuries. To determine whether functional changes of NMDA receptors related to the susceptibility to NMDA toxicity are involved in developing neurons in the pontine nucleus, we have examined the lesion produced by in vivo direct injection of NMDA into the pontine nucleus of rats at postnatal days 1-30, recorded NMDA-induced whole-cell currents from neurons in the pontine nucleus in the developing rat brainstem slices, and performed in situ hybridization for NMDA receptor subunit mRNAs in the pontine nucleus. The susceptibility to NMDA neurotoxicity peaked near postnatal day 15, and the NMDA-induced currents showed prominent reduction of the voltage-dependent block by Mg2+ near postnatal day 15. The pontine nucleus near postnatal day 15 showed distinct expression of the NMDA receptor subunit NR2C mRNA. These results suggest that the susceptibility to NMDA neurotoxicity that is enhanced in the rat pontine nucleus near postnatal day 15 is mediated by the NMDA receptor channels that are relatively insensitive to Mg2+ and that the reduction in the sensitivity of NMDA receptors to Mg2+ correlates with the expression of the NR2C. We present the possibility that functional changes in the NMDA receptor channels play a crucial role in the occurrence of developmentally specific neuronal injury.

Animals↗

PubMed1998

Correlation between potentiation of AP1 DNA binding and expression of c-Fos in association with phosphorylation of CREB at serine133 in thalamus of gerbils with ischemia.

Protein biosynthesis is mainly under the control at the level of gene transcription in eukaryotes. Transcription factors are nuclear proteins with abilities to modulate the activity of RNA polymerase II which is responsible for the formation of messenger RNA from double stranded DNA in the cell nuclei. Binding of a radiolabeled oligonucleotide probe for the transcription factor activator protein-1 (AP1) was transiently potentiated 1 to 6 h after the recirculation of blood supply in the thalamus and striatum, but not in the entorhinal cortex, olfactory bulb, frontal cortex, cerebellar cortex and medulla-pons, in gerbils with transient global forebrain ischemia for 5 min, in addition to the hippocampal subregions. The ischemic insult not only increased the immunoreactivity with an antibody against cyclic AMP response element binding protein (CREB) phosphorylated at serine133, but also induced the expression of both c-Jun and c-Fos family proteins 3 h after the recirculation in the thalamus. Limited proteolysis by Staphylococcus aureus (S. aureus) V8 protease revealed the expression of different partner proteins of AP1 in response to ischemic signals in the thalamus. Moreover, ischemia for 2 min led to more prolonged elevation of AP1 binding in the thalamus at least up to 12 h after the reperfusion than that seen with ischemia for 5 min. These results suggest that potentiation of AP1 DNA binding may at least in part involve mechanisms associated with the expression of c-Fos protein through phosphorylation of CREB at serine133 in the thalamus of gerbils with ischemia.

Amino Acid Sequence↗

PubMed1998

Gene synthesis, expression, and mutagenesis of zucchini mavicyanin: the fourth ligand of blue copper center is Gln.

The gene coding for the 109-amino-acid, non-glycosylated form of mavicyanin was synthesized and expressed in Escherichia coli. The recombinant protein refolded from E. coli inclusion bodies was purified and characterized. Its spectroscopic properties are fully identical to those of mavicyanin isolated from zucchini, even in the absence of its carbohydrate moiety. The blue cooper center of mavicyanin strongly binds three ligands (2His and Cys) as well as many blue copper proteins. To disclose the fourth ligand of mavicyanin, Met was substituted for Gln95 by site-directed mutagenesis. The replacement changes from a rhombic EPR signal to an axial one and exhibits the quite similar absorption and CD spectra to those of plastocyanin. The midpoint potential of Gln95-->Met mavicyanin shows the positive shift of 187 mV compared with the recombinant protein, being close to the values of plastocyanins. The differences of the spectroscopic and electrochemical properties between mavicyanin and its mutant demonstrate that the fourth ligand of mavicyanin is Gln95.

Amino Acid Sequence↗

PubMed1998

Polymeric micelles for drug delivery: solubilization and haemolytic activity of amphotericin B.

Polymeric micelles may serve as nanoscopic, long-circulating carriers of hydrophobic drugs. In this study, we have researched the solubilization of amphotericin B (AmB), an antifungal drug, by micelles of poly(ethylene oxide)-block-poly(beta benzyl-L-aspartate) (PEO-PBLA), the properties of the AmB-loaded PEO-PBLA micelles and the resultant haemolytic activity of AmB. AmB loading takes place during self assembly of PEO-PBLA micelles, and this occurs through a dialysis procedure as an alkaline aqueous solution replaces the selective solvent for the polymer and the drug. In this way, AmB reaches levels of 57 to 141 microg/ml, corresponding to a loading efficiency of 27-30% (loaded AmB/initial amount of AmB). The molar ratio of AmB to PEO-PBLA is 0.40 to 1.0. Pictures by transmission electron microscopy reveal spherical AmB-loaded PEO-PBLA micelles with a mean diameter of 25.8+/-4.2 nm. AmB-loaded PEO-PBLA micelles are nonhaemolytic at an AmB level of 10 microg/ml as assessed by release of haemoglobin, measured by UV-Vis spectroscopy. AmB as Fungizone, its standard formulation, completely lyses red blood cells at a level of 3.0 microg/ml in 30 min. In contrast, there is no haemolysis at 5.5 h for AmB-loaded PEO-PBLA micelles at 3.0 microg/ml of AmB, indicating the gradual release of AmB from PEO-PBLA micelles. PEO-PBLA itself is nonhaemolytic even at a level of 0.70 mg/ml. Most amphiphiles, e.g. sodium deoxycholate, present in Fungizone, are haemolytic. Finally, AmB-loaded PEO-PBLA micelles can be freeze-dried and easily reconstituted in water. Afterwards, AmB is present in the intact PEO-PBLA micelles and remains nonhaemolytic.

Amphotericin B↗

PubMed1998

Prolongation by bifemelane of potentiation of AP1 DNA binding in hippocampal CA1 subfield of gerbils with transient forebrain ischemia.

In eukaryotes, protein de novo synthesis is mainly under the control of transcription factors at the level of gene transcription in cell nuclei. Gel retardation electrophoresis was employed for determination of DNA-binding activity of the transcription factor activator protein-1 (AP1), which is a dimer between c-Fos and c-Jun protein families. Binding of a radiolabeled double-stranded oligonucleotide probe for AP1 was rapidly potentiated in the CA1 and CA3 subfields and the dentate gyrus of the hippocampus of gerbils with forebrain ischemia for 5 min. Similarly marked potentiation was seen in the thalamus and the striatum, but not in the frontal cortex, following the recirculation of blood supply. The potentiation was transient in the vulnerable CA1 subfield, but was rather persistent in the thalamus and the striatum in addition to the resistant CA3 subfield and dentate gyrus. However, administration of the neuroprotective drug bifemelane (10 to 20 mg/kg, i.p.) resulted in prolongation of the potentiation of AP1 binding in the CA1 subfield up to 6 hr after ischemia, without significantly affecting that in other central structures. Limited proteolysis revealed that bifemelane induced expression of the AP1 consisting of constructive proteins different from those expressed in control animals in the CA1 subfield. These results suggest that bifemelane may protect neuronal cells against ischemic injuries through molecular mechanisms associated with prolongation of the potentiation of AP1 binding in the vulnerable CA1 subfield after ischemia.

Animals↗

PubMed1998

Characterization of physical entrapment and chemical conjugation of adriamycin in polymeric micelles and their design for in vivo delivery to a solid tumor.

An anticancer drug adriamycin (ADR) was incorporated into polymeric micelles forming from poly(ethylene glycol)-poly(aspartic acid) block copolymer by chemical conjugation and physical entrapment. Structural stability of the polymeric micelles was found to be dependent on both the contents of chemically conjugated and physically entrapped ADR. The polymeric micelle with high contents of the chemically conjugated ADR and the physically entrapped ADR expressed very high in vivo antitumor activity against murine C 26 tumor, while the polymeric micelle with only the chemically conjugated ADR showed negligible in vivo activity. This indicates that the physically entrapped ADR played a major role in antitumor activity in vivo. For the polymeric micelle with the high ADR contents, it was found that a dimer of adriamycin molecules formed and that this dimer was physically entrapped in the inner core of the micelle as well as intact ADR.

Animals↗

PubMed1998

Matrix-assisted laser desorption/ionization and electrospray ionization mass spectrometry analysis of blue copper proteins. Azurin and mavicyanin.

Two copper proteins azurin-1 and azurin-2 were isolated from denitrifying bacteria Alcaligenes xylosoxidans GIFU1051, and the mass spectrometric analysis of the proteins were carried out by both matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) and electrospray ionization (ESI). The mass spectrometric analysis was also carried out with the recombinant zucchini protein mavicyanin, which was obtained by expression in Escherichia coli. All the proteins were detected as positive ions with the copper atom being eliminated. The molecular weights were determined as 14,017.6 for azurin-1, 13,807.6 for azurin-2 and 11,808.8 for mavicyanin. The observed molecular weight of azurin-1 agrees within two daltons with that calculated from the amino acid composition. Azurin-2 was found to have one different amino acid residue when compared with the known azurin-2 isolated from A. xylosoxidans NCIB11015. The measured molecular weight for the recombinant mavicyanin agrees within two daltons with that of calculated from the amino acid composition of the native protein; therefore, the recombinant mavicyanin is identical to the native protein.

Azurin↗

PubMed1998

Mild hypothermia--a revived countermeasure against ischemic neuronal damages.

Although hypothermia as a means of cerebral protection against and resuscitation from ischemic damage has a history of approximately six decades, extensive studies, both in basic and clinical fields, on the mechanisms, effects and methods of mild hypothermia at temperatures no less than 31 degrees C have started only in the last decade. In experiments on rodents, hypothermia in the postischemic period that is introduced up to several hours after reperfusion and is maintained for one day followed by a slow rewarming, significantly protects hippocampal neurons against damage. The mode of action of hypothermia is apparently non-specific and multi-focal in widely progressing cascade reactions in ischemic cells; namely, suppressing: (1) glutamate surge followed by; (2) intraneuronal calcium mobilization; (3) sustained activation of glutamate receptors; (4) dysfunction of blood brain barrier; (5) proliferation of microglial cells; and (6) production of superoxide anions and nitric oxide. In addition, mild hypothermia modulates processes in ischemic condition at the level of cell nucleus, such as the binding of transcription factor AP-1 to DNA, and ameliorates the depression of protein synthesis. This non-specific and widely affecting manner might explain why hypothermia is superior to any medicine developed. Recent clinical trials of mild hypothermia in various individual institutions have revealed significantly beneficial outcomes in some cases, along with an accumulation of practical knowledge of techniques and treatments. Large scale randomized studies involving multiple institutions as well as exchange of informations and ideas are needed for further development of hypothermia treatment.

Animals↗

PubMed1998

Possible involvement of activator protein-1 DNA binding in mechanisms underlying ischemic tolerance in the CA1 subfield of gerbil hippocampus.

Transcription factors are nuclear proteins with an ability to recognize particular nucleotide sequences on double stranded genomic DNAs and thereby modulate the activity of RNA polymerase II which is responsible for the formation of messenger RNAs in cell nuclei. Gel retardation electrophoresis revealed that transient forebrain ischemia for 5 min led to drastic potentiation of binding of a radiolabelled double-stranded oligonucleotide probe for the transcription factor activator protein-1, in the thalamus as well as the CA1 and CA3 subfields and the dentate gyrus of the hippocampus of the gerbils previously given ischemia for 2 min two days before, which is known to induce tolerance to subsequent severe ischemia in the CA1 subfield. By contrast, ischemia for 5 min resulted in prolonged potentiation of activator protein-1 binding in the vulnerable CA1 subfield of the gerbils with prior ischemia for 5 min 14 days before, which is shown to induce delayed death of the pyramidal neurons exclusively in this subfield. Similar prolongation was seen with activator protein-1 binding in the vulnerable thalamus but not in the resistant CA3 subfield and dentate gyrus of the gerbils with such repeated ischemia for 5 min. Limited proteolysis by Staphylococcus aureus V8 protease as well as supershift assays using antibodies against c-Fos and c-Jun proteins demonstrated the possible difference in constructive partner proteins of activator protein-1 among nuclear extracts of the CA1 subfield obtained from gerbils with single, tolerated and repeated ischemia. These results suggest that de novo protein synthesis may underlie molecular mechanisms associated with acquisition of the ischemic tolerance through modulation at the level of gene transcription by activator protein-1 composed of different constructive partner proteins in the CA1 subfield. Possible participation of glial cells in the modulation is also suggested in particular situations.

Animals↗

PubMed1998

Remarkable increase in nuclease resistance of plasmid DNA through supramolecular assembly with poly(ethylene glycol)-poly(L-lysine) block copolymer.

Supramolecular associates of DNA with Poly(ethylene glycol)-poly(L-lysine) block copolymers (PEG-PLL) were prepared in aqueous milieu, and the water solubility under charge-neutralized (stoichiometric) conditions was maintained. Associates thus prepared achieved remarkable resistance against nuclease attack, probably due to the formation of PEG palisade surrounding the ion-complexed core of the DNA with the PLL segment of the block copolymer. Further, to estimate the stability of the associate, the rate of interexchange reaction of DNA in the associate with poly(vinyl sulfonate) was evaluated from a method using a metachromasy of toluidine blue dye. Of interest, the nuclease resistance of DNA in PEG-PLL/DNA associate had an inverse correlation with the rate of interexchange reaction and, thus, progressively increased with an increase in the molecular weight of PLL segment in the block copolymer, indicating the substantial importance of block copolymer design for DNA stabilization.

Animals↗

PubMed1998

Antioxidative bromoindole derivatives from the mid-intestinal gland of the muricid gastropod Drupella fragum.

Three new bromoindoles, 6-bromo-5-hydroxyindole (1), 6-bromo-4, 5-dihydroxyindole (2), and 6-bromo-4,7-dihydroxyindole (3), were isolated from the midintestinal gland of the muricid gastropod Drupella fragum. The structures of 2 and 3 were elucidated mainly by NMR spectroscopic analyses of their acetyl derivatives, whereas the structure of 1 was established by spectroscopic methods and total synthesis. Antioxidative activity for compounds 1-3 was evaluated by the POV method, and compound 1 was found to have as strong an antioxidative potency as BHT.

Acetates↗

PubMed1998

Visualization of PEO-PBLA-pyrene polymeric micelles by atomic force microscopy.

PURPOSE: To directly visualize and evaluate the aqueous block copolymeric micelles, poly(ethylene oxide)-poly(beta-benzyl L-aspartate) (PEO-PBLA) chemically conjugated with pyrene fluorescence molecule, by nanotechnology of atomic force microscopy (AFM). METHODS: The block copolymers' PEO-PBLA-Pyrene was first synthesized by reacting with pyrene sulfonyl chloride and PEO-PBLA in tetrahydrofuran (THF) solution and were identified by GPC reflect index, UV and fluorescence detectors. The characterization of physical and chemical properties of PEO-PBLA-Pyrene polymeric micellar solution were examined by the dynamic light scattering (DLS) and critical micelles concentrations (CMC). In addition, the nanotechnology of AFM was used to directly visualize the size and shape of nanopolymeric micelles. RESULTS: The pyrene fluorescence molecule were successfully conjugated at the amino group of the end of PBLA chain by GPC with three different detectors. The size of the aqueous PEO-PBLA-Pyrene polymeric micelles was detected around 57 nm with unimodal distribution by DLS measurement. As a result of this finding, the CMC test was also found out that the fluorescence intensity was increasing around 0.01 approximately 0.05 mg/ml. Using AFM evaluation of polymeric micellar solution, the morphology of aqueous PEO-PBLA-Pyrene polymeric micelles was observed on round shape and with the narrow dispersity of size range 50 approximately 80 nm. CONCLUSIONS: The presence of PEO-PBLA copolymers with pyrene in an aqueous system formed in a spherical and nano range of polymeric micelles.

Micelles↗

PubMed1998

Characterization of a human genomic DNA fragment which rescues defective lipid-linked oligosaccharide synthesis in a mutant G258 cell line isolated from the FM3A mouse mammary carcinoma cell line.

The G258 mutant cell line, isolated from the FM3A mouse mammary carcinoma cell line, is temperature-sensitive for both cell growth and asparagine-linked glycosylation due to mutation at a single location. The biochemical defect in the G258 mutant resides in the formation of lipid-linked oligosaccharide, presumably in one of the steps of GDP-mannose-dependent mannosylation (Y. Nishikawa, J. Cell. Physiol. 119, 260-266, 1984; Y. Nishikawa, Biochim. Biophys. Acta 1091, 135-140, 1991). In the present study, we transfected human genomic DNA fragments into the G258 mutant by the radiation hybrid method and isolated transformants (KK-1, -3 and -4) which showed recovery from both temperature-sensitive cell growth and asparagine-linked glycosylation. These transformants contained a common Alu-containing human DNA fragment (1.3 kb) which will be used as a marker for isolating the gene that complements the defect of lipid-liked oligosaccharide synthesis in the G258 mutant.

Alu Elements↗

PubMed1998

Increased vascular permeability by a specific agonist of protease-activated receptor-2 in rat hindpaw.

The present study examined the effect of intraplantar (i.pl.) administration of a selective agonist of protease-activated receptor (PAR)-2, SLIGRL-NH2(PP6-NH2), on vascular permeability in rat hindpaw. PP6-NH2, administered i.pl. at 10-100 nmol per paw, enhanced vascular permeability and caused oedema formation in rat hindpaw. SLIGRL (PP6-OH) and trypsin, by i.pl. administration, also elicited an increase in vascular permeability, although i.pl. administration of the mixture of constituent amino acids of PP6-OH at an equivalent dose did not. The PP6-NH2-induced increase in vascular permeability was abolished by repeated pretreatment with compound 48/80 to deplete bioactive amines in mast cells. These findings suggest that the activation of PAR-2 induces acute inflammation, at least partially, via mast cell degranulation in rat hindpaw.

Administration, Topical↗

PubMed1998

Type 1 Cu structure of blue nitrite reductase from Alcaligenes xylosoxidans GIFU 1051 at 2.05 A resolution: comparison of blue and green nitrite reductases.

The crystal structure of the blue nitrite reductase from Alcaligenes xylosoxidans GIFU 1051 (AxgNIR) has been determined at 2.05 A resolution. AxgNIR contains both type 1 and 2 Cu sites, the geometry of the former being distorted tetrahedral. The superpositioning of the type 1 Cu sites in the blue enzyme and a green nitrite reductase revealed that the orientation of the Met150 side chain differed. The deviation of the Sdelta(Met150) atom from the axial position of the NNS plane formed by two Ndelta(His95 and His145) and one Sgamma(Cys136) atom caused the difference in the colors of the enzymes, i.e. blue and green.

Alcaligenes↗

PubMed1998

Postischemic enhancements of N-methyl-D-aspartic acid (NMDA) and non-NMDA receptor-mediated responses in hippocampal CA1 pyramidal neurons.

Glutamate receptor-mediated responses were investigated by using a whole-cell recording and an intracellular calcium ion ([Ca2+]i) imaging in gerbil postischemic hippocampal slices prepared at 1, 3, 6, 9, 12, and 24 hours after 5-minute ischemia. Bath application of N-methyl-D-aspartic acid (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA), and kainate showed that NMDA-, AMPA- and kainate-induced currents were enhanced in postischemic CA1 pyramidal neurons at 1 to 12 hours after 5-minute ischemia. NMDA and non-NMDA receptor-mediated excitatory postsynaptic currents (EPSC) were examined in postischemic CA1 pyramidal neurons at 3 hours after 5-minute ischemia to confirm whether synaptic responses are enhanced in the postischemic CA1 pyramidal neurons. The amplitudes of NMDA- and non-NMDA-receptor-mediated EPSC were enhanced in the postischemic CA1 pyramidal neurons. NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were also examined to determine whether the enhancement of currents is accompanied by the enhancement of [Ca2+]i elevation. The enhancements of NMDA-, AMPA-, and kainate-induced [Ca2+]i elevations were shown in the postischemic CA1. These results indicate that NMDA and non-NMDA receptor-mediated responses are persistently enhanced in the CA1 pyramidal neurons 1 to 12 hours after transient ischemia, and suggest that the enhancement of glutamate receptor-mediated responses may act as one of crucial factors in the pathologic mechanism responsible for leading postischemic CA1 pyramidal neurons to irreversible neuronal injury.

Animals↗