Conversion of endoscopic nasobiliary drainage to internal drainage by means of endoscopic scissor forceps.
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Bile Duct Neoplasms↗
Biomedical subjects
K Tsutsui
Publications and source records attributed to K Tsutsui.
cDNA cloning and expression of mutant catalase from the hypocatalasemic mouse: comparison with the acatalasemic mutant.
Mutant catalase cDNAs from the hypocatalasemic and acatalasemic mice were cloned and expressed in bacteria. A novel missense mutation, Asp (AAT) to Ser (AGT), was identified at amino acid position 439 of the hypocatalasemic catalase. Analysis of recombinant catalase mutants revealed that the mutation is responsible for the reduced activity of hypocatalasemic catalase and the unstable tetrameric structure of acatalasemic catalase was also suggested.
Acatalasia↗
Detecting genotypes of catalase mutant mice by genomic polymerase chain reaction and restriction analysis.
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Animals↗
Activity and localization of 3beta-hydroxysteroid dehydrogenase/ Delta5-Delta4-isomerase in the zebrafish central nervous system.
Little information is available for neurosteroidogenesis in the central nervous system (CNS) of lower vertebrates. Therefore, in the present study, we examined the enzymatic activity and localization of 3beta-hydroxysteroid dehydrogenase/Delta5-Delta4-isomerase (3betaHSD), a key steroidogenic enzyme, in the CNS of adult male zebrafish to clarify central progesterone biosynthesis. Biochemical studies together with HPLC analysis revealed that the zebrafish brain converted pregnenolone to progesterone, suggesting the enzymatic activity of 3betaHSD. This conversion was significantly reduced by trilostane, a specific inhibitor of 3betaHSD. By using Western immunoblotting with the polyclonal antiserum directed against purified bovine adrenal 3betaHSD, a 3betaHSD-like substance was found in homogenates of the zebrafish brain. Immunocytochemical analysis was then undertaken to investigate the localization of the 3betaHSD-like substance in the zebrafish brain and spinal cord. Clusters of immunoreactive cell bodies were localized in the dorsal telencephalic areas (D), central posterior thalamic nucleus (CP), preoptic nuclei (NPO), posterior tuberal nucleus (PTN), paraventricular organ (PVO), and nucleus of medial longitudinal fascicle (NMLF). 3betaHSD-like immunoreactivity was also observed in somata of cerebellar Purkinje neurons. A widespread distribution of immunoreactive fibers was found throughout the brain and spinal cord. In addition, positively stained cells were restricted to other organs, such as the pituitary and retina. Preabsorbing the antiserum with purified bovine adrenal microsome resulted in a complete absence of 3betaHSD-like immunoreactivity. These results suggest that the fish CNS possesses steroidogenic enzyme 3betaHSD and produces progesterone. The present study further provides the first immunocytochemical mapping of the site of 3betaHSD expression in the fish CNS.
3-Hydroxysteroid Dehydrogenases↗
Large third-order optical nonlinearity of Cu-O chains investigated by third-harmonic generation spectroscopy.
Spectra of the third-order nonlinear susceptibility chi((3)) have been investigated for one-dimensional Mott insulators, Sr(2)CuO(3) and Ca(2)CuO(3), by applying the third-harmonic generation (THG) spectroscopy on their single-crystalline thin films. The three-photon resonance to the lowest charge-transfer (CT) state with odd parity strongly enhances chi((3)), which is of the order of 10(-9) esu. The two-photon resonant structure unravels the even-CT state, located close to the odd-CT state. Two types of chi((3)) spectra obtained from THG and the electroreflectance measurements are explained based on the concept of spin-charge separation.
Journal Article↗
Changes in central steroid receptor expression, steroid synthesis, and dopaminergic activity related to the reproductive cycle of the ring dove.
This review examines possible neural mechanisms involved in the expression of parental behavior in the ring dove, Streptopelia risoria. This avian species has proved an excellent animal model for studies concerning endocrine-behavior interactions for many years. Studies were performed to localize the expression of central androgen and progesterone receptor in both sexes. Expression of androgen receptor (androgen receptor immunoreactivity, AR-ir) was widespread but increased, similarly in both sexes, with increasing day-length. Progesterone receptor-immunoreactivity (PR-ir) was more localized in several discrete areas of the hypothalamus. Similarly, no sex differences were observed in PR-ir, and expression increased in birds maintained on long days. AR-ir demonstrated dramatic changes over the breeding cycle, being greatest in courting birds and almost undetectable in parenting birds of both sexes brooding their young. PR-ir showed a differential expression over the breeding cycle relative to its hypothalamic localization. PR-ir decreased in the tuberal hypothalamic area in brooding birds of both sexes; whereas in the preoptic area, PR-ir was maintained. Significant increases in dopaminergic activity during the parenting phase of the breeding cycle occurred in specific neural regions including the PVM and DMA. Studies demonstrated the ability of the diencephalon of both sexes of the ring dove brain to synthesize progesterone, with indications that in the male brooding dove, synthesis is increased. Finally, a model is presented that proposes a mechanism whereby these central systems may interact to result in the expression of full parental behavior in both sexes of the ring dove.
Animals↗
Plasmids with a mammalian replication origin and a matrix attachment region initiate the event similar to gene amplification.
Gene amplification plays a crucial role in the development of many human malignancies. Amplified genes are frequently localized on double minutes (DMs). We show here that plasmids bearing both a mammalian replication origin and a nuclear matrix attachment region were able to integrate into DMs if transfected to cells having DMs (COLO 320DM). Furthermore, these plasmids triggered the events leading to the de novo formation of the structure similar to DMs if transfected to the cells without DMs (COLO 320HSR or HeLa). Autonomous replication of these plasmids was suggested to be a prerequisite for these events to occur, because the presence of the origin sequences in the plasmids was required. The presence of matrix attachment region in the plasmids is also required for these events to occur, suggesting that matrix attachment plays an indispensable role in extrachromosomal replication. This model system will allow us to investigate the mechanism of gene amplification as well as to analyze the autonomous replication of the plasmid with mammalian replication origins.
Colorectal Neoplasms↗
Expression of amphiphysin I in Sertoli cells and its implication in spermatogenesis.
Amphiphysin I is a protein concentrated in nerve terminals and involved in the endocytosis of synaptic vesicle membrane. We show here that amphiphysin I is expressed in the rat testis, localized exclusively in the Sertoli cells. In the postnatal testicular development, expression of amphiphysin I was not evident at birth, but became significant at postnatal day 15 (P15), coinciding with the onset of spermatogenesis. The expression level of amphiphysin I increased 10-fold between P15 and P25 to reach the adult level. In adult testes reversibly damaged by ethane dimethane sulphonate administration, expression of amphiphysin I did not change following the damage, whereas the protein was transiently converted into its phosphorylated form. The increase in levels of phosphorylated amphiphysin I was closely associated with the severe histological damage to germ cells. The present findings suggest that amphiphysin I in Sertoli cells is involved in spermatogenesis, probably through endocytic processes.
Animals↗
Effects of progesterone synthesized de novo in the developing Purkinje cell on its dendritic growth and synaptogenesis.
De novo steroidogenesis from cholesterol is a conserved property of vertebrate brains, and such steroids synthesized de novo in the brain are called neurosteroids. The identification of neurosteroidogenic cells is essential to the understanding of the physiological role of neurosteroids in the brain. We have demonstrated recently that neuronal neurosteroidogenesis occurs in the brain and indicated that the Purkinje cell actively synthesizes several neurosteroids de novo from cholesterol in vertebrates. Interestingly, in the rat, this neuron actively synthesizes progesterone de novo from cholesterol only during neonatal life, when cerebellar cortical formation occurs most markedly. Therefore, in this study, the possible organizing actions of progesterone during cerebellar development have been examined. In vitro studies using cerebellar slice cultures from newborn rats showed that progesterone promotes dose-dependent dendritic outgrowth of Purkinje cells but dose not affect their somata. This effect was blocked by the anti-progestin RU 486 [mifepristone; 17beta-hydroxy-11beta-(4-methylaminophenyl)-17alpha-(1-propynyl) estra-4,9-dien-3 one-6-7]. In vivo administration of progesterone to pups further revealed an increase in the density of Purkinje spine synapses electron microscopically. In contrast to progesterone, there was no significant effect of 3alpha,5alpha-tetrahydroprogesterone, a progesterone metabolite, on Purkinje cell development. Reverse transcription-PCR-Southern and immunocytochemical analyses showed that intranuclear progesterone receptors were expressed in Purkinje cells. These results suggest that progesterone promotes both dendritic outgrowth and synaptogenesis in Purkinje cells through intranuclear receptor-mediated mechanisms during cerebellar development. Such organizing actions may contribute to the formation of the cerebellar neuronal circuit.
Animals↗
High incidence of antinuclear antibodies that recognize the matrix attachment region.
The matrix attachment region (MAR) is a distinctive genomic DNA involved in a variety of nuclear processes through association with the nuclear matrix. Recent studies suggest that nuclear matrix is altered in the process of apoptosis and presented to the immune system, leading to the production of autoantibodies against its protein components. To see whether MARs are also recognized by autoantibodies, a collection of human sera containing antinuclear antibodies was screened for the presence of binding activities against cloned MARs. We found that MAR-binding activities are quite common in these sera. There was a positive correlation among the MAR-binding titers for three different MAR probes. As expected, the MAR-binding activity was copurified with serum IgG, and subclass analysis with affinity-purified IgG on MAR-Sepharose showed a predominance of IgG2 isotype. Several lines of evidence implied that the anti-MAR antibodies detected here is distinct from the ordinary anti-DNA antibodies that are reactive to bulk DNA.
Antibodies, Antinuclear↗
Expression and localization of cytochrome P450 17 alpha-hydroxylase/c17,20-lyase in the avian brain.
Steroids synthesized de novo from cholesterol in the brain are generally called neurosteroids. We have recently demonstrated, using biochemical and molecular biological methods, that certain structures in the quail brain possess cytochrome P450 side-chain cleavage enzyme (P450scc) and 3beta-hydroxysteroid dehydrogenase/Delta(5)-Delta(4)-isomerase (3beta-HSD) and produce pregnenolone, pregnenolone sulfate and progesterone. To clarify the biosynthetic pathway of neurosteroids in the avian brain, therefore, we examined the expression of messenger RNA (mRNA) encoding for the enzyme cytochrome P450 17alpha-hydroxylase/c17,20-lyase (P450(17alpha,lyase)), which converts pregnenolone to dehydroepiandrosterone via 17alpha-hydroxypregnenolone or progesterone to androstenedione via 17alpha-hydroxyprogesterone. RT-PCR analysis followed by Southern hybridization indicated the expression of P450(17alpha,lyase) mRNA in the brain of sexually mature birds without a clear-cut sex difference. Employing biochemical techniques combined with HPLC analysis, the conversion of progesterone to 17alpha-hydroxyprogesterone was also found in brain slices of the mature male. P450(17alpha,lyase) mRNA was detected in various brain regions, but there was a clear regional difference in the expression. The expressions of P450(17alpha,lyase) mRNA in the diencephalon and mesencephalon were significantly higher than those in the cerebrum and cerebellum, unlike 3beta-HSD mRNA, which showed no regional difference in the expression. In situ hybridization revealed the cellular localization of P450(17alpha,lyase) mRNA. The cells expressing P450(17alpha,lyase) mRNA were detected several diencephalic and mesencephalic regions, such as the preoptic area, the anterior hypothalamus, the dorsolateral thalamus, the optic tectum and the ventral midbrain. The expression was also localized in the septum, the hyperstriatum accessorium, and the ventral portions of the archistriatum in the telencephalon. Cerebellar Purkinje cells also expressed P450(17alpha,lyase) mRNA. These results suggest that the avian brain possesses P450(17alpha,lyase) as well as P450scc and 3beta-HSD in both sexes. The expression of P450(17alpha,lyase) in the avian brain may be region-dependent.
17-Hydroxysteroid Dehydrogenases↗
Developmental changes in progesterone biosynthesis and metabolism in the quail brain.
We have recently demonstrated that the quail brain possesses the cholesterol side-chain cleavage enzyme (cytochrome P450scc) and 3beta-hydroxysteroid dehydrogenase/Delta5-Delta4-isomerase (3beta-HSD) and produces pregnenolone, pregnenolone sulfate and progesterone from cholesterol. To elucidate the developmental changes in progesterone biosynthesis and its metabolism in the quail brain, we examined the expression and activity of 3beta-HSD and progesterone metabolite(s) during embryonic and post-hatched ages. Both the progesterone concentration and 3beta-HSD mRNA expression in the brain were almost constant during embryonic and post-hatched ages. The conversion of pregnenolone to progesterone (net 3beta-HSD enzymatic activity) was also constant during development and at maturity. However, without radioinert progesterone, the production of progesterone was drastically reduced in the embryonic brain, indicating active progesterone metabolism at the embryonic stage. Biochemical analysis together with HPLC and TLC revealed that only the embryonic brain actively produced 5beta-dihydroprogesterone from progesterone. Thus, progesterone production may be constant during embryonic and post-hatched development and in adulthood, whereas 5beta-dihydroprogesterone may be produced actively only in embryonic life due to 5beta-reductase.
Aging↗
Distribution of novel RFamide-related peptide-like immunoreactivity in the mouse central nervous system.
Recently, novel mammalian RFamide-related peptides (RFRPs) have been identified and suggested to increase prolactin release in the rodent. To assess possible functions of RFRPs, we investigated the distribution of RFRPs in the mouse central nervous system by immunochemical analyses. The quantitative analysis revealed that the concentration of RFRP-like substances was much higher in the brainstem and spinal cord than in other regions. Immunohistochemistry showed that RFRP-like immunoreactive perikarya were localized in the dorsomedial hypothalamic nucleus, the lateral superior olive, and the nucleus of the solitary tract. The dense networks of immunoreactive fibers were found in the lateral parabrachial nucleus, the lateral reticular nucleus, and the superficial layer of spinal trigeminal nucleus and dorsal horn of the spinal cord. Thus, RFRPs may participate not only in neuroendocrine functions but also in behavioral, sensory, and autonomic functions.
Animals↗
Immunohistochemical analyses of DNA topoisomerase II isoforms in developing rat cerebellum.
In mammalian cells, two isoforms of DNA topoisomerase II (topo IIalpha and topo IIbeta) have been identified. Topo IIalpha is essential in mitotic cells, whereas the function of topo IIbeta remains unclear. In the present study, we investigated the developmental control of topo II isoforms in two different neuronal lineages, cerebellar Purkinje cells and granule cells, by immunohistochemical analysis with isoform-specific monoclonal antibodies. As expected, proliferating cells in the neuroepithelium and in the external germinal layer (EGL) were topo IIalpha immunopositive. The migrating as well as differentiating Purkinje cells and granule cells showed an enhanced topo IIbeta immunoreactivity. The postmitotic granule cells in the postnatal EGL showed an abrupt transition of expressed topo II isoforms from IIalpha to IIbeta. The transition was clearly coincident with the completion of final cell division and the initiation of terminal differentiation because no increase of the topo IIbeta immunoreactivity was observed in the spreading EGL cells that are still in the cell division cycle. The topo IIbeta signal was detected in both nucleoplasm and nucleolus of differentiating cells. However, the nucleoplasmic signal decreased significantly as the cells reached terminal differentiation. The residual topo IIbeta in nucleoli was shown to occupy an unique location with respect to other nucleolar proteins, nucleolin and DNA topoisomerase I. Our findings indicate that both Purkinje cells and granule cells express the topo II isoforms in a similar timing during the cerebellar development and also suggest that topo IIbeta localized in nucleoplasm is the functional entity involved in neuronal differentiation.
Age Factors↗
Characterization of a cDNA encoding a novel avian hypothalamic neuropeptide exerting an inhibitory effect on gonadotropin release.
We previously isolated a novel dodecapeptide containing a C-terminal -Arg-Phe-NH(2) sequence, SIKPSAYLPLRF-NH(2) (RFamide peptide), from the quail brain. This quail RFamide peptide was shown to decrease gonadotropin release from the cultured anterior pituitary and to be located at least in the quail hypothalamo-hypophysial system. We therefore designated this RFamide peptide gonadotropin inhibitory hormone (GnIH). In the present study we characterized the GnIH cDNA from the quail brain by a combination of 3' and 5' rapid amplification of cDNA ends ('RACE'). The deduced GnIH precursor consisted of 173 amino acid residues, encoding one GnIH and two putative gene-related peptide (GnIH-RP-1 and GnIH-RP-2) sequences that included -LPXRF (X=L or Q) at their C-termini. All these peptide sequences were flanked by a glycine C-terminal amidation signal and a single basic amino acid on each end as an endoproteolytic site. Southern blotting analysis of reverse-transcriptase-mediated PCR products demonstrated a specific expression of the gene encoding GnIH in the diencephalon including the hypothalamus. Furthermore, mass spectrometric analyses detected the mass numbers for matured GnIH and GnIH-RP-2, revealing that both peptides are produced from the precursor in the diencephalon as an endogenous ligand. Taken together, these results lead to the conclusion that GnIH is a hypothalamic factor responsible for the negative regulation of gonadotropin secretion. Furthermore, the presence of a novel RFamide peptide family containing a C-terminal -LPXRF-NH(2) sequence has been revealed.
Amino Acid Sequence↗
Comparison of chromatographic and spectroscopic methods used to rank compounds for aqueous solubility.
Rapid methods for ranking the solubility of compounds in aqueous media using commercial, 96-well ultraviolet-visible (UV-vis) and nephelometric plate readers are described. The methods were evaluated using commercially available compounds from a variety of structural classes as well as a series of structurally related compounds derived from combinatorial synthesis. Samples were predissolved in dimethyl sulfoxide (DMSO) and then added to the study solvent to attain a final concentration of DMSO in the aqueous solution of 5%. Comparison of filtration of the samples through nylon and poly(tetrafluoroethylene) (PTFE) membranes is also described. The solubility of the compounds determined using the UV-vis plate reader in the absorption mode (with samples filtered with the PTFE filter) as well as in the light scattering mode was in good agreement with that determined by high-performance liquid chromatography, with an average correlation of 0.95. Solubility data obtained using a 96-well nephelometer was also comparable (r(2) = 0.97). The nonequilibrium methods described in this study can be used to rapidly rank compounds from combinatorial libraries for solubility and can also give a general assessment of solubility prior to running additional high throughput screens in a drug discovery environment.
Chromatography, High Pressure Liquid↗
Cortical areas related to attention to 3D surface structures based on shading: an fMRI study.
The aim of the present study was to determine which cortical areas are activated in relation to attention to a three-dimensional (3D) structure of a surface based on shading. Cortical activities were examined using functional magnetic resonance imaging while subjects discriminated whether the central part of the surface protruded or was recessed based on shading without any binocular disparity cues. Relatively broad cortical areas including both dorsal and ventral visual pathways were recruited when shading was used as a crucial cue for the perception of the 3D structure of a surface. In these cortical areas, however, the right intraparietal area was shown to be commonly activated in all subjects and in all sessions by multisubject conjunction analysis. These results strongly suggest that the intraparietal area plays an important role in perception of the 3D structure of a surface, even when based only on monocular depth cues without binocular disparity cues.
Adult↗
Responses to task-irrelevant visual features by primate prefrontal neurons.
The primate brain is equipped with prefrontal circuits for interpreting visual information, but how these circuits deal with competing stimulus-response (S-R) associations remains unknown. Here we show different types of responses to task-irrelevant visual features in three functionally dissociated groups of primate prefrontal neurons. Two Japanese macaques participated in a go/no-go task in which they had to discriminate either the color or the motion direction of a visual target to make a correct manual response. Prior to the experiment, the monkeys had been trained extensively so that they acquired fixed associations between visual features and required responses (e.g., "green = go"; "downward motion = no-go"). In this design, the monkey was confronted with a visual target from which it had to extract relevant information (e.g., color in the color-discrimination condition) while ignoring irrelevant information (e.g., motion direction in the color-discrimination condition). We recorded from 436 task-related prefrontal neurons while the monkey performed the multidimensional go/no-go task: 139 (32%) neurons showed go/no-go discrimination based on color as well as motion direction ("integration cells"); 192 neurons (44%) showed go/no-go discrimination only based on color ("color-feature cells"); and 105 neurons (24%) showed go/no-go discrimination only based on motion direction ("motion-feature cells"). Overall, however, 162 neurons (37%) were influenced by irrelevant information: 53 neurons (38%) among integration cells, 71 neurons (37%) among color-feature cells, and 38 neurons (36%) among motion-feature cells. Across all types of neurons, the response to an irrelevant feature was positively correlated with the response to the same feature when it was relevant, indicating that the influence from irrelevant information is a residual from S-R associations that are relevant in a different context. Temporal and anatomical differences among integration, color-feature and motion-feature cells suggested a sequential mode of information processing in prefrontal cortex, with integration cells situated toward the output of the decision-making process. In these cells, the response to irrelevant information appears as a congruency effect, with better go/no-go discrimination when both the relevant and irrelevant feature are associated with the same response than when they are associated with different responses. This congruency effect could be the result of the combined input from color- and motion-feature cells. Thus these data suggest that irrelevant features lead to partial activation of neurons even toward the output of the decision-making process in primate prefrontal cortex.
Animals↗