Biomedical subjects

Y Takeda

Publications and source records attributed to Y Takeda.

At least 163 records · Page 9Linked to original sources

PubMed2000

Angiotensin II stimulates cyclic ADP-ribose formation in neonatal rat cardiac myocytes.

To examine the role of cyclic ADP-ribose (cADP-ribose) as a second messenger downstream of angiotensin II (Ang II) receptor activation in the heart, ADP-ribosyl cyclase activity was measured in a crude membrane fraction of ventricular myocytes. Ang II at 10-100 nM increased ADP-ribosyl cyclase activity by 40-90% in the ventricular muscle of neonatal (2-4-day-old) rats, but not in fetal or adult hearts. This increase was inhibited by the Ang II antipeptide. Stimulation of ADP-ribosyl cyclase was reproduced by GTP and guanosine 5'-[gamma-thio]triphosphate, and prevented by guanosine 5'-[beta-thio]diphosphate. Prior treatment of the rats with cholera toxin A and B subunits also blocked the Ang II-induced activation. The density of Ang II receptors detected as [(3)H]Ang II binding was higher in neonatal than adult rats. These results demonstrate the existence of a signalling pathway from Ang II receptors to membrane-bound ADP-ribosyl cyclase in the ventricular muscle cell and suggest that the Ang II-induced increase in cADP-ribose synthesis is involved in the regulation of cardiac function and development.

Adenosine Diphosphate Ribose↗

PubMed2000

Total synthesis of the antitumor antibiotic (+/-)-fredericamycin A by a linear approach.

A linear approach to the total synthesis of racemic fredericamycin A (1) through the oxidative intramolecular [4 + 2] cycloaddition of a (phenylthio)acetylene-cobalt complex is described, which is applicable for the asymmetric total synthesis of naturally occuring 1. The highlight of this work is the aromatic Pummerer-type reaction with 1-ethoxyvinyl chloroacetate, which effects the introduction of the oxygen functional group to the internal B-ring of the highly functionalized, congested polyaromatic ABC-ring moiety.

Antibiotics, Antineoplastic↗

PubMed2000

DNA ligase IV and XRCC4 form a stable mixed tetramer that functions synergistically with other repair factors in a cell-free end-joining system.

Repair of DNA double-strand breaks in mammalian cells occurs via a direct nonhomologous end-joining pathway. Although this pathway can be studied in vivo and in crude cell-free systems, a deeper understanding of the mechanism requires reconstitution with purified enzymes. We have expressed and purified a complex of two proteins that are critical for double-strand break repair, DNA ligase IV (DNL IV) and XRCC4. The complex is homogeneous, with a molecular mass of about 300,000 Da, suggestive of a mixed tetramer containing two copies of each polypeptide. The presence of multiple copies of DNL IV was confirmed in an experiment where different epitope-tagged forms of DNL IV were recovered simultaneously in the same complex. Cross-linking suggests that an XRCC4.XRCC4 dimer interface forms the core of the tetramer, and that the DNL IV polypeptides are in contact with XRCC4 but not with one another. Purified DNL IV.XRCC4 complex functioned synergistically with Ku protein, the DNA-dependent protein kinase catalytic subunit, and other repair factors in a cell-free end-joining assay. We suggest that a dyad-symmetric DNL IV.XRCC4 tetramer bridges the two ends of the broken DNA and catalyzes the coordinate ligation of the two DNA strands.

Animals↗

PubMed2000

Human NB-2 of the contactin subgroup molecules: chromosomal localization of the gene (CNTN5) and distinct expression pattern from other subgroup members.

NB-2 is one of the neural recognition molecules in the contactin subgroup, which belongs to the immunoglobulin superfamily. In rat, the six molecules in this subgroup that have been reported to date are contactin, TAG-1, BIG-1, BIG-2, NB-2, and NB-3. We have isolated cDNAs encoding the two splicing isoforms of human NB-2. The long isoform of human NB-2 consists of 1100 amino acids residues that are 91% homologous to rat NB-2 at the amino acid sequence level. The short isoform lacks 74 amino acid residues between residues 19 and 93 of the long isoform. Among various regions of the adult human brain, high-level expression of NB-2 was detected in the amygdala and occipital lobe, whereas expression was low in the corpus callosum, caudate nucleus, and spinal cord. Although there were some differences, the expression pattern of NB-2 was the most similar to that of BIG-1 in the brain. Likewise, contactin and BIG-2 exhibited similar expression patterns. The expression of TAG-1 showed the least regional differences. The human NB-2 gene (CNTN5) was mapped to chromosome 11q21-q22.2 by fluorescence in situ hybridization. Our results suggest that the NB-2 gene may contribute to human neurological disorders.

Alternative Splicing↗

PubMed2000

A case of infantile neuroblastoma with intramucosal metastasis in a paranasal sinus.

An 8-year-old boy with neuroblastoma of the right adrenal gland is reported. His initial treatment included chemotherapy and surgery, with complete response (CR) being achieved at the initial site. A metastatic lesion was found in the right maxillary sinus 32 months after his initial treatment. A mass in the right soft palate was detected and was clinically suspected of being a metastasis. The results of biopsy were negative and the differential diagnosis from the imaging studies of CT included odontogenic disease, fungal infection, paranasal sinus cyst or hematoma, and benign tumors. Open transantral biopsy was done under general anesthesia, revealing severe inflammation in the right maxillary sinus as well as bone erosion. The histopathological diagnosis was metastatic neuroblastoma from the adrenal lesion. The local field was irradiated with 20 Gy of linear accelerator (linac) radiation, then the local field was eradicated. Extensive skeletal metastases were subsequently found by bone scintigraphy. Despite further treatment his general condition deteriorated rapidly and he died 24 months after starting treatment. We review the previous reports and discuss metastasis to the sinuses.

Adrenal Gland Neoplasms↗

PubMed2000

The O139 serogroup of Vibrio cholerae comprises diverse clones of epidemic and nonepidemic strains derived from multiple V. cholerae O1 or non-O1 progenitors.

Sixty-four representative strains of Vibrio cholerae O139 were analyzed, to re-examine the origin of this serogroup. Ribotyping differentiated the strains into 3 HindIII and 7 BglI ribotypes. One HindIII and 5 BglI ribotypes were shared by all toxigenic O139 strains. Of 6 nontoxigenic O139 strains, 3 shared ribotypes with the toxigenic strains, carried genes encoding toxin coregulated pilus, and were susceptible to the cholera toxin-converting bacteriophage CTXPhi. The remaining 3 strains belonged to 2 different ribotypes distinct from toxigenic O139 strains and were resistant to CTXPhi and JA-1, an O139-specific lytic bacteriophage. Polymerase chain reaction amplicons corresponding to the gmhD gene carried by these 3 strains also differed from those of the toxigenic O139 strains but were identical to those of 15 environmental non-O1-non-O139 strains. Thus, the O139 antigen is present in different lineages, and this serogroup appears to comprise epidemic and nonepidemic strains derived separately from different progenitors.

Cholera↗

PubMed2000

Dynamic changes of NADH fluorescence images and NADH content during spreading depression in the cerebral cortex of gerbils.

To elucidate the changes in the mitochondrial redox state during spreading depression (SD), tissue NADH content was measured in 20 anesthetized gerbils by the enzymatic cycling assay in a small cortical region (0.30+/-0.07 mg) where the direct-current (DC)-potential was measured. Sequential imaging of NADH fluorescence with a CCD camera and continuous monitoring of DC-potential and regional CBF were also performed in another 5 gerbils. Biphasic fluorescence waves propagating at the rate of 3 mm/min were observed using the CCD camera. An initial narrow (1.6+/-0.4 mm) wave, which showed a modest increase in fluorescence (108+/-6.4%), was observed simultaneously with the onset of negative DC-deflection. During depolarization, CBF was unchanged and tissue NADH content increased to 25.3+/-7.9 micromol/kg brain, which was higher than the value in the sham-control (11.0+/-2.5 micromol/kg brain). At 30 s after the deflection, a subsequent wide (7.0+/-2.1 mm) wave, which showed a moderate decrease in fluorescence (87.1+/-5.7%), was observed simultaneously with the increase in CBF and repolarization in DC-potential. Then NADH fluorescence recovered along with normalization of CBF at 152.2+/-38.6 s after the onset of DC-deflection. Tissue NADH concentration sampled at 120 s after the deflection was 11.6+/-4.6 micromol/kg brain. Since NADH fluorescence is absorbed by hemoglobin, the initial increase and subsequent decrease in fluorescence seem to have been induced by increases in NADH content and CBF, respectively. These findings indicate that the mitochondrial redox state transiently inclines to the reduction side synchronous to the onset of DC-deflection and that it normalizes within 120 s after deflection.

Animals↗

PubMed2000

Polyglutamation of antifolates is not required for induction of extracellular release of adenosine or expression of their anti-inflammatory effects.

Methotrexate (MTX) exerts an anti-inflammatory effect, reportedly by enhancing the release of adenosine, through an accumulation of 5-amino-4-imidazolecarboxamide ribonucleotide (AICAR). To examine the role of polyglutamation in promoting anti-inflammatory effects by antifolates, we tested whether a new antifolate designed to be resistant to intracellular polyglutamation (MX-68) exhibited anti-inflammatory effects and stimulated adenosine release. Both MX-68 and MTX (at concentrations greater than 0.1 microM) increased the release of adenosine from Daudi cells in vitro. Inhibition of AICAR synthesis suppressed adenosine release by MX-68 and MTX. The anti-inflammatory effects of antifolates were estimated using the murine air pouch model, in which a BALB/c mouse was intraperitoneally injected with MX-68 or MTX once a week for 3 weeks. MX-68 (0.5 mg kg(-1) week(-1)), as well as MTX, inhibited infiltration of leukocytes into the air pouch. This inhibitory effect was suppressed in the presence of an adenosine A(2) receptor antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX). These results suggest that MX-68, like MTX, exerts anti-inflammatory effects through the accumulation of AICAR and release of adenosine. These results suggest that polyglutamation of antifolate is not required for expression of anti-inflammatory effects.

2-Aminoadipic Acid↗

PubMed2000

Immunohistochemical localization of substance P receptors in the midline glia of the developing rat medulla oblongata with special reference to the formation of raphe nuclei.

Immunohistochemical localization of the substance P receptor (SPR) was examined in the developing rat medulla oblongata, with special reference to the development of substance P (SP)-immunoreactive neurons which form the medullary raphe nuclei. During development, SPR immunoreactivity was detected in cells lying lateral to the medullary midline from embryonic day 13 (E13) to postnatal day 5 (P5). The SPR-positive cell bodies were located close to the fourth ventricle, and bore long processes extending to the ventral pial surface. This SPR immunoreactivity co-localized with staining for monoclonal antibody 1D11, a specific marker of immature astrocytes. Substance P (SP)-immunoreactive neurons were first detected at E14 in the ventrolateral part of the medulla. By E16 their number had increased and they were arrayed in two rows closely parallel to the SPR-immunoreactive processes of non-neuronal cells. By P1, two separate SP-immunoreactive cell clusters could be recognized at the midline, representing dorsally the nascent raphe pallidus and ventrally the raphe obscurus. In addition, many SP-immunoreactive fibers traveled rostrocaudally in the medulla oblongata, juxtaposed to the midline sheets of SPR-immunoreactive long processes. SPR-immunoreactive processes at the midline were also immunoreactive for S-100, a glia-specific calcium-binding protein that is known to promote axonal growth of raphe neurons. These results suggest that SPR-expressing immature glial cells at the medullary midline are involved in the development of SP-immunoreactive raphe neurons, both in the formation of the medullary raphe nuclei and in axon guidance and growth.

Animals↗

PubMed2000

Enterotoxin adjuvants have direct effects on T cells and antigen-presenting cells that result in either interleukin-4-dependent or -independent immune responses.

In an in vitro study, Escherichia coli heat-labile toxin (LT) was shown to directly affect activated CD4(+) T cells and support interleukin (IL)-5 production in IL-4-deficient (IL-4(-/-)) mice, whereas cholera toxin (CT) did not. Both LT and CT enhanced B7-2 expression on B cells and macrophages. These effects were not influenced by CD40-CD40 ligand cosignaling. Addition of LT- or CT-treated antigen-presenting cells to anti-CD3-triggered CD4(+) T cells resulted in the induction of T cell proliferative responses. Further, these responses were inhibited by anti-B7-2 monoclonal antibody. Cocultivation of CD4(+) T cells with LT- or CT-treated antigen-presenting cells and anti-CD3 enhanced Th1- and IL-4-mediated Th2-type cytokine production. The results from in vitro studies were supported by in vivo studies in IL-4(-/-) mice, in which LT induced mucosal IgA responses but CT did not. Thus, although both LT and CT induce mucosal adjuvant responses via IL-4-dependent Th2-type responses, LT also elicits Th1- and IL-4-independent Th2-type responses.

Animals↗

PubMed2000

Induction of unresponsiveness to islet xenograft by MMC treatment of graft and blockage of LFA-1/ICAM-1 pathway.

BACKGROUND: Induction of unresponsiveness to graft is one of major interest in xenotransplantation. Two different modalities [direct graft treatment by mitomycin C (MMC) and blockage of the lymphocyte function-associated antigen-1/intracellular adhesion molecule-1 (LFA-1/ICAM-1) pathway in recipients by species-specific mAbs] were tested for their ability to produce unresponsiveness to secondary islet xenografts. METHODS: Collagenase-digested WS (RT1k) rat islets, purified by Ficoll density gradient, were incubated for 30 min with MMC 10 microg/ml, cultured for 20 hr, and transplanted into the renal subcapsular space of streptozotocin-induced diabetic C57BL/6 (H-2b) mice. Recipient mice were divided into experimental groups according to anti-rat ICAM-1 and/or anti-mouse LFA-1 mAb treatment and transplantation of MMC-treated or nontreated islets. RESULTS: MMC pretreatment alone prolonged graft survival, with a mean survival time (MST) of 23.0+/-7.4 days, compared with that of cultured islets (12.4+/-2.7 days; P<0.01). MMC treatment of islets significantly augmented graft survival, compared with that of crude islet grafts under treatment with anti-donor ICAM-1 mAb (MST: >41.3+/-30 vs. 16.6+/-5.4 days, P<0.01), anti-recipient LFA-mAb (MST: >70.3+/-28.9 vs. 30.4+/-10.4 days, P<0.001), or both mAbs (MST: >88.1+/-24.1 vs. 23+/-7.4 days, P<0.0001). One of six, four of nine, and six of eight animals accepted MMC-treated islet xenografts over 100 days after treatment with anti-rat ICAM-1, anti-mouse LFA-1, or both mAbs treatments, respectively, whereas none of the animals accepted nontreated islets under the same treatment. When the mice bearing long-term functioning xenografts were challenged with the secondary graft from the original donor strain, the animals previously treated with anti-recipient LFA-1 and anti-donor ICAM-1 mAbs were more prone to accept it than animals given anti-recipient LFA-1 mAb alone (MST: 55.8+/-25.7 vs. 15+/-2.4 respectively; P<0.001), although they rejected the third-party xenograft and allograft acutely. CONCLUSIONS: In the xenogeneic islet transplantation model, MMC graft pretreatment and blockage of the ICAM-1/LFA-1 pathway constitute a potent protocol for inducing unresponsiveness to islet xenografts.

Animals↗

PubMed2000

Expression and regulation of a gene encoding neural recognition molecule NB-3 of the contactin/F3 subgroup in mouse brain.

NB-3 is a neural recognition molecule which is a member of contactin/F3 subgroup in the immunoglobulin superfamily. We report here the developmental expression pattern and localization of NB-3 mRNA in mouse brain, determination of the NB-3 gene organization and identification of the promoter region. We also describe a splicing isoform of mouse NB-3. Mouse NB-3 exhibited 96% identity with rat NB-3 at the amino acid sequence level. The splicing isoform lacked the amino acid residues between 62 and 78 of the original NB-3, which constituted a part of the first immunoglobulin-like domain. The expression of NB-3 mRNA was evident after birth, reaching a maximum at the postnatal seventh day, and declined thereafter in the cerebrum, whereas the mRNA increased in the cerebellum to adulthood. In situ hybridization demonstrated that NB-3 mRNA was preferentially expressed in the accessory olfactory bulb, layers II/III and V of the cerebral cortex, piriform cortex, anterior thalamic nuclei, locus coeruleus of the pons and mesencephalic trigeminal nucleus, and in Purkinje cells of the cerebellum. The mouse NB-3 gene consisted of 23 exons spanning more than 130kb. The overall organization of the gene was similar to those of the F11, axonin-1 and TAX-1 genes of the subgroup. By reporter gene analysis with the 5'-flanking region of the gene, we found a basal promoter activity in the 1.2kb fragment upstream of the putative transcription initiation site. This study provides a basis for elucidating the biological significance of the contactin/F3 subgroup molecules.

Alternative Splicing↗

PubMed2000

Analysis of hepatic oxidative stress status by electron spin resonance spectroscopy and imaging.

Real-time detection of free radicals generated within the body may contribute to clarify the pathophysiological role of free radicals in disease processes. Of the techniques available for studying the generation of free radicals in biological systems, electron spin resonance (ESR) has emerged as a powerful tool for detection and identification. This article begins with a review of spin trapping detection of oxygen-centered radicals using X-band ESR spectroscopy and then describes the detection of superoxide and hydroxyl radicals by the spin trap 5,5-dimethyl-1-pyrroline-N-oxide and ESR spectroscopy in the perfusate from isolated perfused rat livers subjected to ischemia/reperfusion. This article also reviews the current status of ESR for the in vivo detection of free radicals and in vivo imaging of exogenously administered free radicals. Moreover, we show that in vivo ESR-computed tomography with 3-carbamoyl-2,2,5, 5-tetramethylpyrrolidine-1-oxyl may be useful for noninvasive anatomical imaging and also for imaging of hepatic oxidative stress in vivo.

Animals↗

PubMed2000

Genomic diversity among Vibrio cholerae O139 strains isolated in Bangladesh and India between 1992 and 1998.

In order to assess the extent of genomic diversity among Vibrio cholerae O139 strains, restriction fragment length polymorphisms in two genetic loci, rrn and ctx, were studied. Analysis of 144 strains isolated from different regions of Bangladesh and India between 1992 and 1998 revealed the presence of at least six distinct ribotypes (B-I through B-VI) of which three were new ribotypes, and one of these was represented by a nontoxigenic O139 strain. Strains of ribotypes B-I through B-V shared 11 different CTX genotypes (A through K). Antimicrobial resistance patterns of the strains varied independently of their ribotypes and CTX genotypes. Results of this study suggest that V. cholerae O139 is undergoing rapid genetic changes leading to the origination of new variants, and temporal changes in antimicrobial resistance patterns may be contributing to the selection of different variants.

Bacterial Typing Techniques↗

PubMed2000

Macrophyllols A and B, two unusual novel sesquiterpene and monoterpene dimers from the bark of Inula macrophylla.

[structure: see text] Two novel sesquiterpene and monoterpene dimers, macrophyllols A (1) and B (2), were isolated from the bark of Inula macrophylla. Their structures were determined on the basis of spectral evidence (especially HREIMS and 2D NMR) as well as chemical transformation. The structure of macrophyllol A (1) was confirmed by X-ray analysis. The possible biosynthetic pathways of macrophyllols A (1) and B (2) are discussed.

Dimerization↗

PubMed2000

Protection by vascular endothelial growth factor against sinusoidal endothelial damage and apoptosis induced by cold preservation.

BACKGROUND: It is well known that sinusoidal endothelial cell (SEC) damage during cold preservation of liver tissue is closely involved in early graft failure. The objective of this study was to investigate the involvement of apoptosis in the SEC damage induced by cold preservation and to demonstrate the protective effect of vascular endothelial growth factor (VEGF) on SEC injury, including apoptotic changes. METHODS: Isolated SECs and liver tissue of Wistar rats were cold-preserved in University of Wisconsin (UW) solution, and the protective effect of VEGF was then investigated. Isolated SECs were cultured for 24 hr, and divided into the following 3 groups: Group A, in which the cells were cultured for an additional 27 hr, Group B, in which the cells were cold-preserved in UW solution for 3 hr, and then recultured for 24 hr, and Group C, in which 20 ng/ml of VEGF was added to both the culture medium and the UW solution of cells cultured according to the Group B protocol. Each group of SECs was morphologically examined using the phase contrast microscopic method and the transmission electron microscopic method (TEM), and quantitatively analyzed using the WST-1 assay. Rat livers were cold-preserved in UW solution and divided into the VEGF(+) group and the VEGF(-) group, depending on whether VEGF was added or not. Each group of livers were analyzed by scanning electron microscopic method (SEM) after 24 hr of preservation. The hyaluronic acid uptake rate (HUR) was also determined after 6 hr of preservation. After 24 hr of preservation and 6 hr of reperfusion, tissues were examined by TEM and by the terminal deoxynucleotidyl transferase d-uridine triphosphate nick end labeling (TUNEL) assay. RESULTS: The phase contrast microscopic method and the WST-1 assay showed a protective effect of VEGF against the injury to isolated SECs during cold preservation and subsequent reculturing. Apoptosis was detected immediately by TEM after isolation of SECs, and the number of apoptotic cells increased with the incubation time. This increase was accelerated after cold preservation. The scanning electron microscopic method and the hyaluronic acid uptake rate showed a protective effect of VEGF against SEC damage in the cold-preserved livers. In the liver tissue, the TEM and the TUNEL assay detected apoptosis of SECs only after cold preservation and subsequent reperfusion. VEGF suppressed the apoptosis of SECs induced by cold preservation in both isolated cells and liver tissue. CONCLUSIONS: We demonstrated that SEC damage in the cold preservation of liver tissue was caused mainly by apoptosis, which required subsequent reperfusion. Moreover, isolated SECs showed spontaneous occurrence of apoptotic changes during culture, and these changes were accelerated by the preceding cold preservation. This is the first report to demonstrate the apoptotic changes of SECs seen here were inhibited by VEGF.

Animals↗

PubMed2000

Diversity in the arrangement of the CTX prophages in classical strains of Vibrio cholerae O1.

This study reports the results of a molecular analysis of the CTX prophages in classical biotype strains of Vibrio cholerae O1 of clinical origin isolated between 1970 and 1979 in India. All strains were sensitive to group IV classical phage and polymyxin B but resistant to group 5 El Tor phage. These phenotypic traits are consistent to that exhibited by the classical biotype. PCR studies reconfirmed their biotype assignment and showed the presence of intact CTX prophages and the presence of the recently described toxin linked cryptic plasmid. Restriction fragment length polymorphism of rRNA genes and pulsed-field gel electrophoresis showed clonal diversity among the strains. The most notable observation was the finding that one strain (GP13) has three CTX prophages while another (GP147) has four CTX prophages. This is the first time heterogeneity is reported in the arrangement of the CTX prophages among classical strains of V. cholerae O1.

Bacteriophage Typing↗

PubMed2000

Mutational analysis of ATP7B and genotype-phenotype correlation in Japanese with Wilson's disease.

The gene ATP7B responsible for Wilson's disease (WD) produces a protein which is predicted to be a copper-binding P-type ATPase, homologous to the Menkes disease gene (ATP7A). Various mutations of ATP7B have been identified. This study aimed to detect disease-causing mutations, to clarify their frequency and distribution, to determine whether genotype correlates with phenotype, and to determine the rate of abnormal findings in heterozygotes for the WD gene. We analyzed 41 unrelated Japanese WD families, including 47 patients. Twenty-one mutations, including nine novel ones, were identified. 2871delC (15.9%), 1708-5T-->G (11. 0%), and Arg778Leu (13.4%) were the most common mutations. 2871delC was detected mainly in eastern Japan and 1708-5T-->G in western Japan. The homozygotes for the 1708-5T-->G, 2871delC, or Arg778Leu mutations did not show a correlation with their phenotypes. Ceruloplasmin and copper levels were abnormally low in 28.6% and 35. 0% of heterozygotes, respectively. When patients and their families are screened for WD, a high rate of abnormal laboratory data in heterozygotes must be taken into account.

Adenosine Triphosphatases↗