Biomedical subjects

M Okuda

Publications and source records attributed to M Okuda.

At least 55 records · Page 3Linked to original sources

PubMed2001

Cancer chemopreventive activity of odorine and odorinol from Aglaia odorata.

In the course of our continuing search for novel cancer chemo-preventive agents from natural sources, we have carried out a primary screening in vitro assay of the compounds isolated from Aglaia odorata. Consequently, aminopyrrolidine-diamides, odorine and odorinol, were obtained as active constituents. These compounds exhibited potent anti-carcinogenic effects in a two-stage carcinogenesis test of mouse skin induced by 7,12-dimethylbenz[a]anthracene (DMBA) as an initiator and 12-O-tetradecanoylphorbol-13-acetate (TPA) as a promoter. Further, both compounds showed remarkable inhibitory effects in two-stage mouse skin carcinogenesis models induced by nitric oxide (NO) donors such as (+/-)-(E)-methyl-2-[(E)-hydroxyimino]-5-nitro-6-methoxy-3-hexenamide (NOR-1) or peroxynitrite as an initiator and TPA as a promoter. From these results, it was concluded that odorine and odorinol inhibited both the initiation and promotion stages of two-stage skin carcinogenesis.

Animals↗

PubMed2001

Survey of tick infestation and tick-borne ehrlichial infection of dogs in Ishigaki Island, Japan.

Twelve (54.5%) of 22 free-roaming dogs in Ishigaki Island had tick infestation identified as Rhipicephalus sanguineus. There were 121 ticks recovered and consisted of 28 females, 58 males, 22 nymphs and 3 larvae. Infection of dogs possibly with canine ehrlichial pathogens was examined by both indirect immunofluorescence assay and polymerase chain reaction (PCR). Two dogs of the 13 examined were sero-positive for the human granulocytic ehrlichia agent, and one of two dogs was PCR positive for Ehrlichia platys. This dog had platelet numbers slightly lower than normal value, however, no morulae were found within platelet on peripheral blood smear stained with Giemsa.

Animals↗

PubMed2001

Urine-based enzyme-linked immunosorbent assay for the detection of Helicobacter pylori infection in children.

OBJECTIVE: Serology and (13)C-urea breath test have been widely used as noninvasive tests to detect Helicobacter pylori infection. However, easier collection of samples and lower costs are desirable for diagnosis of the individual patient or for use in epidemiologic studies. Our aim was to study the diagnostic accuracy of a recently developed urine-based enzyme-linked immunosorbent assay (ELISA) kit for the detection of H pylori-specific immunoglobulin G (IgG) antibodies in children. STUDY DESIGN: Specimens of serum and randomly voided urine were collected from 816 children (0-15 years old) and were analyzed using 2 serum-based ELISA kits and a urine-based ELISA kit, respectively. Based on results of serology, the sensitivity, specificity, and accuracy of the urine-based ELISA kit were evaluated. With regard to false-positive and false-negative results, urinary IgG concentrations and IgG/creatinine levels were studied. RESULTS: Both serum-based ELISAs were positive in 41 children and were negative in 666, who were enrolled in this study. The remaining 109 children were excluded because of disagreement between the results of the 2 serum-based ELISAs, including indeterminate values. Overall sensitivity, specificity, and accuracy of urine-based ELISA test compared with serology were 85.4%, 95.5%, and 94.9%, respectively. On positivity rates, the urine-based ELISA was closely coincident with the serum-based ELISA in each age group. There was no correlation between antibody levels detected by urine-based ELISA and each serum-based ELISA. Urinary IgG concentrations and IgG/creatinine levels were significantly higher in false-positives and were lower in false-negatives than in true-positives plus true-negatives for serology. Most of those with false-positive results had trace to moderate proteinuria. CONCLUSIONS: The urine-based ELISA is an alternative to serum-based ELISA for diagnosis of H pylori infection in children and should be suitable for large-scale epidemiologic studies concerning the organism. In children with proteinuria, results of the test should be interpreted with caution. It is possible that the urine-based ELISA method would be applicable to diagnosis of other infectious diseases.

Adolescent↗

PubMed2001

Inhibitory effect of 1,1,1-trichloroethane on calcium channels of neurons.

Peripheral neuropathy may remain for some time after 1,1,1-trichloroethane exposure. A variety of Ca2+ channels gives sensory neurons many kinds of transmitting sensory information. We measured calcium currents in sensory neurons from neonatal rat dorsal root ganglion using whole-cell patch-clamp recordings. Trichloroethane reversibly reduced the low-voltage-activated (LVA) and high-voltage-activated (HVA) calcium. The half-inhibitory concentration (IC50) of the HVA and LVA currents was 5.76 x 10(-3) M and 3.99 x 10(-3) M, respectively. The Hill coefficient of the HVA and LVA currents was 0.61 and 1.04, respectively. In assessing voltage dependence for activation and inactivation of calcium currents, only the HVA calcium current was inactivated at greater negative potentials. This may be one of the mechanisms to reduce HVA current. However, activation and inactivation of the LVA currents were not affected by trichlorothane, so inhibition of the LVA currents may have other mechanisms. Calcium currents are thought to be involved in the control of neuronal excitability and neurotransmitter release. The inhibitory effect of trichloroethane on calcium currents may be involved in trichloroethane-induced sensory discomfort.

Animals↗

PubMed2001

Results of hyperamplification of centrosomes in naturally developing tumors of dogs.

OBJECTIVE: To evaluate results of centrosome hyperamplification in naturally developing tumors of dogs. SAMPLE POPULATION: Tumor specimens from 9 dogs with tumors (rhabdomyosarcoma, osteosarcoma, chondrosarcoma, myxosarcoma, and mammary gland tumor) and 2 canine osteosarcoma cell lines. PROCEDURE: 3 antibodies for centrosome proteins (ie, anti-gamma-tubulin, anti-BRCA1, and anti-pericentrin) were used for immunohistochemical analysis. Double immunostaining for centrosomes was used to confirm the specificity of these antibodies for centrosomes. Mutational analysis of the canine p53 gene was carried out by polymerase chain reaction-single-strand conformation polymorphism analysis, and expression of canine MDM2 protein was evaluated by use of immunohistochemical analysis, using anti-MDM2 antibody. RESULTS: Immunohistochemical analysis of dog osteosarcoma cell lines with apparent aneuploidy revealed frequent hyperamplification of centrosomes in the osteosarcoma cell lines. Similar hyperamplified centrosomes were detected in the tumor tissues from all of the 9 tumors. The frequency of cells with hyperamplified centrosomes (3 to 20/cell) in each tumor tissue ranged from 9.50 to 48.1%, whereas centrosome hyperamplification was not observed in normal lymph nodes from these dogs. In 8 of the 9 tumors, mutation of p53 gene or overexpression of MDM2, or both, was detected. CONCLUSIONS AND CLINICAL RELEVANCE: Various types of naturally developing tumors in dogs often have hyperamplification of centrosomes associated with chromosome instability. Hyperamplification of centrosomes is a novel tumor marker for use in cytologic and histologic examinations of clinical specimens obtained from dogs.

Animals↗

PubMed2001

Telomere length and telomerase activity in canine mammary gland tumors.

OBJECTIVE: To measure telomere length and telomerase activity in naturally occurring canine mammary gland tumors. SAMPLE POPULATION: 27 mammary gland tumor specimens obtained during resection or necropsy and 12 mammary gland tissue specimens obtained from healthy (control) dogs. PROCEDURE: Telomere length in tissue specimens was measured by use of restriction endonuclease digestion and Southern blot analysis. Telomerase activity was measured by use of a telomeric repeat amplification protocol assay. RESULTS: Telomere length in mammary gland tumors ranged from 11.0 to 21.6 kilobase pairs (kbp; mean +/- SEM, 14.5+/-0.5 kbp) but did not differ among tumor types. Telomeres in mammary gland tumors were slightly shorter than in normal tissue specimens, but telomere length could not be directly compared between groups, because mean age of dogs was significantly different between groups. Age was negatively correlated with telomere length in control dogs but was not significantly correlated with length in affected dogs. Telomerase activity was detected in 26 of 27 mammary gland tumors and in 4 of 12 normal tissue specimens. However, telomerase activity and telomere length were not correlated in tumor specimens. CONCLUSIONS AND CLINICAL RELEVANCE: Telomere length is maintained in canine mammary gland tumors regardless of the age of the affected dog. Measurement of telomere length may be a useful tool for monitoring the in vivo effects of telomerase inhibitors in dogs with tumors.

Adenocarcinoma↗

PubMed2001

Aberrations of the p53 tumor suppressor gene in various tumors in dogs.

OBJECTIVE: To evaluate aberrations of the p53 tumor suppressor gene in naturally developing tumors in dogs. SAMPLE POPULATION: Tumor specimens from 15 dogs with various tumors, including malignant lymphoma (7 dogs), monocytic leukemia (1), mammary gland adenoma (1), mammary gland benign mixed tumor (1), rhabdomyosarcoma (1), colon cancer (1), and osteosarcoma (3). PROCEDURE: Aberrations of the p53 gene in these tumor tissues were examined by reverse transcriptase-polymerase chain reaction and single-strand conformation polymorphism analysis, using 3 fragments that covered the entire open reading frame of the canine p53 gene, followed by nucleotide sequencing of the abnormal bands. RESULTS: Point mutations, deletions, and insertions resulting in a number of amino acid substitutions of wild-type p53 were detected in 7 of the 15 tumor specimens from dogs with malignant lymphoma, monocytic leukemia, rhabdomyosarcoma, colon cancer, and osteosarcoma. Of these 7 dogs, 2 had aberrations of the p53 gene on both alleles, whereas 5 had aberrations of the p53 gene on 1 allele and concurrently lacked the wild-type p53 transcript. Many of the aberrations of the p53 gene detected in these tumors were located in the transactivation, DNA binding, and oligomerization domains. CONCLUSIONS AND CLINICAL RELEVANCE: Various naturally developing tumors in dogs often have inactivation of the p53 tumor suppressor gene, which may be 1 of the multiple step-wise genetic changes during tumorigenesis. This study indicates that p53 gene can be a target for gene therapy for tumors in dogs.

Animals↗

PubMed2001

Relationship between bond strengths and nanoleakage: evaluation of a new assessment method.

PURPOSE: To test if there was a correlation between resin-dentin bond strengths and nanoleakage of total etch and self-etching primer bonding systems. MATERIALS AND METHODS: Ten extracted third molars were ground flat with 600 grit SIC paper under running water to expose middle dentin. Clearfil Liner Bond 2V or Single Bond was applied to the dentin surface according to the manufacturers' instructions. A crown was built-up with Clearfil AP-X resin-based composite, and the specimens stored in water for 24 hrs at 37 degrees C. The bonded assemblies were cut mesiodistally perpendicular to the interface into approximately 0.7 mm thick slabs, and trimmed for microtensile bond strength testing. Alternate slices from each tooth were either entirely double-coated with nail varnish leaving a 0.5 mm window around the interface, or left uncoated to be immersed in water for 13 hrs as the control. The varnished (experimental) slabs were immersed in 50% AgNO3 for 1 hr, and then immersed in a photo developing solution for 12 hrs. All specimens were subjected to microtensile bond test at a crosshead speed of 1 mm/min. After debonding, the specimens were embedded in epoxy resin and observed under a confocal laser scanning microscope for determination of lateral silver penetration within the interface. Silver penetration was further analyzed under the SEM. Data were analyzed with one- and two-way ANOVA, Fisher's PLSD test (P < 0.05), and regression analysis. RESULTS: The bond strengths of Clearfil Liner Bond 2V and Single Bond were not significantly different (P > 0.05); however the bond strengths of the specimens that were soaked in 50% AgNO3 for 1 hr significantly increased (P < 0.01). No correlation was observed between bond strengths and nanoleakage.

Acid Etching, Dental↗

PubMed2001

[Experience of specific immunotherapy with standardized Japanese cedar pollen extract].

A study was conducted in 165 subjects with Japanese cedar pollinosis (JC) to evaluate the switching to the new standardized extract (SE) for patients who are going on specific immunotherapy (SP-IT) with conventional non-standardized extracts, products of Trii Co. or Hollister-Stier Co. Eight of 137 subjects exhibited adverse systemic reactions such as general skin eruption and despnea when JC allergen extract of Hollister-Stier Co. was switched to JC-SE. There were 6 cases where concentration of the extract had to be decreased due to extraordinary late reactions, while none of 28 subjects exhibited adverse side effect, when conventional allergen extract by Torii Pharmaceutical Co. was switched to JC-SE. Careful switching to high concentration of SE is required, because eight cases with adverse side effect were shown in switching to 200 JAU/ml and 2000 JAU/ml. 20 subjects started with SP-IT by JC-SE after spread of JC in the year of 2000 exhibited no adverse side effects in our protocol.

Adolescent↗

PubMed2000

Molecular cloning and sequencing of equine cDNA encoding serum amyloid A (SAA).

The serum amyloid A (SAA) protein is a characteristic and sensitive acute phase reactant in all vertebrates investigated. We molecularly cloned the equine cDNA encoding SAA from the liver of a healthy horse by polymerase chain reaction (PCR). The cloned cDNA is 480 bases in length, and contains an open reading frame (ORF) of 387 nucleotides encoding a precursor SAA protein of 128 amino acids. The precursor of horse SAA seems to have an 18-residue signal peptide and differs from the reported amino acid sequences of the horse SAA by substitution of valine at residue 81. It shows high homology with SAA amino acid sequence of other species such as dog (80.6%), mink (77.5%), human (76.9%) and duck (71.9%). An insertion of eight amino acids at residues between 85 and 92, as compared to human SAA, has also been found in horse SAA. The availability of the equine SAA cDNA will provide a useful reagent for studying its role in diseased horses.

Amino Acid Sequence↗

PubMed2000

Novel oxidatively stable subtilisin-like serine proteases from alkaliphilic Bacillus spp.: enzymatic properties, sequences, and evolutionary relationships.

The genes for five subtilisin-like serine proteases from alkaliphilic strains of Bacillus exhibiting resistance to oxidative inactivation were cloned and sequenced. The deduced amino acid sequences of the enzymes were highly homologous (greater than 88% identity). They were composed of 638 or 639 amino acids, including a possible approximately 200-amino acid prepro-peptide, and unique stretches of approximately 160 amino acids were found in the C-terminal regions. The molecular masses of mature enzymes (433 or 434 amino acids) were approximately 45 kDa for all. Amino acid sequence comparison and phylogenetic analysis indicated that these enzymes are far removed from other known subtilisins in the line of molecular evolution. We propose that these novel proteases be categorized as a new class of subtilisins, named oxidatively stable, alkaline protease.

Amino Acid Sequence↗

PubMed2000

Antitumor-promoting effects of cyclic diarylheptanoids on Epstein-Barr virus activation and two-stage mouse skin carcinogenesis.

Eleven cyclic diarylheptanoids were screened as potential antitumor promoters by examining the ability of the compounds to inhibit Epstein-Barr virus early antigen activation (induced by 12-O-tetradecanoylphorbol-13-acetate) in Raji cells. 13-Oxomyricanol and myricanone showed the highest activity and also exhibited remarkable inhibitory effects on mouse skin tumor promotion in an in vivo two-stage carcinogenesis test. These data suggest that certain diarylheptanoids might be valuable antitumor promoters and/or chemopreventors.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2000

Nucleophosmin/B23 is a target of CDK2/cyclin E in centrosome duplication.

In animal cells, duplication of centrosomes and DNA is coordinated. Since CDK2/cyclin E triggers initiation of both events, activation of CDK2/cyclin E is thought to link these two events. We identified nucleophosmin (NPM/B23) as a substrate of CDK2/cyclin E in centrosome duplication. NPM/B23 associates specifically with unduplicated centrosomes, and NPM/B23 dissociates from centrosomes by CDK2/cyclin E-mediated phosphorylation. An anti-NPM/B23 antibody, which blocks this phosphorylation, suppresses the initiation of centrosome duplication in vivo. Moreover, expression of a nonphosphorylatable mutant NPM/ B23 in cells effectively blocks centrosome duplication. Thus, NPM/B23 is a target of CDK2/cyclin E in the initiation of centrosome duplication.

3T3 Cells↗

PubMed2000

Antitumor promoting activities of 3-O-acyl-(-)epigallocatechins.

As an exploratory investigation of antitumor promoting compounds, 3-O-acyl-(-)-epigallocatechins possessing a straight-, branched-, phenyl-inserted- or 1,4-phenylene-inserted-acyl chain of varying length from C4 to C18 were synthesized and evaluated their inhibitory effects against the activation of the Epstein-Barr virus early antigen (EBV-EA). It was indicated that the epigallocatechin derivatives having the straight- or branched-acyl chain of C8 to C11 carbon atoms achieve marked effects.

Anticarcinogenic Agents↗

PubMed2000

Genomic convergence and suppression of centrosome hyperamplification in primary p53-/- cells in prolonged culture.

Chromosome instability, a major property of cancer cells, is believed to promote mutations that establish malignant phenotypes. Centrosome hyperamplification and the consequential increase in the frequency of aberrant mitoses are the major causes of chromosome instability in cancer cells that lack the functional p53 tumor suppressor protein. Here, we examined dynamic changes of chromosome and centrosome behaviors during long-term culturing of primary epithelial cells derived from p53-null mice. The heterogeneity in the number of chromosomes per cell in the early to mid passage cell population diminished in late passage cells, giving rise to distinct subpopulations of cells. Concomitantly, centrosome hyperamplification that was observed at a high frequency in early to mid passage cells was suppressed in late passage cells. These results provide an explanation for the frequent observations that some cancer cell lines and tissues that lack functional p53 show normal centrosome behaviors and altered, yet relatively stable, chromosomes. Moreover, our in vitro findings may provide a model for possible genomic convergence in cultured cells. This may be analogous to the genomic convergence model proposed for in vivo tumor progression in which chromosome instability initially imposed during tumorigenesis becomes suppressed when neoplastic cells have acquired chromosome compositions that promise an optimal growth in a given environment.

Animals↗

PubMed2000

Hormonal regulation of organic cation transporter OCT2 expression in rat kidney.

Rat (r) OCT2 was identified as the second member of the organic cation transporter (OCT) family, and is predominantly expressed in the kidney. We reported previously that rOCT2 was responsible for the gender differences in renal basolateral membrane organic cation transport activity. As renal rOCT2 expression in males is much higher than that in females, we hypothesized that rOCT2 expression may be under the control of sex hormones. Treatment of male and female rats with testosterone significantly increased the expression levels of rOCT2 mRNA and protein in the kidney, whereas estradiol treatment moderately decreased the expression levels of rOCT2. There was no regulation of renal rOCT1 mRNA expression by testosterone or estradiol. Treatment of male and female rats with testosterone significantly stimulated the tetraethylammonium (TEA) accumulation by renal slices, whereas estradiol treatment caused a decrease in the TEA accumulation by slices from male but not female rats. The present findings suggested that testosterone up-regulates renal rOCT2 expression and estradiol moderately down-regulates rOCT2.

Animals↗

PubMed2000

Synergistic induction of centrosome hyperamplification by loss of p53 and cyclin E overexpression.

Centrosome hyperamplification and the consequential mitotic defects contribute to chromosome instability in cancers. Loss or mutational inactivation of p53 has been shown to induce chromosome instability through centrosome hyperamplification. It has recently been found that Cdk2-cyclin E is involved in the initiation of centrosome duplication, and that constitutive activation of Cdk2-cyclin E results in the uncoupling of the centrosome duplication cycle and the DNA replication cycle. Cyclin E overexpression and p53 mutations occur frequently in tumors. Here, we show that cyclin E overexpression and loss of p53 synergistically increase the frequency of centrosome hyperamplification in cultured cells as well as in tumors developed in p53-null, heterozygous, and wildtype mice. Through examination of cells derived from Waf1-null mice, we further found that Waf1, a potent inhibitor of Cdk2-cyclin E and a major target of p53's transactivation function, is involved in coordinating the initiation of centrosome duplication and DNA replication, suggesting that Waf1 may act as a molecular link between p53 and Cdk2-cyclin E in the control of the centrosome duplication cycle.

Animals↗

PubMed2000

Structure of the central core domain of TFIIEbeta with a novel double-stranded DNA-binding surface.

Human general transcription factor TFIIE consists of two subunits, TFIIEalpha and TFIIEbeta. Recently, TFIIEbeta has been found to bind to the region where the promoter starts to open to be single-stranded upon transcription initiation by RNA polymerase II. Here, the central core domain of human TFIIEbeta (TFIIEbetac) has been identified by a limited proteolysis. This solution structure has been determined by NMR. It consists of three helices with a beta hairpin at the C-terminus, resembling the winged helix proteins. However, TFIIEbetac shows a novel double-stranded DNA-binding activity where the DNA-binding surface locates on the opposite side to the previously reported winged helix motif by forming a positively charged furrow. A model will be proposed that TFIIE stabilizes the preinitiation complex by binding not only to the general transcription factors together with RNA polymerase II but also to the promoter DNA, where double-stranded DNA starts to open to be single-stranded upon activation of the preinitiation complex.

Amino Acid Sequence↗