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Biomedical subjects

K Kawamura

Publications and source records attributed to K Kawamura.

At least 37 records · Page 2Linked to original sources
PubMed2001

Capillary electrochromatographic enantioseparations using a packed capillary with a 3 microm OD-type chiral packing.

Enantiomer separations by capillary electrochromatography (CEC) using a packed capillary were investigated. As a chiral stationary phase, an OD-type packing material of 3 microm particle diameter, which is a silica-gel based material coated with cellulose tris(3,5-dimethylphenylcarbamate), was employed. The chiral packing was packed into a 100 microm I.D. fused-silica capillary by a pressurized method. Several racemic enantiomers were successfully resolved with acidic or basic buffers containing acetonitrile as mobile phases. The separation efficiencies obtained in this chiral CEC system were superior to those in the previously reported chiral CEC system using 5 microm particles. The typical plate heights obtained for several enantiomers were 4.6-6.6 microm or reduced plate heights of 1.5-2.2. Good repeatabilities in the migration time, peak height, and corrected peak area were observed as well as for the plate number. As an application study of this CEC system, an optical purity test was carried out by using benzoin enantiomers. A 1% content of one enantiomer in the other enantiomer was successfully separated and detected.

Chromatography, Micellar Electrokinetic Capillary↗
PubMed2001

DNA polymerase kappa, implicated in spontaneous and DNA damage-induced mutagenesis, is overexpressed in lung cancer.

DNA polymerase kappa (Pol kappa) is a newly identified low-fidelity polymerase implicated in spontaneous and DNA damage-induced mutagenesis. As an initial study to investigate its possible involvement in tumorigenesis, we compared the expression level of Pol kappa in tumors and adjacent nontumorous tissues by Northern blot, semiquantitative RT-PCR, and Western blot analyses. In this study, paired tumor and normal specimens from 29 patients with stages I to IIIb non-small cell lung cancer (NSCLC), including 13 adenocarcinomas, 15 squamous cell cancers, and 1 adenosquamous carcinoma, were analyzed, among which different levels of tumor-associated Pol kappa overexpression were observed in 21 of 29 matched specimens. In addition, five matched specimens exhibited elevated Pol kappa expression in both tumor and control tissues, whereas only one nontumorous tissue expressed a higher level of Pol kappa than its tumor counterpart. The preferential up-regulation of Pol kappa expression in tumors was highly significant (P < 0.001). There was no apparent correlation of Pol kappa expression levels with tumor histology, grade, and stage or with smoking history. Southern blot analysis did not show amplification of the Pol kappa gene, indicating that the elevated Pol kappa expression is likely attributable to dysregulated transcription. Our data suggest that Pol kappa may contribute to lung tumor development by accelerating the accumulation of mutations.

Aged↗
PubMed2001

A comparison of intraperitoneal and oral gavage administration in comet assay in mouse eight organs.

One of the important advantages of the comet assay is its ability to detect genotoxicity in many different organs. Since the exposure route of the test compounds is likely to influence the genotoxicity detected in a given organ, it is an important factor to consider when conducting the assay. In this study, we compared the effects of numerous model compounds on eight organs when administered to mice by intraperitoneal (i.p.) injection and oral (p.o.) gavage. Groups of four mice were treated once i.p. or p.o. at the identical proportion of LD50 for each route, and the stomach, colon, liver, kidney, bladder, lung, brain, and bone marrow were sampled 3, 8, and 24h after treatment. For 19 of the 20 tested mutagens with various modes of action, genotoxicity in some organs varied with treatment route; only the genotoxicity of methyl methane sulfonate was not affected. Treatment route, however, did not produce a qualitative difference in the genotoxicity of promutagens at the sites of conversion to ultimate mutagens, with aromatic hydrocarbons as the exception. When chemicals with positive responses in at least one organ were considered to be comet assay-positive, the administration route made no difference. Since azo reduction is mediated by azo reductase synthesized in the gastrointestinal wall and by gut microflora and i.p.-administered azo dyes bypass their activation site (colon), the administration route is expected to make a difference in their in vivo genotoxicity. Direct-acting mutagens are expected to affect the mucosa of the gastrointestinal tract when given p.o. For those mutagens, however, the administration route did not make a qualitative difference in gastrointestinal tract genotoxicity. Moreover, although the gastrointestinal mucosa is the first site to be exposed to p.o. administered agents, the peak times in the stomach tended to be the same as in most other organs. Based on those results, we concluded that the genotoxicity at high exposures was due to a systemic effect, and that both routes are acceptable for the comet assay when the liver and gastrointestinal organs are sampled, so long as appropriate dose levels for systemic exposure are selected for each route.

Administration, Oral↗
PubMed2001

cAMP and calcium ionophore induce outgrowth of neuronal processes in PC12 mutant cells in which nerve growth factor-induced outgrowth of neuronal processes is impaired.

During continuous culturing, PC12 cells are subject to spontaneous mutations. We obtained PC12m3 cells, clone cells in which outgrowth of neuronal processes (dendrites and axons) under the condition of nerve growth factor (NGF) treatment was highly stimulated by various inducers, such as cyclic adenosine monophosphate (cAMP), calcium ionophore, steroid and high osmolarity. The number of cells with neuronal processes in the presence of cAMP was approximately twenty-fold greater than PC12 parental cells and other PC12 mutant cells. In PC12m3 cells, NGF-induced outgrowth of neuronal processes was reduced by cytotoxic solanine, whereas the effect of NGF was unaffected by hyaluronic acid. In PC12m3 cells, various inducers of neurite outgrowth, such as cAMP, calcium ionophore and high osmolarity, activated mitogen activated protein (MAP) kinase, whereas solanine and hyaluronic acid did not cause any significant activation of MAP kinase. However, PC12m3 cells, in which NGF-induced outgrowth of neuronal processes were impaired, had strong NGF-induced MAP kinase activity as PC12 parental cells had. These findings suggest that cAMP, calcium influx and high osmolarity induce outgrowth of neuronal processes in PC12m3 cells through activation of the downstream target of MAP kinase or through a novel pathway independent of NGF activation.

Animals↗
PubMed2001

The pharmacological profile of delta opioid receptor ligands, (+) and (-) TAN-67 on pain modulation.

We designed the nonpeptidic highly selective delta opioid receptor agonist on the basis of message address concept and the accessory site theory and synthesized (+/-) TAN-67. In spite of highly potent agonistic activity in in vitro assay, (+/-) TAN-67 (racemate) afforded a weak antinociceptive effect in the mouse tail-flick test. This result led us to separate (+/-) TAN-67 to optical pure compounds, (+) and (-) TAN-67. An i.t.-treatment with (-) TAN-67 produced profound antinociceptive effects through specifically acting on delta1 receptors. Unlike (-) TAN-67, i.t.-administered (+) TAN-67 displayed dose-related nociceptive behaviors such as scratching, biting and licking. The effect of (+) TAN-67 was blocked by i.t.-treatment with NTI (delta receptor antagonist) and (-) TAN-67 (delta1 receptor agonist), but not by morphine (mu receptor agonist). The mechanisms involved in spinal pain modulation induced by (+) and (-) TAN-67 were also described.

Analgesics↗
PubMed2001

Specific phosphorylation of nucleophosmin on Thr(199) by cyclin-dependent kinase 2-cyclin E and its role in centrosome duplication.

The kinase activity of cyclin-dependent kinase 2 (CDK2)-cyclin E is required for centrosomes to initiate duplication. We have recently found that nucleophosmin (NPM/B23), a phosphoprotein primarily found in nucleolus, associates with unduplicated centrosomes and is a direct substrate of CDK2-cyclin E in centrosome duplication. Upon phosphorylation by CDK2-cyclin E, NPM/B23 dissociates from centrosomes, which is a prerequisite step for centrosomes to initiate duplication. Here, we identified that threonine 199 (Thr(199)) of NPM/B23 is the major phosphorylation target site of CDK2-cyclin E in vitro, and the same site is phosphorylated in vivo. NPM/T199A, a nonphosphorylatable NPM/B23 substitution mutant (Thr(199) --> Ala) acts as dominant negative when expressed in cells, resulting in specific inhibition of centrosome duplication. As expected, NPM/T199A remains associated with the centrosomes. These observations provide direct evidence that the CDK2-cyclin E-mediated phosphorylation on Thr(199) determines association and dissociation of NPM/B23 to the centrosomes, which is a critical control for the centrosome to initiate duplication.

3T3 Cells↗
PubMed2001

Myelin basic protein is necessary for the regulation of myelin-associated glycoprotein expression in mouse oligodendroglia.

Myelin-associated glycoprotein (MAG) has currently been proposed to be a possible igniter of myelination in the central nervous system. We propose here that myelin basic protein (MBP) is prerequisite, which is more diffusely expressed than MAG in early stages of myelination. MBP-deficient mice show significant reduction of MAG-positive immature oligodendroglia at early stages of myelination, although significant increase of such cells is noted in the adult. MBP/Fyn-double deficient adult mice also show moderate increase of the same type of immature glia, but to a lesser degree than MBP-deficiency alone. The present study suggests that MBP is required for the regulation of MAG expression in oligodendroglia, involving Fyn therein. We discuss the possibility that hitherto unknown molecule, not MAG, may be in responsible for the ignition of the myelination.

Animals↗
PubMed2001

Defect in SHAP-hyaluronan complex causes severe female infertility. A study by inactivation of the bikunin gene in mice.

Hyaluronan (HA) associates with proteins and proteoglycans to form the extracellular HA-rich matrices that significantly affect cellular behaviors. So far, only the heavy chains of the plasma inter-alpha-trypsin inhibitor (ITI) family, designated as SHAPs (serum-derived hyaluronan-associated proteins), have been shown to bind covalently to HA. The physiological significance of such a unique covalent complex has been unknown but is of great interest, because HA and the ITI family are abundant in tissues and in plasma, respectively, and the SHAP-HA complex is formed wherever HA meets plasma. We abolished the formation of the SHAP-HA complex in mice by targeting the gene of bikunin, the light chain of the ITI family members, which is essential for their biosynthesis. As a consequence, the cumulus oophorus, an investing structure unique to the oocyte of higher mammals, had a defect in forming the extracellular HA-rich matrix during expansion. The ovulated oocytes were completely devoid of matrix and were unfertilized, leading to severe female infertility. Intraperitoneal administration of ITI, accompanied by the formation of the SHAP-HA complex, fully rescued the defects. We conclude that the SHAP-HA complex is a major component of the HA-rich matrix of the cumulus oophorus and is essential for fertilization in vivo.

Alpha-Globulins↗
PubMed2001

Primary lung cancer occurring concomitantly with the cicatrized and calcified ova of a parasite: report of a case.

We report herein a rare case of primary lung cancer that occurred concomitantly with the calcified ova of a parasite. A 58-year-old man was referred to our department after a pulmonary abnormal shadow had been seen on a chest X-ray done at mass screening. A transbronchial lung biopsy (TBLB) revealed the calcified ova of a parasite. Because the possibility of concomitant lung cancer could not be ruled out, a lung biopsy was taken via video-assisted thoracic surgery (VATS). The pathological diagnosis was squamous cell carcinoma, and a left upper lobectomy was serially performed through a posterolateral thoracotomy. The patient recovered uneventfully and has remained in good health without any sign of recurrence for over 9 months. Following this case report, we review three other cases of this unusual disease combination documented in the literature.

Calcinosis↗
PubMed2001

Vectorial delivery of macromolecules into cells using peptide-based vehicles.

The ability to direct the import of therapeutic agents into cells and target them to specific organelles would greatly enhance their functional efficacy. The available spectrum of peptide-based import signals and intracellular routing signals might provide practical solutions towards achieving a guided or vectorial delivery of molecules. Multiple cell-targeting signals and routing domains can be efficiently displayed on branched peptides. These constructs are typically nonimmunogenic in the absence of adjuvant and can be easily assembled using solid phase synthesis. The vectorial delivery of larger complexes, however, will necessitate the development of alternate templates that favor the optimal presentation of all functional signals.

Cell Nucleus↗
PubMed2001

[11C]Raclopride binding was reduced in vivo by sigma(1) receptor ligand SA4503 in the mouse brain, while [11C]SA4503 binding was not by raclopride.

[11C]Raclopride is widely used as a representative dopamine D(2)-like receptor ligand in positron emission tomography (PET) studies, and [11C]1-(3,4-dimethoxyphenethyl)-4-(3-phenylpropyl)piperazine dihydrochloride ([11C]SA4503) is a recently developed selective ligand for mapping sigma(1) receptors in the brain. The striatal uptake of [11C]raclopride in mice was reduced by co-injection of an excess amount of SA4503, in spite of the fact that raclopride had no effect on the brain uptake of [11C]SA4503 as shown in a previous study. The blocking effect of SA4503 on the striatal uptake of [11C]raclopride was dose-dependent, but disappeared by 1 h or 6 h after intraperitoneal injection of SA4503. The brain uptake of [11C]SA4503 was not affected by a dopamine transporter inhibitor GBR 12909, nor was [11C]beta-CIT-FP inhibited by SA4503. The IC(50) values of raclopride for sigma(1) and sigma(2) receptor subtypes measured in vitro were 11800 nM and 4950 nM, respectively, suggesting that the affinity was too low for [11C]raclopride to bind in vivo to sigma receptors. On the other hand, the IC(50) value of SA4503 for dopamine D(2) receptors was 470 nM, that is approximate 1/25 of the affinity of raclopride for the dopamine D(2) receptors. Therefore, possible explanations for the partial blocking effects of SA4503 on the striatal uptake of [11C]raclopride are: (1) an excess amount of SA4503 may reduce the [11C]raclopride uptake due to its low affinity for dopamine D(2) receptors, or (2) SA4503 may enhance endogenous dopamine release, which results in the competitive inhibition of the [11C]raclopride uptake. These findings support that both [11C]raclopride and [11C]SA4503 are selective in vivo ligands for dopamine D(2)-like receptors and sigma(1) receptors, respectively, in spite of the partial blocking effect of SA4503 on the striatal uptake of [11C]raclopride.

Animals↗
PubMed2001

Efficacy of the bystander effect in the herpes simplex virus thymidine kinase-mediated gene therapy is influenced by the expression of connexin43 in the target cells.

Tumoricidal "bystander effect" observed in the herpes simplex virus thymidine kinase (HSVtk)/ganciclovir (GCV) gene therapy was studied between different rat glioma cell lines (9L and C6 cells) under both in vitro and in vivo conditions. For that purpose, mixed populations of wild-type cells (9Lwt and C6wt) and respective HSVtk gene-transduced cells (9Ltk and C6tk) were examined for their sensitivity to GCV. A potent in vitro bystander effect was observed in 9Lwt/9Ltk and 9Lwt/C6tk combinations but not in C6wt/9Ltk and C6wt/C6tk combinations. In vivo bystander effect studied in a subcutaneous tumor model in athymic nude mice was also potent in 9Lwt/9Ltk and 9Lwt/C6tk combinations. Because the expression of connexin43, a major protein in the connexin family gene products, in 9L cells is much higher than that in C6 cells, the results suggest that the amount of connexin in target (wild-type) cells but not in effector (HSVtk gene-bearing) cells is important for the generation of the bystander effect. This hypothesis was further confirmed by the observation that in vitro bystander effect in C6wt/C6tk combination was potentiated by transduction of the connexin43 gene to the target cells.

Animals↗
PubMed2001

Endometrioid adenocarcinoma arising from adenomyosis: report and immunohistochemical analysis of an unusual case.

A case of endometrioid adenocarcinoma arising from adenomyosis is reported. The patient was a 53-year-old woman who complained of vulvar itching. Smear cytology of the endometrium revealed adenocarcinoma. Magnetic resonance imaging of the pelvis revealed a lesion with a slightly high intensity in the uterine fundus on a T2-weighted image. Semiradical total hysterectomy and bilateral adnexectomy were performed, followed by chemotherapy. Histologically, the lesion in the uterine fundus was composed mostly of adenocarcinoma with stromal invasion. There were many adenomyotic foci in and around the carcinoma, including some showing transition to adenocarcinoma. There was no malignant finding in the normally situated endometrium. The carcinoma invaded in the myometrium, involving the uterine serosa, but no dissemination to the peritoneal cavity was found. The carcinoma was, therefore, considered to be endometrioid adenocarcinoma arising from adenomyosis. Immunohistochemistry showed expression of p53 oncoprotein and Ki-67 antigen in the carcinoma cells. The value of immunohistochemistry in predicting prognosis is discussed.

Adenocarcinoma↗
PubMed2001

Evaluation of neurotoxicity of TIQ and MPTP and of parkinsonism-preventing effect of 1-MeTIQ by in vivo measurement of pre-synaptic dopamine transporters and post-synaptic dopamine D(2) receptors in the mouse striatum.

Parkinsonism-inducing neurotoxicity of 1,2,3,4-tetrahydroisoquinoline (TIQ), as contrasted to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and parkinsonism-preventing effect of 1-methyl-1,2,3,4-tetrahydroisoquinoline (1-MeTIQ) have been investigated in mice by measuring their effects on the in vivo binding of radioligand to pre-synaptic dopamine transporters (DATs) or to dopamine D(2) receptors (D2R) in the striatum. A significant reduction of the ligand-DATs binding was found in the mice treated with MPTP, but not with TIQ, under the dosage inducing behavioral abnormality and loss of tyrosine hydroxylase-positive cells in the substantia nigra. A slight decrease in the ligand-DATs binding was observed in the mice given a larger dose of TIQ. Compensatory up-regulation in the post-synaptic D2Rs was found in the MPTP-treated mice. Pre-treatment with (S)-enantiomer, but not (R)-enantiomer, of 1-MeTIQ prevented the degeneration of DATs to some extent. We concluded that the TIQ-induced parkinsonism model is different from the MPTP-induced model as evaluated by the radioligand-DATs binding and that (S)-1-MeTIQ has a preventing effect for the degeneration of the DATs to a certain extent.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed2001

Expression of intercellular adhesion molecule 1 (ICAM-1) in Plasmodium falciparum-infected placenta.

We investigated the expression of intercellular adhesion molecule 1 (ICAM-1) in malarial placenta and related histological changes. Thirty-two malarial and 40 control term placentae were collected at Tanga, Tanzania and examined histologically and immunohistochemically. Malaria infected placentae were further divided into acute (15) and chronic (17) cases according to the presence of malarial pigment. The expression of ICAM-1 on monocyte, syncytio- and cytotrophoblasts, endothelial and stromal cells was assessed. Birthweight was lower and leukocyte counts higher in placentae with chronic infection. Many monocytes were present within the intervillous spaces, especially in placentae with chronic infection, and aggregated with parasitized erythrocytes. Some monocytes were adhesive to the surface of fibrinoid deposits. ICAM-1 expression on monocytes of malarial placentae was significantly conspicuous and correlated to the degree of intervillous leukocyte infiltration. Syncytiotrophoblasts often did not show ICAM-1, even though ICAM-1 was expressed by endothelium and weakly by cytotrophoblasts and stromal cells in both infected and control placentae. These results suggest that the expression of ICAM-1 on monocytes contributes to sequestration of infected erythrocytes within the intervillous spaces and their adhesion to fibrin masses and that ICAM-1 is unlikely to be associated with the direct adhesion of infected erythrocytes to the syncytiotrophoblasts.

Birth Weight↗