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Biomedical subjects

K Kawamura

Publications and source records attributed to K Kawamura.

At least 271 records · Page 15Linked to original sources
PubMed1995

Production of thrombopoietin (TPO) by rat hepatocytes and hepatoma cell lines.

Recently, we purified rat thrombopoietin (TPO) from plasma of irradiated rats (XRP) by measuring its activity that stimulated the production of megakaryocytes from megakaryocyte progenitor cells (CFU-MK) in vitro. We then cloned the cDNAs for rat and human TPO. In this study, we found the production of TPO by hepatocytes isolated with the collagenase perfusion method from both normal and thrombocytopenic rats, by a two-step fractionation of hepatocyte culture medium (CM). Subsequently, CM of rat hepatoma cell lines was screened for the presence of TPO; three cell lines, H4-II-E, McA-RH8994, and HTC, were found to produce TPO. According to the purification procedure for TPO from XRP, TPO was partially purified from 2 L CM of each of three cell lines with a six-step procedure. In the final reverse-phase column, TPO from each cell line was eluted with the same retention time as that from XRP, and the TPO fraction exhibited megakaryocyte colony-stimulating activity (Meg-CSA). TPO-active fraction eluted from the final reverse-phase column was separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), extracted from the gel, and assayed. TPO activity from each cell line was found in the respective molecular weight region, indicating the heterogeneity of the TPO molecule. Using reverse transcriptase-polymerase chain reaction (RT-PCR), we detected the expression of TPO mRNA in hepatocytes, three hepatoma cell lines, normal rat liver, and X-irradiated rat liver. Northern blot analysis showed that TPO mRNA was expressed mainly in liver among the various organs tested. These data demonstrate that TPO is produced by rat hepatocytes and hepatoma cell lines and suggest that liver may be the primary organ that produces TPO.

Animals↗
PubMed1995

[Diagnostic significance of QRST isointegral map in patients with premature ventricular contraction].

Identification of the focus of premature ventricular contraction (PVC) has been reported by various body surface mapping methods: i.e. PVC isopotential map, ventricular activation time map, QRS isointegral map, PVC T wave map, etc. For evaluating the clinical usefulness and diagnostic significance of QRST isointegral map in patients with PVC, 23 patients without apparent underlying heart disease (group N), and 44 patients with dilated cardiomyopathy (group D), were studied. The significant changes in QRST values at maximal points during two activation sequences (basic rhythm and PVC) were detected in group D (N:D = 26.6 +/- 17.6:52.6 +/- 27.0%, p < 0.01), which were frequently directed to the PVC focus and influenced by the short coupling interval (Q-Q interval: 300-390 ms). In conclusion, QRST isointegral map reflecting ventricular gradient may be useful for identifying the presence of underlying heart disease and may correlate with the focus and coupling interval of PVC.

Body Surface Potential Mapping↗
PubMed1995

The effects of neurotoxins 6-hydroxydopamine and 5,7-dihydroxytryptamine into the rat nucleus accumbens on the alcohol drinking behavior.

To elucidate the relationship between drinking behavior and brain monoamines in the brain rewarding system, the effects of injection of 6-hydroxydopamine (6-OHDA) and 5,7-dihydroxytryptamine (5,7-DHT) into the rat nucleus accumbens (ACC), which would affect dopamine- and indoleamine-containing neurons, respectively, on the rat alcohol preference score were examined. The 6-OHDA (4 micrograms/side)-treated rats showed a higher alcohol preference score for 2 weeks following the treatment, and a lowered dopamine (DA) level in the ACC and midbrain after ACC injection of 6-OHDA. On the other hand, although the serotonin (5-HT) and DA levels in the midbrain and 5-HT level in ACC were lowered after the injection of 5,7-DHT, the alcohol preference score was not significantly changed in the 5,7-DHT (4 micrograms/side)-treated rats. Taken together, these findings suggest that the alcohol drinking behavior is more influenced by the ACC DA activity than 5-HT activity. Changes in alcohol drinking behavior might be related to the compensatory mechanism for the rat to restore the original rewarding properties.

5,7-Dihydroxytryptamine↗
PubMed1995

[QRST isointegral map in dilated cardiomyopathy].

To evaluate the diagnostic usefulness and prognostic significance of QRST isointegral map in dilated cardiomyopathy (DCM), we performed body surface mapping, signal averaged electrocardiogram and thallium-201 myocardial scintigraphy in 41 DCM patients. Late potentials (LP) were detected in 17 patients (41%). QRST isointegral subtraction map (QRST I-sub map: departure map, calculated from 40 normal subjects) showed -2 standard deviation area and departure indices at minimal point (DImin) were calculated in each case. Significant coefficients of correlation were observed between the DImin and root mean square voltage in last 40 ms (r = -0.435, p < 0.001). The abnormal scintigraphic patterns were discriminated into 3 groups: anteroseptal, inferoapical and posterolateral defects, which revealed separate distribution of minimal point in QRST I-sub map: F-G 4, G-H 2 and I-J 4, respectively. These results suggest that QRST isointegral map is useful for detecting the presence of LP (prognostic value) and the location of scintigraphic perfusion defect (diagnostic value) in DCM.

Body Surface Potential Mapping↗
PubMed1995

Localization of matrix metalloproteinase 9 (92-kilodalton gelatinase/type IV collagenase = gelatinase B) in osteoclasts: implications for bone resorption.

BACKGROUND: Matrix metalloproteinase 9 (MMP-9, 92-kD gelatinase/type IV collagenase = gelatinase B) is a member of the MMP gene family and implicated in tissue destruction in the various pathophysiologic conditions. Our previous study showed that MMP-9 purified from human fibrosarcoma cells can cleave the cross-link-containing NH(2)-terminal telopeptides of the alpha 2 chain of type I collagen and collagen types III, IV, and V as well as gelatins. EXPERIMENTAL DESIGN: To investigate the role of MMP-9 in bone resorption we have examined its localization in the human bone tissues by immunohistochemistry and in situ hybridization. The enzymic properties were also biochemically studied. RESULTS: Immunohistochemistry using monoclonal antibodies against MMP-1 (interstitial collagenase), MMP-2 (72-kD gelatinase/type IV collagenase = gelatinase A), MMP-3 (stromelysin-1), MMP-9, and tissue inhibitor of metalloproteinases-1 demonstrated that MMP-9 is localized exclusively in osteoclasts of the bone tissues from normal subjects and patients with rheumatoid arthritis or metastatic carcinoma whereas some osteoclasts are also labeled by anti-(MMP-1) antibody. Northern blot and in situ hybridizations of rheumatoid bone tissues using a RNA probe for MMP-9 exhibited strong signals for the mRNA within osteoclasts. MMP-9 depolymerized acid-insoluble polymers of type I collagen and digested collagen fibrils in the demineralized bone. The gelatinolytic activity of the proteinase was optimal at pH 7.5, but 50 to 80% of the full activity was retained at pH 5.5 to 6.0. It was also 90% active in the presence of 100 mM Ca2+. Degradation of acid-soluble and -insoluble type I collagens by MMP-9 was enhanced at higher concentrations of Ca2+. The zymogen of MMP-9 was activated up to approximately 85% of full activity by incubation at pH 2.3. CONCLUSIONS: These results demonstrate that MMP-9 is produced by osteoclasts in the human bone tissues and suggest that it can degrade bone collagens in concert with MMP-1 and cysteine proteinases in the subosteoclastic microenvironment. This proteinase may play a role in the normal bone remodeling and pathologic bone resorption in the human diseases.

Adolescent↗
PubMed1995

[Migration, differentiation and integration of an immortalized neural cell line transplanted into the neonatal and adult mouse brain].

An immortalized neural cell line V1 was transplanted stereotaxically into the cerebellum and hippocampus of developing and adult mice, and the mode of migration, differentiation and arrangement of the grafted cells were examined by labeling the grafted cells with DiI (1, 1'-dioctadecyl-3, 3, 3', 3'-tetramethylindocarbocyanine perchlorate) and immunohistochemical staining. This cell line was established by transduction of the temperature-sensitive allele tsA58 of SV40 large T antigen oncogene into mouse hypothalamic cells. Grafted cells did not show any tumorigenicity for a long time. Some of the cells grafted into the neonatal cerebellum and hippocampus were arranged along the host cortical layer and showed neuronal or glial differentiation according to the grafted site. The cells grafted into adult cerebellum also showed migration and arrangement along the host cortical layer as well as morphological differentiation into glial cells in a manner similar to that of transplantation to the neonate. On the other hand, the cells grafted into the adult hippocampus made only clusters without forming an organized arrangement. These findings suggest that the grafted cells are integrated into the developmental processes of the host brain, and the mode of differentiation and arrangement of the grafted cells depends on the microenvironment of the different developmental stages of the host brain. The involvement of host blood vessels and astroglial framework in the migration and arrangement of the grafted cells was also suggested. Furthermore, these findings suggest the plasticity of the host brain in response to the grafted cells and the possibility of reconstructing the host brain with this multipotential neural cell line.

Animals↗
PubMed1995

A chorioretinal lesion in a patient with Kimura's disease.

A 47-year-old male patient with Kimura's disease showed a white and slightly elevated choroidal lesion surrounded by serous retinal detachment in his left eye. Serological examination revealed esinophilia. Swelling in his left cheek had been increasing during a period of 20 years, and diagnosis of Kimura's disease had been made earlier by biopsy. Fluorescein angiography suggested disturbance of choroidal circulation and dysfunction of retinal pigment epithelium. After corticosteroid therapy, improvement was seen in the fundus findings, the swelling in the cheek and eosinophilia.

Angiolymphoid Hyperplasia with Eosinophilia↗
PubMed1994

cDNA cloning and expression of a novel human desmocollin.

We have cloned a human cDNA encoding for a novel transmembrane protein from a bladder carcinoma cell line (HT-1376) cDNA library. Sequence analysis of this clone revealed an open reading frame of 2,691 bases encoding a protein of 896 amino acids. Sequence comparison showed that this clone has significantly homology to desmocollins, intracellular adhesion molecules belonging to the cadherin superfamily. This protein consists of a signal peptide of 30 amino acids, a precursor segment of 105 amino acids, and a mature protein of 761 amino acids. We expressed this cDNA clone in COS1 cells and found that the predicted cell adhesion molecule remained in the membrane fraction. Antibodies recognizing the predicted mature adhesion molecule of the protein stained antigens along the cell boundaries of both HT-1376 cells and normal human keratinocytes resembling the pattern of desmosome localization. These findings suggest that our clone might be a new member of the desmocollin family, and we have tentatively called this clone human desmocollin type 4.

Alternative Splicing↗
PubMed1994

Molecular cloning and chromosomal localization of the human thrombopoietin gene.

The complete gene for human thrombopoietin (TPO) has been cloned by screening a human genomic library using human TPO cDNA as a probe. This gene is 6.2 kb in length and contains six exons and five introns. It is shown that the human genome contains a single copy of the human TPO gene according to Southern blotting analysis. The transcription initiation site was determined by S1 nuclease mapping. The human TPO gene expressed TPO activity when transfected into COS-1 cells. The human TPO gene has been mapped to chromosome 3q27 by in situ hybridization using a biotin-labeled probe.

Amino Acid Sequence↗
PubMed1994

Molecular cloning of complementary DNA for a novel human hepatoma-derived growth factor. Its homology with high mobility group-1 protein.

A novel hepatoma-derived growth factor was purified from the conditioned medium of human hepatoma-derived cell line, HuH-7, by the assay of [3H]thymidine incorporation into Swiss 3T3 cells. Molecular cloning of a complementary DNA from the cDNA library of HuH-7 cells was done on the basis of the N-terminal amino acid sequence. This protein was acid- and heat-labile heparin-binding protein and inactivated by reducing condition. The cDNA is 2.4 kilobase pairs long, and the deduced amino acid sequence from cDNA contained 240 amino acids without a signal peptide-like N-terminal hydrophobic sequence. Heparin column-eluted fraction of the conditioned medium of Cos-7 cells transfected by the cDNA stimulated the DNA synthesis. Northern blot analysis revealed its ubiquitous expression in several tumor-derived cell lines, as well as in normal tissues. The primary sequence shares homology with the high mobility group (HMG)-1 protein (23.4% amino acid identity and 35.6% similarity). However, its hydrophobic profile was distinct from that of HMG-1 except for the C-terminal acidic region, and it contained no apparent HMG box motif. Immunofluorescence study showed that it is localized in the cytoplasma of hepatoma cells. These findings suggest that this factor is a novel heparin-binding protein, with mitogenic activity for fibroblasts.

Amino Acid Sequence↗
PubMed1994

Expression of neuronal src mRNA as a favorable marker and inverse correlation to N-myc gene amplification in human neuroblastomas.

Neuron-specific src mRNA, which is expressed in human brain tissue by alternative splicing, is associated with neural differentiation. Neuronal c-srcNI expression may be associated with the ability of neuroblastomas to mature; furthermore, c-srcN2 mRNA is induced in chemically differentiated neuroblastoma cells in vitro. The prognosis of a patient with a neuroblastoma is strongly affected by the ability of the tumor to differentiate in vivo. In order to clarify the relationship between neuronal src mRNA expression and the clinical outcome of a neuroblastoma, we analyzed the expression of src mRNA in neuroblastoma tissues from 28 patients by SI-nuclease-protection assay. N-myc gene amplification was also examined by Southern blot hybridization. The clinical significance of neuronal src mRNA expression and its relevance to N-myc gene amplification was also investigated. A high ratio (more than 10%) of c-srcN2 mRNA expression was observed in all early-stage tumors and in advanced neuroblastomas with a favorable prognosis. In contrast, in advanced neuroblastomas with an aggressive clinical phenotype, c-srcN2 mRNA expression was found at a low ratio (below 10%). Genomic amplification of the N-myc gene and expression of c-srcN2 mRNAs were inversely correlated. When combined with other prognostic markers such as N-myc gene amplification, the expression of c-srcN2 mRNA may be a new biological marker to predict the prognosis of patients with neuroblastomas.

Blotting, Southern↗
PubMed1994

Analysis of lung surfactant in the metamorphosing bullfrog (Rana catesbeiana).

In bullfrog (Rana catesbeiana) tadpoles, the lung begins to function at an advanced stage of metamorphosis. As a preliminary step for investigation of the mechanisms involved in lung maturation, pulmonary surfactant was prepared from tadpoles at advanced stages of metamorphosis and its biochemical properties were analyzed. Surfactant phospholipid analysis revealed that the major constituent was phosphatidylcholine (PC), as examined in the animals at late climax (stage 24). Other detectable phospholipids were phosphatidylethanolamine, phosphatidylserine, sphingomyelin, and phosphatidylglycerol, a marker lipid in mammalian surfactant. As in mammals, PC in the surfactant was rich in saturated fatty acids, about 50% of fatty acid moieties being palmitic acid. The content of surfactant PC in the lung increased moderately around mid-climax and markedly at the end of climax. The effect of antiserum against bullfrog prolactin (PRL) on the pulmonary surfactant was studied in climactic tadpoles. The content of surfactant PC in the lung of the antiserum-treated larvae was lower than that in the lung of the normal rabbit serum-injected larvae, whereas the content of PC in the whole lung did not differ between the antiserum-treated and control groups. The results suggest that synthesis of surfactant in the amphibian lung is enhanced as metamorphosis progresses and that PRL is involved in lung maturation.

Animals↗
PubMed1994

The development of noradrenaline-, acetylcholinesterase-, neuropeptide Y- and vasoactive intestinal polypeptide-containing nerves in human cerebral arteries.

The development of cerebrovascular nerves containing noradrenaline (NA), acetylcholinesterase (AChE), neuropeptide Y (NPY) and vasoactive intestinal polypeptide (VIP) in the basilar artery was histochemically or immunohistochemically studied in human fetuses of various gestational ages. At the 12th week of gestation, nerve fibers containing NA and AChE appeared. Both types of nerve fibers consisted exclusively of the longitudinal nerve fibers (LNF) running along the basilar arteries. Subsequently, the circular nerve fibers (CNF) gradually increased and such development was then completed by the 20th week of gestation. NPY- and VIP-containing nerve fibers were detected in human fetal cerebral arteries in the 16th week of gestation, in the form of LNF. Subsequently, CNF gradually increased and development was completed by the 24th week of gestation. These results indicate that neuropeptide-containing nerve fibers, together with NA- and AChE-containing nerves, therefore contribute to the control of cerebral circulation in the later stages of embryogenesis in human cerebral arteries.

Acetylcholinesterase↗
PubMed1994

Canine Lyme disease: clinical and serological evaluations in 21 dogs in Japan.

Of 21 dogs from Sapporo, Hokkaido that had been recognised as having been bitten by ticks, 16 were seropositive to Borrelia burgdorferi by ELISA. Thirteen of the seropositive dogs showed signs such as fever, astasia, convulsions, anorexia, fatigue, abnormal gait, nervous signs, diarrhoea, corneal opacity and conjunctivitis. These signs subsided as a result of antibiotic treatment within five days. The plasma concentrations of creatinine in the 21 dogs were higher than in control dogs. Seven ticks that were removed from seven of the dogs were Ixodes persulcatus, and B burgdorferi was isolated from the midgut of two of the ticks.

Animals↗
PubMed1994

Inhibition by aurintricarboxylic acid of von Willebrand factor binding to platelet GPIb, platelet retention, and thrombus formation in vivo.

Commercial aurintricarboxylic acid (ATA) was separated into molecular-weight (MW) fractions of < 210 to > 25,000, using gel permeation chromatography. Fractions with MW > 1,300 effectively inhibited both botrocetin-induced vWF and bovine vWF binding to fixed human platelets. These activities decreased with a MW > 17,000. Platelet retention for a human in vitro was reduced by ATA at 150 microM, as was that for rats ex vivo at 3 mg/kg. ATA prolonged tail transection bleeding time in rats but had only a weak effect on buccal mucosal bleeding time in dogs. ATA had no effect on platelet count but markedly prolonged PTT. ATA at 10 mg/kg exhibited antithrombotic activity and caused a marked improvement in patency status following successful thrombolysis by t-PA in electrically and copper coil-induced thrombosis models. These results suggest that specific inhibitors of the vWF-GPIb interaction such as ATA may prove useful as antithrombotic agents.

Animals↗
PubMed1994

Predominant induction of kinetochore-containing micronuclei by extracts of diesel exhaust particulates in cultured human lymphocytes.

The aneuploidy-inducing activity of extracts of diesel exhaust particulates from light duty (LD) and heavy duty (HD) engines was investigated in cultured peripheral blood lymphocytes of 8 healthy donors using the cytokinesis-block micronucleus test with the kinetochore labelling modification. A majority of the subjects tested showed a significant kinetochore-positive micronucleus induction after treatment with the highest dose (150 micrograms/ml) of LD extract, although some subjects also showed induction of kinetochore-negative micronuclei. Only one subject had significantly increased numbers of kinetochore-positive micronuclei at a dose of 400 micrograms/ml of HD extract. These results suggest that diesel extract, at least LD extract, possesses the ability to induce whole chromosome loss (aneuploidy) preferentially, although there are also chromosome breaks.

Adult↗
PubMed1994

Interactions between growing thalamocortical afferent axons and the neocortical primordium in normal and reeler mutant mice.

The interactions between growing thalamocortical afferent axons and the neocortical primordium were examined during neocortical development of the mouse cerebrum, by labeling the afferents with the carbocyanine fluorescent dye, DiI, which was introduced into the dorsal thalamus of the fixed brains of control and reeler mutant mice. In the neocortical primordium of the control mouse, the labeled afferents running tangentially in the intermediate zone formed a dense plexus in the subplate, the layer below the cortical plate, as early as the 16th gestational day (E16). Small numbers of the afferents invaded the lower cortical plate at E16 and increasing numbers of labeled growing axons extended into the cortical plate at E17. At the 4th postnatal day (P4), labeled afferents grew radially up to the upper cortical plate and terminal arborizations of the afferents were evident in the forming layer IV. In contrast, in the E16 cerebrum of the reeler mutant mouse, in which the cortical layers are inverted, the labeled afferents traversed the neocortical primordium directly towards the superplate, the superficial layer above the cortical plate and the equivalent of the subplate in the control mouse. Thick bundles of labeled axons reached the superplate and made contact with the superplate neurons. At P4 in the reeler neocortex, the afferent axons that had reached the superplate began to change their direction of growth and turned towards the deeper layer. Electron-microscopic observations at E16 revealed that immature synapses were formed on the somata of the subplate neurons in the control mouse, and similar immature synapses were also formed on the superplate neurons of the reeler mutant. At E16 in the control, NGF receptor immunoreactivity was expressed in the intermediate zone, subplate and lower cortical plate, and the mode of expression corresponded to the distribution of thalamocortical afferents. At the same stage of the reeler mutant, expression of NGF receptor immunoreactivity was confined to the afferent axons that had grown through the neocortical primordium towards the superplate. In the control at E17, highly polysialylated NCAM (NCAM-H), a homophilic cell adhesion molecule, was expressed in the subplate, marginal zone and afferent axons. In the reeler mutant at the same stage, this adhesion molecule was expressed in both the superplate and the bundles of the afferent axons. These findings suggest that the subplate and the superplate, which are composed of neurons generated at the earliest stage, attract growing thalamocortical afferent axons specifically by a chemotropic mechanism through the expression of NGF receptor.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent Pathways↗