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Biomedical subjects

Y Kawazoe

Publications and source records attributed to Y Kawazoe.

At least 19 recordsLinked to original sources
PubMed2001

Signal transducer and activator of transcription (STAT)-induced STAT inhibitor 1 (SSI-1)/suppressor of cytokine signaling 1 (SOCS1) inhibits insulin signal transduction pathway through modulating insulin receptor substrate 1 (IRS-1) phosphorylation.

Signal transducer and activator of transcription (STAT)-induced STAT inhibitor 1 (SSI-1) is known to function as a negative feedback regulator of cytokine signaling, but it is unclear whether it is involved in other biological events. Here, we show that SSI-1 participates and plays an important role in the insulin signal transduction pathway. SSI-1-deficient mice showed a significantly low level of blood sugar. While the forced expression of SSI-1 reduced the phosphorylation level of insulin receptor substrate 1 (IRS-1), SSI-1 deficiency resulted in sustained phosphorylation of IRS-1 in response to insulin.Furthermore, SSI-1 achieves this inhibition both by binding directly to IRS-1 and by suppressing Janus kinases. These findings suggest that SSI-1 acts as a negative feedback factor also in the insulin signal transduction pathway through the suppression of IRS-1 phosphorylation.

Animals↗
PubMed2000

Hepatocarcinogen quinoline induces G:C to C:G transversions in the cII gene in the liver of lambda/lacZ transgenic mice (MutaMouse).

Quinoline is carcinogenic to the liver in rodents, but it is not clear whether it acts by a genotoxic mechanism. We previously demonstrated that quinoline does induce gene mutation in the liver of lambda/lacZ transgenic mice. In the present report, we reveal the molecular nature of the mutations induced by quinoline in the lambda cII gene, which is also a phenotypically selectable marker in the lambda transgene. (The cII gene has 294bp, which enables much easier sequence analysis than the original lacZ gene (3kb)). The liver cII mutant frequency was nine times higher in quinoline-treated mice than in control mice. Sequence analysis revealed that quinoline induced primarily G:C to C:G transversions (25 of 34). Thus, we have confirmed that quinoline is genotoxic in its target organ, and the G:C to C:G transversion is the molecular signature of quinoline-induced mutations.

Animals↗
PubMed2000

Origin of anomalous lattice expansion in oxide nanoparticles

Anomalous lattice expansions have been measured for the first time in monodisperse CeO2-x nanoparticles and in BaTiO3 single nanoparticles by electron diffraction. X-ray photoelectron spectroscopy studies on CeO2-x nanoparticles and ab initio computer simulation on BaTiO3 clusters show that the origin of expansion is the decrease of electrostatic force caused by valence reduction of Ce ions and the increase in ionicity of Ti ions, respectively. The lattice constant change of oxide (ionic) nanoparticles with the increase in ionicity would depend on the structure of the particles. Hence, first-principles calculations of large ionic clusters are indispensable.

Journal Article↗
PubMed2000

Defective thymocyte development and perturbed homeostasis of T cells in STAT-induced STAT inhibitor-1/suppressors of cytokine signaling-1 transgenic mice.

Previous experiments have shown that STAT-induced STAT inhibitor-1 (SSI-1; also named suppressors of cytokine signaling-1 (SOCS-1) or Janus kinase binding protein) is predominantly expressed in lymphoid organs and functions in vitro as a negative regulator of cytokine signaling. To determine the function of SOCS-1 in vivo, we generated SSI-1 transgenic mice using the lck proximal promoter that drives transgene expression in T cell lineage. In thymocytes expressing SSI-1 transgene, tyrosine phosphorylation of STATs in response to cytokines such as IFN-gamma, IL-6, and IL-7 was inhibited, suggesting that SSI-1 suppresses cytokine signaling in primary lymphocytes. In addition, lck-SSI-1 transgenic mice showed a reduction in the number of thymocytes as a result of the developmental blocking during triple-negative stage. They also exhibited a relative increase in the percentage of CD4+ T cells, a reduction in the number of gammadelta T cells, as well as the spontaneous activation and increased apoptosis of peripheral T cells. Thus, enforced expression of SSI-1 disturbs the development of thymocytes and the homeostasis of peripheral T cells. All these features of lck-SSI-1 transgenic mice strikingly resemble the phenotype of mice lacking common gamma-chain or Janus kinase-3, suggesting that transgene-derived SSI-1 inhibits the functions of common gamma-chain-using cytokines. Taken together, these results suggest that SSI-1 can also inhibit a wide variety of cytokines in vivo.

Animals↗
PubMed2000

Adenoviral vector-mediated GDNF gene transfer prevents death of adult facial motoneurons.

We examined neuroprotective effects of an adenoviral vector encoding glial cell line-derived neurotrophic factor (AxCAhGDNF) on the lesioned adult rat facial motoneurons. After facial nerve avulsion, animals locally injected into the facial canal with AxCALacZ (adenovirus encoding beta-galactosidase gene) or AxCAhGDNF showed expression of beta-galactosidase activity or intense immunolabeling for GDNF in lesioned facial motoneurons, respectively. The treatment with AxCAhGDNF after avulsion significantly prevented the loss of lesioned facial motoneurons, ameliorated choline acetyltransferase immunoreactivity, and suppressed the activity of nitric oxide synthase in these neurons. These results indicate that the adenovirus-mediated gene transfer of GDNF may prevent the degeneration of motoneurons in adult humans with peripheral nerve injury and motor neuron diseases.

Adenoviridae↗
PubMed2000

Rescue of lesioned adult rat spinal motoneurons by adenoviral gene transfer of glial cell line-derived neurotrophic factor.

Glial cell line-derived neurotrophic factor (GDNF) has been shown to protect cranial and spinal motoneurons, that suggests potential uses of GDNF in the treatment of spinal cord injury and motor neuron diseases. We examined neuroprotective effect of human GDNF encoded by an adenovirus vector (AxCAhGDNF) on the death of lesioned adult rat spinal motoneurons. The seventh cervical segment (C7) ventral and dorsal roots and dorsal root ganglia of adult Fisher 344 rats were avulsed, and AxCAhGDNF, AxCALacZ (adenovirus encoding beta-galactosidase gene) or PBS was inoculated in C7 vertebral foramen. One week after the avulsion and treatment with AxCALacZ, the animals showed expression of beta-galactosidase activity in lesioned spinal motoneurons. Animals avulsed and treated with AxCAhGDNF showed intense immunolabeling for GDNF in lesioned spinal motoneurons and expression of virus-induced human GDNF mRNA transcripts in the lesioned spinal cord tissue. Nissl-stained cell counts revealed that the treatment with AxCAhGDNF significantly prevented the loss of lesioned ventral horn motoneurons 2 to 8 weeks after avulsion, as compared to AxCALacZ or PBS treatment. Furthermore, the AxCAhGDNF treatment ameliorated choline acetyltransferase immunoreactivity in the lesioned motoneurons after avulsion. These results indicate that the adenovirus-mediated gene transfer of GDNF may prevent the degeneration of motoneurons in adult humans with spinal cord injury and motor neuron diseases.

Adenoviridae↗
PubMed2000

Signals transducers and activators of transcription (STAT)-induced STAT inhibitor-1 (SSI-1)/suppressor of cytokine signaling-1 (SOCS-1) suppresses tumor necrosis factor alpha-induced cell death in fibroblasts.

Signal transducers and activators of transcription (STAT)-induced STAT inhibitor-1 [SSI-1; also known as suppressor of cytokine signaling-1 (SOCS-1)] was identified as a negative feedback regulator of Janus kinase-STAT signaling. We previously generated mice lacking the SSI-1 gene (SSI-1 -/-) and showed that thymocytes and splenocytes in SSI-1 -/- mice underwent accelerated apoptosis. In this paper, we show that murine embryonic fibroblasts lacking the SSI-1 gene are more sensitive than their littermate controls to tumor necrosis factor-alpha (TNF-alpha)-induced cell death. In addition, L929 cells forced to express SSI-1 (L929/SSI-1), but not SSI-3 or SOCS-5, are resistant to TNF-alpha-induced cell death. Furthermore L929/SSI-1 cells treated with TNF-alpha sustain the activation of p38 mitogen-activated protein (MAP) kinase. In contrast, SSI-1 -/- murine embryonic fibroblasts treated with TNF-alpha show hardly any activation of p38 MAP kinase. These findings suggest that SSI-1 suppresses TNF-alpha-induced cell death, which is mediated by p38 MAP kinase signaling.

Androstadienes↗
PubMed2000

Safe and efficient gene transfer into porcine hepatocytes using Sendai virus-cationic liposomes for bioartificial liver support.

Establishment of a bioartificial liver support system using genetically modified hepatocytes is a potential approach to improve the treatment of severe liver failure. We describe the development of an efficient ex vivo method of gene transfer into a large number of porcine hepatocytes using hemagglutinating virus of Japan (HVJ)-liposome. The transfection efficiency of HVJ-liposome into isolated porcine hepatocytes attached to microcarrier beads was evaluated by beta-galactosidase (beta-gal) staining, fluorescence activated cell sorting analysis for beta-gal and luciferase assay, respectively. To examine the function and cellular damage of transduced hepatocytes, we used enzyme-linked immunosorbent assay for porcine albumin synthesis, lidocaine clearance test (P-450 activity), aspartate aminotransferase, and lactic dehydrogenase release assays. The optimal conditions for gene transfer into the beads-attached hepatocytes using HVJ-liposome included 4 microg of deoxyribonucleic acid with 200 microg of lipid/2 x 105 cells and exposure duration of 90 min. Under these conditions, beta-gal and luciferase genes were transduced to 2.5 x 108 isolated porcine hepatocytes following attachment to the beads. Positive beta-gal staining was observed in more than 30% of the beads-attached hepatocytes. The gene transfer activity of HVJ-liposome method determined by luciferase activities was about 100-fold of that of the lipofection method. Transfected porcine hepatocytes remained functional without any significant cell damage. Our results demonstrated that HVJ-liposome mediated gene transfer into microcarrier-attached porcine hepatocytes is an efficient and nontoxic method suitable for a bioartificial liver support sytem.

Animals↗
PubMed2000

Clastogenicity of quinoline derivatives tested by micronucleus induction in vivo in the hepatocytes of partially hepatectomized mice.

The induction of micronucleated liver cells (MN-liver cell) was examined with halogenated and hydroxylated quinolines using partially hepatectomized mice. Among the chloroquinolines, 8-chloroquinoline demonstrated a significantly higher level of induction than the control. All the fluorinated derivatives examined, except for 6-fluoroquinoline, induced significantly higher levels, and there were no appreciable differences in MN-liver cell induction among the fluorinated quinolines, regardless of their mutagenic potencies in the Ames test. Of the hydroxylated quinolines examined, 2- and 4-isomers, which are not mutagenic, induced MN-liver cells to the same extent as a mutagenic isomer, 8-hydroxyquinoline. It seems that clastogenicity was not satisfactorily correlated with mutagenicity in the Ames test as far as this class of compounds is concerned.

Animals↗
PubMed1999

Ubiquitous and cell-specific members of the avian small heat shock protein family.

Small heat shock proteins (sHsps) have been suggested to act as molecular chaperones for many kinds of substrates and have protective roles in cells exposed to external stresses. Unlike other major Hsps such as Hsp70 and Hsp90, expression of many vertebrate sHsps is restricted to the muscle tissues and/or eye lens. Among the sHsps, the heat-inducible human Hsp27 (hHsp27) homologue is believed to be expressed ubiquitously in various cell types. Here, we distinguished the chicken homologue of hHsp27 (cHsp24) from the chicken major heat-inducible protein of molecular size 25 kDa (cHsp25). cHsp25 is not expressed in the absence of stress, but is highly expressed after hyperthermia in all tissues of developing embryos. In contrast, expression of cHsp24 is restricted to some specific tissues even in the presence of stress. Thus, cHsp25 is the first member of the sHsps in vertebrates the expression of which is ubiquitous in tissues exposed to external stresses similar to Hsp70.

Animals↗
PubMed1999

Regression of hepatocellular carcinoma in vitro and in vivo by radiosensitizing suicide gene therapy under the inducible and spatial control of radiation.

To improve the efficacy and selectivity of gene therapy for hepatocellular carcinoma (HCC), we designed a strategy for suicide gene therapy in conjunction with radiation therapy using an HVJ-liposome vector system. The radio-inducible suicide gene was constructed by insertion of the early growth response gene 1 (Egr-1) promoter upstream of the HSV-tk gene (EGF-tk). First, to test the tumor specificity of Egr-1, RT-PCR and immunohistochemistry were performed. The Egr-1 gene was highly expressed in HCC compared with normal liver, where expression was barely detectable. Next, radiation-inducible activity of the Egr-1 promoter was examined in primary cultured normal hepatocytes and human hepatoma cell lines Huh7, HepG2, and PLC/PRF/5 by luciferase assay as a reporter gene system. Egr-1 promoter activity was markedly increased in hepatoma cell lines in a radiation dose-dependent manner, with maximum activation (15- to 28-fold) 12 hr after irradiation. In contrast, only a twofold increase in activation was noted in normal hepatocytes. An in vitro gene therapy experiment showed that EGR-tk-transduced hepatoma cells became highly sensitive to ganciclovir (GCV) after irradiation, but not without irradiation. GCV with or without irradiation did not show any cytotoxic effects against control gene-transfected cells. In addition, a "radiosensitization effect" was also demonstrated by combination therapy with the HSV-tk/GCV system and irradiation. To examine the efficacy of this EGR-tk/GCV gene therapy in vivo, xenografted liver tumors in nude mice were targeted using the HVJ-liposome vector system. EGR-tk-transfected tumors regressed significantly after a combination therapy of irradiation and GCV in all mice (n = 8), and almost disappeared in 3 weeks without any side effects. In comparison, tumors continued to grow in all mice (n = 8 in each group) treated by transfer of EGR-tk followed by either irradiation without GCV or GCV without irradiation. Our data indicate that HSV-tk gene therapy under the control of a radioinducible promoter is effective, and might be selective for hepatoma cells because of its inducible and radiosensitive capacity after radiation exposure as well as its tumor-specific activation.

Aged↗
PubMed1999

Substituent effect of a fluorine atom on the mutagenicity of nitroquinolines.

Some 16 nitroquinolines (NQs) and their fluorinated derivatives were tested for mutagenicity in Salmonella typhimurium TA100 without S9 mix to investigate the effect of fluorine-substitution on the mutagenicity. These NQs consist of 5-NQs, 5-nitroquinoline N-oxides (5-NQOs), N-methyl-5-nitroquinolinium methanesulfonates (N-Me-5-NQs) and 8-NQs, including three ortho-F-NQs, one meta-F-NQ, four para-F-NQs and four 3-F-NQs. For this purpose, eight F-NQs were newly synthesized. The data indicated that the ratio of the mutagenic activities (revertants/plate/nmol) of fluorinated NQs to those of the corresponding parent non-fluorinated compounds ranged from 0.6- to 119-fold. The fluorine atom located para to the nitro group markedly enhanced the mutagenicity (24-fold and more), while three ortho-fluorinated derivatives showed no significant increase in mutagenicity (enhancement ratio were 0.6, 0.8 and 1.7). With respect to 8-NQs, its meta-fluorinated derivative also had an enhanced mutagenicity over the parent compound (53-fold). In addition, although N-Me-5-NQ was less mutagenic than 5-NQ and 5-NQO, the mutagenicity of N-Me-5-NQ was most significantly enhanced by fluorine-substitution. These results suggest that introduction of a fluorine atom to the molecule in question may be a useful tool to modify their mutagenic potency and to better understand the mechanism of mutation.

Chromatography, Gel↗
PubMed1999

Effects of oligofluorine substitution on the mutagenicity of quinoline: a study with twelve fluoroquinoline derivatives.

A total of 12 variously fluorinated derivatives of quinoline (Q) were tested for their mutagenicity in Salmonella typhimurium TA100 in the presence of S9 mix to investigate the structure-mutagenicity relationship in oligofluorinated quinolines. Nine of them, 3,7-di-, 5,6-di-, 6,7-di-, 6,8-di-, 7,8-di-, 3,5,7-tri-, 5,6,8-tri-, 6,7, 8-tri-, and 5,6,7,8-tetrafluoroquinolines (FQs), were newly synthesized for this purpose. Those fluorinated at position 3 were all non-mutagenic. Mutagenicity was enhanced by fluorine-substitution at position 5 or 7, but not in 3-FQs (i.e., 3, 5-di-, 3,7-di-, and 3,5,7-triFQs). Some of the 6-fluorinated derivatives showed less maximum induced-revertants with more mutagenic potencies in terms of induced-revertants per dose than quinoline. No marked change occurred by fluorine-substitution at position 8. These results show that the effect of di- and trifluoro-substitution on mutagenicity is generally additive, while that of tetrafluorination approaches the deactivating effect of perfluorination. Our study suggests that 3-fluorine-substitution in the pyridine moiety may be a useful means of antimutagenic structural modification in pyridine-fused aromatic chemicals for medicinal and agricultural use.

Animals↗
PubMed1999

HSF3 is a major heat shock responsive factor duringchicken embryonic development.

The expression of heat shock genes in vertebrates is regulated mainly at the level of transcription by four heat shock transcription factors (HSFs 1-4). Avian cells express at least three HSFs (HSFs 1-3). HSF1 is rapidly activated by even mild heat shock, while HSF3 is activated only by severe heat shock. In contrast, HSF2 is not activated by heat stress and has been speculated to have developmental functions. Here, we examined the temporal and spatial profiles of changes in the levels of these three HSFs in various tissues during chicken development. We found that HSF3 was almost constantly expressed in various tissues during early to late chicken development. The expression of HSF1 was equally high in most tissues early in development and thereafter declined to different levels in a tissue-dependent manner. Thus, HSF3 became the dominant heat-responsive factor in all tissues examined. The magnitude of heat shock response determined by Northern blotting did not always correlate with the level of HSF1 expression, suggesting that not only HSF1 but also HSF3 may be a major factor mediating stress signals to heat shock gene expression in the chicken. In addition, the high-level and ubiquitous expression of HSF2 as well as HSF1 and HSF3 in early embryogenesis suggested the involvement of these factors in all developmental processes.

Animals↗
PubMed1999

Effects of anticoagulants on porcine hepatocytes in vitro: implications in the porcine hepatocyte-based bioartificial liver.

For the clinical treatment with porcine hepatocyte-based bioartificial liver (BAL), the use of an anticoagulant in the extracorporeal system is essential. In this experiment, we studied the effect of various anticoagulants on cultured porcine hepatocytes. Porcine hepatocytes were isolated and seeded at a density of 2 x 10(5) cells on a collagen-coated plate in Dulbecco's modified Eagle's medium (DMEM) with 10% fetal calf serum (FCS). Twenty-four hours later, the medium was changed to DMEM with various anticoagulants such as nafamostat mesilate (NM), sodium heparin (SH) and sodium citrate (SC) at concentration used clinically. As a control, the hepatocytes were cultured in only DMEM. After culturing for 6 hours, the viability of the porcine hepatocytes, lactate dehydrogenase (LDH) release, lidocaine clearance (cytochrome p450 function) and albumin synthesis were investigated. SC did not affect either the viability or the p450 function of the hepatocytes. In the NM group, the viability of porcine hepatocytes and lidocaine clearance were decreased significantly more than in the other groups. SH did not affect the viability of porcine hepatocytes, however, it seemed to reduce the p450 function. In conclusion, SC may therefore be the optimal anticoagulant available for hepatocyte-based BAL circuit in terms of its cell toxicity.

Albumins↗