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Tsutomu Takegami

Publications and source records attributed to Tsutomu Takegami.

10 recordsLinked to original sources
PubMed2006

RhoGDIbeta lacking the N-terminal regulatory domain suppresses metastasis by promoting anoikis in v-src-transformed cells.

Rho guanine nucleotide dissociation inhibitors (RhoGDIs) regulate the activity of Rho family GTPases. RhoGDIbeta (LyGDI/GDID4/RhoGDI2) has two caspase cleavage sites after Asp19 and Asp55. The resulting cleavage products, DeltaN(1-19)RhoGDIbeta and DeltaN(1-55)RhoGDIbeta, are expressed in cells under conditions that activate caspases. DeltaN(1-19)RhoGDIbeta, which can inhibit GDP dissociation, is implicated in the process of apoptosis, whereas the physiological roles for DeltaN(1-55)RhoGDIbeta, which lacks the ability to inhibit GDP dissociation, are largely unknown. To explore the roles of DeltaN(1-55)RhoGDIbeta, we examined the phenotypes of v-src-transformed metastatic fibroblasts transfected with plasmids for expressing DeltaN(1-55)RhoGDIbeta. Although the expression of DeltaN(1-55)RhoGDIbeta had no effect on the rate of growth in vitro, it suppressed experimental metastasis and decreased the rate of growth in vivo. In addition, DeltaN(1-55)RhoGDIbeta-expressing cells had enhanced adhesion to fibronectin, laminin, and collagens but reduced retention in the lung after intravenous injection. Also, the expression of DeltaN(1-55)RhoGDIbeta promoted anoikis without affecting the levels of activated Rac1 or Cdc42. Furthermore, DeltaN(1-55)RhoGDIbeta did not affect the expression or phosphorylation of focal adhesion kinase, p44/p42 mitogen-activated protein kinases, or Akt1 before or after induction of anoikis. Thus, DeltaN(1-55)RhoGDIbeta appears to promote anoikis by undefined mechanisms, thereby suppressing metastasis in v-src-transformed fibroblasts.

Animals↗
PubMed2006

Activation of Rac1 by Rho-guanine nucleotide dissociation inhibitor-beta with defective isoprenyl-binding pocket.

Rho-guanine nucleotide dissociation inhibitor-beta (RhoGDIbeta), a regulator for Rho GTPases, is implicated in cancer cell progression. We reported that C-terminal truncated RhoGDIbeta (DeltaC(166-201)-RhoGDIbeta) promoted metastasis through activating Rac1 signaling pathway in ras-transformed fibroblast cells. To better understand the mechanism of Rac1 activation by DeltaC(166-201)-RhoGDIbeta during metastasis, the amount of GTP-bound Rac1 was measured as the activation level of Rac1 in cells expressing various mutant RhoGDIbeta with sequential C-terminal deletions. Three C-terminal hydrophobic amino acid residues (Trp191, Leu193, and Ile195) supposed to interact with isoprenyl groups of Rac1, was indispensable for a proper regulation of Rac1 activation/inhibition. Deletion of this region led RhoGDIbeta to continuously associate with GTP-bound Rac1, provoking constitutive activation of Rac1. Thus, impaired interaction of RhoGDIbeta with Rac1 isoprenyl groups possibly makes RhoGDIbeta function as a positive regulator for Rac1 during metastasis.

Amino Acid Sequence↗
PubMed2006

Microvessel density: correlation with 18F-FDG uptake and prognostic impact in lung adenocarcinomas.

UNLABELLED: Although researched for many years, the prognostic value of tumor angiogenesis reflected by microvessel density (MVD) is still controversial, and there have been no previous reports regarding the correlation with 18F-FDG uptake in lung adenocarcinomas. Therefore, in the present study, we investigated the correlation between MVD determined with different endothelial cell antibodies and 18F-FDG uptake and compared the prognostic impact of those factors in lung adenocarcinomas. METHODS: Forty-four patients with 45 lung adenocarcinomas underwent 18F-FDG PET before surgery. Consecutive paraffin-embedded sections obtained from each resected tumor were immunostained for CD31 (a panendothelial cell marker), CD105 (a proliferation-related endothelial cell marker), and CD34/alpha-SMA (for double labeling of endothelial cells and mural cells). Four high-power fields in the area with the highest MVD were selected for analysis. Computer-assisted image analysis was used to assess MVD. RESULTS: MVD staining results for panendothelial cell markers can be classified into 3 microvessel patterns: diffuse, alveolar, and mixed. The highly ordered alveolar pattern is believed to represent preexisting alveolar vessels trapped in lung adenocarcinomas and may have no significant meaning for the aggressiveness of tumors. Preexisting alveolar cells also do not contribute to 18F-FDG uptake. CD105 staining of MVD (CD105-MVD) showed a significantly positive correlation with 18F-FDG uptake (P < 0.0001), whereas CD31 staining of MVD (CD31-MVD) showed a marginally negative correlation with it (P = 0.057). Although CD105-MVD correlated negatively with prognosis, patients with low CD105-MVD, compared with those with high or moderate CD105-MVD, had a much better prognosis in both disease-free and overall survival analyses (P = 0.017 and P = 0.013, respectively). Patients with low CD31-MVD had the worst prognosis (P = 0.032 for disease-free survival analysis and P = 0.179 for overall survival analysis). CONCLUSION: There is no positive correlation between 18F-FDG uptake and MVD determined with panendothelial cell markers (CD31 and CD34); in contrast, there is a marginally negative correlation between them. MVD determined with CD105, which is a proliferation-related endothelial cell marker, reflects active angiogenesis, correlates positively with 18F-FDG uptake, and is a better indicator of prognosis in lung adenocarcinomas.

Adenocarcinoma↗
PubMed2005

MRI features of extraskeletal myxoid chondrosarcoma.

OBJECTIVE: To describe the MRI features of extraskeletal myxoid chondrosarcoma in comparison with clinicopathologic findings. DESIGN AND PATIENTS: The study comprised 12 male subjects and seven female subjects with a mean age of 53 years (range 16-76 years). MRI findings, evaluated by two radiologists with agreement by consensus, were compared for histopathologic features. RESULTS: The tumor size ranged from 2.0 cm to 20.0 cm (mean 8.9 cm). Fusion gene transcripts could be detected in 13 (68%) of the 19 cases: EWS-CHN in nine cases, TAF2N-CHN in three, and TFG-TCH in one. There were six fusion-negative cases. Signal characteristics on T1-weighted and T2-weighted MR images were non-specific with regard to each cytogenetic variant. Peripheral enhancement was seen more frequently in tumors with the EWS-CHN variant than in those with other cytogenetic variants. The characteristic pattern of enhancement corresponded to the presence of fibrous septa and peripheral areas of high cellularity within lobules, by correlation with pathologic findings. All cases with TAF2N-CHN or TFG-TCH variants showed invasion of extracompartmental structure, bone, or vessels. CONCLUSION: Extraskeletal myxoid chondrosarcoma is an uncommon soft-tissue malignancy that may be recognized by MRI features of multi-lobular soft-tissue mass often invading extracompartmental, bony, and vascular structures.

Adolescent↗
PubMed2005

Capacitative Ca2+ entries and mRNA expression for TRPC1 and TRPC5 channels in human epidermoid carcinoma A431 cells.

In human epidermoid carcinoma A431 cells, capacitative Ca2+ entries in response to intracellular Ca2+ store depletion with thapsigargin, an endoplasmic reticulum Ca(2+)-ATPase inhibitor, and uridine 5'-triphosphate, a phospholipase C-linked agonist, were inhibited by trivalent cations such as Gd3+ and La3+, and by the store-operated Ca2+ channel inhibitor, 2-aminoethoxydiphenyl borate. Of the seven types of canonical transient receptor potential (TRPC) channels as molecular candidates for store-operated Ca2+ channels, mRNAs for TRPC1 and TRPC5 were detected in the cells with the reverse transcription-polymerase chain reaction. Western blotting confirmed the protein expressions of TRPC1 and TRPC5 in A431 cells. The present results suggest that TRPC1 and/or TRPC5 channels serve as store-operated Ca2+ channels in A431 cells, and may function as regulators for intracellular Ca2+ signaling.

Base Sequence↗
PubMed2005

Inhibitory effect of RNAi on Japanese encephalitis virus replication in vitro and in vivo.

Flaviviruses include many insect-mediated small viruses and still cause serious problems in the world. In humans, JEV can cause acute meningioencephalomyelitis, resulting in fatality rates of 5 to 40%. RNA-interference (RNAi) as an antiviral mechanism was originally discovered in plants and then found in the specific suppression of gene expression of other organisms such as Caenorhabditis elegans, Drosophila and vertebrates. As JEV is an RNA virus, RNAi could be a reasonable approach for therapeutic purposes to use against Japanese encephalitis. In this study, we examined the effect of RNAi on JEV replication. Viral reproduction in Vero cells was decreased to 7.2% and 39.0% of control by the transfection of small interference RNAs, JCR and JN3R at 250 n M, respectively. Under the transfection of 5 microg/ml pJRi which produces stem-loop RNAi, viral reproduction was decreased to about 10% of control. Western blot analysis indicated that RNAi inhibited the translation level. We used pJRi in the animal experiment. After the inoculation of viruses at 5 x 10(3) PFU, pJRi at 1.0 and 5.0 microg/g was injected into mice i.p. JEV-infected control mice (n=5) died within 15 days. pJRi (1.0 or 5.0 microg/g)-medicated mice survived 40 or 80% at 15 days. The data clearly indicate that pJRi has highly potent inhibitory activity against JEV replication in vivo. The results in vivo and in vitro provide evidence that JEV replication was efficiently inhibited by RNAi and RNAis could be used as an antiviral drug against JEV infection.

Animals↗
PubMed2003

Hepatitis C virus nonstructural protein NS3 binds to Sm-D1, a small nuclear ribonucleoprotein associated with autoimmune disease.

Hepatitis C virus (HCV) causes a persistent infection, chronic hepatitis, and leads to hepatocellular carcinoma. Nonstructural protein 3 (NS3) of HCV possesses protease, nucleoside triphosphatase, and helicase activities. Using the yeast two hybrid assay, we identified Sm-D1, a host protein that binds to NS3. Sm-D1 is a component of small nuclear ribonucleoprotein (snRNP) complexes which are associated with autoimmune disease. Sm-D1 has Gly-Arg (GR) repeats at the C terminus, which contains dimethylarginine modified by post-translational modification and may constitute an immunoreactive determinant. Deletion mutants revealed that the C-terminal region of Sm-D1 containing the GR repeats was the binding region for NS3, and the expression feature of Sm-D1 was affected by co-expression of NS3. Immunostaining assay demonstrated that NS3 was also present in the nucleus of cells overexpressing Sm-D1, although it was usually found in cytoplasm. The localization of NS3 could change following interaction with Sm-D1 and affect the function of Sm-D1.

Autoimmune Diseases↗
PubMed2002

Characterization and E protein expression of mutant strains during persistent infection of KN73 cells with Japanese encephalitis virus.

OBJECTIVE: To study the character of mutants originating from Japanese encephalitis viruses and the relationship between the characterization of mutant strains and E protein expression. METHODS: Persistent infection was established with standard strains of Japanese encephalitis viruse, known as parental viruse, in a human hepatoma cell line, KN73. Cells were subcultured weekly using trypsinization techniques. Cell-associated viruses of persistently infected cells were collected by a freeze and thaw method. Virus titers were examined by plaque method using baby hamster kidney (BHK) cells. Indirect immunofluorescence assays were used to examine E and NS3 protein antigens. Western blot analysis was used to test expression of E and NS3 proteins. RESULTS: In the early phase (24 - 36 h) post-infection, virus titer in culture fluid from KN73 cells infected with parental viruses were 10(6) PFU/ml. They were 10(3 - 4) PFU/ml in the late phase (3 years) post-infection. The titer of cell-associated viruse was 10(2 - 3) PFU/ml. A virus super-infection assay found that virus titers in culture fluid from persistently infected KN73 cells acutely super- infected with parental viruses were much lower than that of culture fluids in acutely infected normal KN73 at the same phase. Indirect immunoflurescence assay revealed that the quantity of viral antigens in persistently infected KN73 cells was lower than that in acutely infected KN73 cells with parental viruses. Western blot analyses indicated that the molecular weights of E and NS3 proteins were 53 kD and 73 kD, respectively. Expression of NS3 protein in persistently infected KN73 cells was stable but expression of E protein was markedly suppressed. CONCLUSIONS: The virulence and reproduction of viruses obtained from persistently infected KN73 cells, which have some features of DI viruses and were involved in persistent infection, was lower than that of parental viruses. These mutants may have be related to the decrease in E protein expression.

Carcinoma, Hepatocellular↗