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Biomedical subjects

Kouichi Itoh

Publications and source records attributed to Kouichi Itoh.

At least 19 recordsLinked to original sources
PubMed2006

Critical role of PICT-1, a tumor suppressor candidate, in phosphatidylinositol 3,4,5-trisphosphate signals and tumorigenic transformation.

The tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN) regulates diverse cellular functions by dephosphorylating the lipid second messenger, phosphatidylinositol 3,4,5-trisphosphate (PIP(3)). Recent study revealed that PICT-1/GLTSCR2 bound to and stabilized PTEN protein in cells, implicating its roles in PTEN-governed PIP(3) signals. In this study, we demonstrate that RNA interference-mediated knockdown of PICT-1 in HeLa cells down-regulated endogenous PTEN and resulted in the activation of PIP(3) downstream effectors, such as protein kinase B/Akt. Furthermore, the PICT-1 knockdown promoted HeLa cell proliferation; however the proliferation of PTEN-null cells was not altered by the PICT-1 knockdown, suggesting its dependency on PTEN status. In addition, apoptosis of HeLa cells induced by staurosporine or serum-depletion was alleviated by the PICT-1 knockdown in the similar PTEN-dependent manner. Most strikingly, the PICT-1 knockdown in HeLa and NIH3T3 cells promoted anchorage-independent growth, a hallmark of tumorigenic transformation. Furthermore, PICT-1 was aberrantly expressed in 18 (41%) of 44 human neuroblastoma specimens, and the PICT-1 loss was associated with reduced PTEN protein expression in spite of the existence of PTEN mRNA. Collectively, these results suggest that PICT-1 plays a role in PIP(3) signals through controlling PTEN protein stability and the impairment in the PICT-1-PTEN regulatory unit may become a causative factor in human tumor(s).

Animals↗
PubMed2006

Monoclonal antibody Rip specifically recognizes 2',3'-cyclic nucleotide 3'-phosphodiesterase in oligodendrocytes.

The antigen recognized with monoclonal antibody (mAb) Rip (Rip-antigen) has been long used as a marker of oligodendrocytes and myelin sheaths. However, the identity of Rip-antigen has yet to be elucidated. We herein identified the Rip-antigen. No signal recognized by mAb-Rip was detected by immunoblot analyses in the rat brain, cultured rat oligodendrocytes, or the oligodendrocyte cell line CG-4. As this antibody worked very well on immunocytochemistry and immunohistochemistry, Rip-antigen was immunopurified with mAb-Rip from the differentiated CG-4 cells. Eight strong-intensity bands thus appeared on 5-20% SDS-PAGE with SYPRO ruby fluorescence staining. To identify these molecules, each band extracted from the gel was analyzed by MALDI-QIT/TOF mass spectrometry. We found an interesting molecule in the oligodendrocytes from an approximately 44-kDa band as 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP). To test whether CNP was recognized by mAb-Rip, double-immunofluorescence staining was performed by using Alexa Fluor 488-conjugated mAb-Rip and Alexa Fluor 568-conjugated mAb-CNP in the rat cerebellum, mouse cerebellum, cultured rat oligodendrocytes, and CG-4 cells. The Rip-antigen was colocalized with CNP in these cells and tissues. To provide direct evidence that CNP was recognized by mAb-Rip, rat Cnp1-transfected HEK293T cells were used for double-immunofluorescence staining with mAb-Rip and mAb-CNP. The Rip-antigen was colocalized with CNP in rat Cnp1-transfected HEK293T cells, but the antigen was not detected by mAb-Rip and mAb-CNP in mock-transfected HEK293T cells. Overall, we have demonstrated that the antigen labeled with mAb-Rip is CNP in the oligodendrocytes.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed2006

Expression of new human inorganic pyrophosphatase in thyroid diseases: its intimate association with hyperthyroidism.

Inorganic pyrophosphatase (PPase) controls the level of inorganic pyrophosphate produced by biosynthesis of protein, RNA, and DNA. Thus, PPase is essential for life. PPase expression is unclear in the thyroid. We cloned a new human PPase, phospholysine phosphohistidine inorganic pyrophosphate phosphatase (LHPPase), and established a rabbit polyclonal anti-LHPPase antibody. This is the first study to determine the PPase expression by immunohistochemistry and Western blot. Intranuclear LHPPase expression of thyrocytes was enhanced most prominently in Graves' disease and autonomously functional thyroid nodule. To estimate a regulating factor of subcellular localization of LHPPase, we examined its expression of Graves' disease-derived thyrocytes in vitro with the disease-originated serum. Nuclear expression of LHPPase was lost in cultured thyrocytes even with the serum, while its cytoplasmic expression was retained. The data suggest that increased expression of LHPPase is associated with hyperthyroidism. Intranuclear expression of LHPPase may not be regulated by Graves' disease-derived serum factors.

Antibodies↗
PubMed2006

Sentinel node detection in breast cancer using contrast-enhanced sonography with 25% albumin--Initial clinical experience.

PURPOSE: To determine the clinical usefulness of a sentinel lymph node (SLN) identification technique using contrast-enhanced sonography (CEUS) with 25% albumin. METHODS: The subjects were 23 women with breast cancer. Each was injected subcutaneously with 5 ml of 25% albumin solution as a negative contrast agent directly superficial to the breast tumor. The area was massaged, and the inferior axillary hairline was examined continuously using gray-scale sonography with a 7.5- or 10-MHz transducer. Any contrast-enhanced lymph node was considered a CEUS-detected SLN, was differentiated from other level I and II nodes, and was resected and pathologically assessed. RESULTS: In all 23 patients, 1 or 2 CEUS-detected SLNs (mean, 1.3 SLNs) were identified. Their sizes ranged from 5 mm to 25 mm (mean, 11.3 mm), and their depths (from the skin surface) ranged from 5mm to 20 mm (mean, 12.6 mm). Pathologic examination revealed a metastasis in 5 of the 23 patients, all in CEUS-detected SLNs. CONCLUSION: In a clinical study using CEUS with 25% albumin, contrast-enhanced nodes were identified in all subjects. The pathologic findings suggested that any metastatic nodes observed were SLNs, indicating that this technique may represent a new modality for SLN identification.

Adult↗
PubMed2006

Development of an ESR/MR dual-imaging system as a tool to detect bioradicals.

A system combining electron spin resonance imaging (ESRI) with another imaging modality capable of enabling visualization of the distribution of bioradicals on an anatomical map of the specimens would be a superior biomedical imaging system. We describe the development of an ESR/MR dual-imaging system with one permanent magnet and the biomedical applications of this system. The magnetic circuit developed for the ESR/MR dual-imaging system consisted of the permanent magnet made of Fe-Nd-B, pole pieces, and poke. The permanent magnet was installed on the MR side only, and the ESR side was made of pole pieces only. The magnetic field was adjusted to 0.5T at MR and to 0.042T at ESR. The overall dimensions of the magnet developed for the ESR/MR imaging system were 460 (W)x440 (D)x460 (H) mm, and it weighed 220 kg. The distance of each center for the magnet for ESR and MR imaging could be set as close as 200 mm. The entire ESR/MR imaging system can be installed in a common laboratory without magnetic shielding. MR images of plants (myoga) and small animals (mice and rats) were successfully acquired with or without ESR operation. ESR spectra of nitroxyl spin probes were also measured, even with MRI operation. ESR signals of triarylmethyl derivatives with narrow line-width (0.026 mT) were observed in living mice while MRI was operating. The ESR/MR imaging dual functions work properly with no electric or magnetic interference. The ESR/MR dual images demonstrate that this system enables visualization of the distribution of bioradicals on the anatomical map of the object.

Animals↗
PubMed2005

L1-mediated branching is regulated by two ezrin-radixin-moesin (ERM)-binding sites, the RSLE region and a novel juxtamembrane ERM-binding region.

We investigated how the neural cell adhesion molecule L1 mediates neurite outgrowth through L1-L1 homophilic interactions. Wild-type L1 and L1 with mutations in the cytoplasmic domain (CD) were introduced into L1 knock-out neurons, and transfected neurons were grown on an L1 substrate. Neurite length and branching were compared between wild-type L1 and L1CD mutations. Surprisingly, the L1CD is not required for L1-mediated neurite outgrowth but plays a critical role in neurite branching, through both the juxtamembrane region and the RSLE region. We demonstrate that both regions serve as ezrin-moesin-radixin-binding sites. A truncation mutant that deletes 110 of 114 amino acids of the L1CD still supports neurite outgrowth on an L1 substrate, suggesting that a coreceptor binds to L1 in cis and mediates neurite outgrowth and that L1-ankyrin interactions are not essential for neurite initiation or outgrowth. These data are consistent with a model in which L1 can influence L1-mediated neurite outgrowth and branching through both the L1CD and a coreceptor.

Amino Acid Motifs↗
PubMed2005

The Human Anatomic Gene Expression Library (H-ANGEL), the H-Inv integrative display of human gene expression across disparate technologies and platforms.

The Human Anatomic Gene Expression Library (H-ANGEL) is a resource for information concerning the anatomical distribution and expression of human gene transcripts. The tool contains protein expression data from multiple platforms that has been associated with both manually annotated full-length cDNAs from H-InvDB and RefSeq sequences. Of the H-Inv predicted genes, 18 897 have associated expression data generated by at least one platform. H-ANGEL utilizes categorized mRNA expression data from both publicly available and proprietary sources. It incorporates data generated by three types of methods from seven different platforms. The data are provided to the user in the form of a web-based viewer with numerous query options. H-ANGEL is updated with each new release of cDNA and genome sequence build. In future editions, we will incorporate the capability for expression data updates from existing and new platforms. H-ANGEL is accessible at http://www.jbirc.aist.go.jp/hinv/h-angel/.

Database Management Systems↗
PubMed2005

Involvement of the tumor necrosis factor (TNF)/TNF receptor system in leukemic cell apoptosis induced by histone deacetylase inhibitor depsipeptide (FK228).

Inhibition of histone deacetylase (HDAC) is a novel strategy for the treatment of leukemias via restoration of aberrantly silenced genes. In this study, we conducted a detailed analysis of anti-leukemic effects of an HDAC inhibitor (HDI), depsipeptide (FK228), using myeloid leukemia cell lines HL-60 and K562. DNA chip analysis revealed upregulation of TNF-alpha mRNA and a number of molecules involved in TNF-signaling such as TRAF-6, caspases-10, and -7 in depsipeptide-treated HL-60 cells, which prompted us to examine the involvement of the TNF/TNF receptor system in the anti-leukemic effects of the drug. Upregulation of TNF-alpha was induced by depsipeptide in HL-60 and K562 cells, which expressed type I TNF receptors (TNF-RI). Depsipeptide activated caspases-8 and -10, which in turn cleave caspases-3 and -7, leading to apoptotic cell death in both cell lines. Anti-TNF-alpha neutralizing antibody and short interfering RNA (siRNA) against TNF-RI alleviated the activation of the caspase cascade and the induction of apoptosis, indicating the presence of an autocrine loop. Finally, we demonstrated that the enhanced production of TNF-alpha by depsipeptide was due to transcriptional activation of the TNF-alpha gene through hyperacetylation of histones H3 and H4 in its promoter region (-208 to +35). These results suggest that autocrine production of TNF-alpha plays a role in the cytotoxicity of depsipeptide against a subset of leukemias.

Acetylation↗
PubMed2005

Dephosphorylation and internalization of cell adhesion molecule L1 induced by theta burst stimulation in rat hippocampus.

The neural cell adhesion molecule L1 may participate in initiating and maintaining synaptic changes during learning in the hippocampus. One prominent form of synaptic change in the hippocampus is long-term potentiation (LTP) that occurs following specific patterns of synaptic activity. We present evidence that Y1176 of the YRSL motif within L1 cytoplasmic domain is dephosphorylated in LTP-induced hippocampus. The dephosphorylated L1 is associated with AP-2 and AP180 that are required for clathrin-mediated internalization of L1. These data suggest that clathrin-mediated recycling of L1 at presynaptic sites is enhanced by certain kinds of neural activity, and that maintenance of LTP-induced synaptic changes is regulated by L1 recycling.

Adaptor Protein Complex 2↗
PubMed2005

The effects of Levovist and DD-723 in activating platelets and damaging hepatic cells of rats.

OBJECTIVE: The purpose of this study was to compare platelet activation and hepatic cell damage produced by 2 ultrasonographic contrast agents with flow cytometric and ultrastructural analysis. METHODS: Suspension samples were made by mixing Levovist (SH U508A; Schering AG, Berlin, Germany) or DD-723 (Nycomed; Amersham Health, Princeton, NJ) with whole blood. The final concentrations of Levovist in citrated whole blood were 0, 15, and 75 mg/mL, and those of DD-723 were 0, 5, and 50 microL/mL. After exposure to ultrasound in vitro, flow cytometric analysis was performed to determine the concentration of the CD62P activation-specific antigen. To compare the hepatic cell damage associated with these 2 agents, we divided 15 rats into 5 groups as follows: group 1, sham operation; group 2, Levovist injection only; group 3, DD-723 injection only; group 4, Levovist injection (contrast agent) and ultrasound exposure; and group 5, DD-723 injection and ultrasound exposure. The ultrasonographic contrast agents Levovist and DD-723 were administered through the femoral vein and sonicated continuously for the first minute; this was followed by sweeping for 5 minutes 10 seconds after the contrast agent was injected. The rats were perfused via the heart with a fixative solution immediately after the sweeping, and then the liver was excised; the specimens were studied with electron and light microscopy. RESULTS: The percentage of CD62P-expressing platelets increased in both contrast agent-ultrasound exposure groups, and the percentage of CD62P-expressing platelets was greater in the Levovist group. We observed vacuolation and round deposits in the hepatocytes in both contrast agent-ultrasound exposure groups. Microbubbles were observed in the rat Kupffer cells, and a few hepatocytes were seen unexpectedly in the DD-723 group but were found in neither the Kupffer cells nor the hepatocytes in the Levovist group. CONCLUSIONS: Both contrast agents, Levovist and DD-723, produced platelet activation and structural change in the rat hepatic cells, but only the microbubbles of DD-723 were taken up by the Kupffer cells and a few hepatocytes.

Animals↗
PubMed2005

[In vitro susceptibilites to levofloxacin and various antibacterial agents of 11,475 clinical isolates obtained from 52 centers in 2002].

The susceptibilities of bacteria to fluoroquinolones (FQs), especially levofloxacin, and other antimicrobial agents were investigated using 11,475 clinical isolates collected in Japan during 2002. Methicillin susceptible staphylococci, Streptococcus pyogenes, Streptococcus pneumoniae, Moraxella catarrhalis, the family of Enterobactericeae, Haemophilus influenzae and Acinetobacter spp. exhibited stable and high susceptibilities to FQs. The rate of FQs-resistant MRSA was 80 approximately 90%, being markedly higher than that of FQs-resistant MSSA. The FQs-resistance rate of MRCNS was also higher than that of MSCNS, however, it was lower than that of MRSA. No FQs-resistant clinical isolates of Salmonella spp. were detected in any of the surveys. Thirteen of Escherichai coli 696 isolates, 8 of Klebsiella pneumoniae 630 isolates and 33 of Proteus mirabilis 373 isolates produced extended-spectrum beta-lactamase (ESBL), furthermore 6 of 13 in E. coli, 1 of 8 in K. pneumoniae and 14 of 31 ESBL-producing isolates, and in P. mirabilis were FQs resistant. Attention should be focused in the future on the emergence of ESBL in relation to FQs resistance. The rate of FQs-resistant P. aeruginosa isolated from urinary tract infection (UTI) was 40 approximately 60%, while 15 approximately 25% of isolates from respiratory tract infection (RTI) were resistant. IMP-1 type metallo beta-lactamase producing organisms were found in 49 of P. aeruginosa 1,095 isolates, 7 of S. marcescens 586 isolates and 4 of Acinetobacter spp. 474 isolates, respectively. Glycopeptide-resistant enterococci or S. aureus was not found.

Anti-Bacterial Agents↗
PubMed2005

[Nationwide surveillance of parenteral antibiotics containing meropenem activities against clinically isolated strains in 2004].

The antibacterial activity of meropenem (MEPM) and other parenteral antibiotics against clinical isolates of 907 strains of Gram-positive bacteria, 1790 strains of Gram-negative bacteria, and 192 strains of anaerobic bacteria obtained from 30 medical institutions during 2004 was measured. The results were as follows; 1. MIC90 of MEPM for almost all of enterobacteriaceae and Haemophilus influenzae were 4-fold to 32-fold lower than those of other carbapenems. MEPM was more active than other carbapenem antibiotics against Gram-negative bacteria, especially against enterobacteriaceae and H. influenzae. MEPM were active against most of the species tested in Gram-positive and anaerobic bacteria, except for multi-drug resistant strains including methicillin-resistant Staphylococcus aureus. 2. As for Pseudomonas aeruginosa, imipenem (IPM) showed high cross-resistant rate againt meropenem-resistant P. aeruginosa (87.9%). MEPM showed low cross-resistant rate both againt IPM-resistant P. aeruginosa (49.2%) and ciprofloxacin-resistant P. aeruginosa (38.0%). 3. The proportion of extended-spectrum beta-lactamase (ESBL) strains was 3.1% (4 strains) in Escherichia coli, 8.0% (2 strains) in Citrobacter koseri, 2.5% (3 strains) in Klebsiella pneumoniae, 2.5% (2 strains) in Enterobacter cloacae, 0.9% (1 strains) in Serratia marcescens, and 2.2% (2 strains) in Proteus mirabilis. The proportion of metallo-beta-lactamase strains was 1.6% (5 strains) in P. aeruginosa. 4. Of all species tested, Peptostreptococcus spp. was the only species, which MIC90 of MEPM was more than 4-fold higher than that in our previous study using clinical isolates during 2002 (0.25 microg/ml --> 1 microg/ml). Therefore, there is almost no siginificant decrease in susceptibility of clinical isolates to meropenem. In conclusion, the results from this surveillance study suggest that MEPM retains its potent and broad antibacterial activity and therefore is a clinically useful carbapenem at present, 9 years after available for commercial use.

Anti-Infective Agents↗
PubMed2004

Essential role for phospholipase D2 activation downstream of ERK MAP kinase in nerve growth factor-stimulated neurite outgrowth from PC12 cells.

The signaling pathway that triggers morphological differentiation of PC12 cells is mediated by extracellular signal-regulated kinase (ERK), the classic mitogen-activated protein (MAP) kinase. However, mediators of the pathway downstream of ERK have not been identified. We show here that phospholipase D2 (PLD2), which generates the pleiotropic signaling lipid phosphatidic acid (PA), links ERK activation to neurite outgrowth in nerve growth factor (NGF)-stimulated PC12 cells. Increased expression of wild type PLD2 (WT-PLD2) dramatically elongated neurites induced by NGF stimulation or transient expression of the active form of MAP kinase-ERK kinase (MEK-CA). The response was activity-dependent, because it was inhibited by pharmacological suppression of the PLD-mediated PA production and by expression of a lipase-deficient PLD2 mutant. Furthermore, PLD2 was activated by MEK-CA, whereas NGF-stimulated PLD2 activation and hypertrophic neurite extension were blocked by an MEK-specific inhibitor. Taken together, these results provide evidence that PLD2 functions as a downstream signaling effector of ERK in the NGF signaling pathway, which leads to neurite outgrowth by PC12 cells.

Actins↗
PubMed2004

Decoupling of automatic control systems in a continuous-wave electron paramagnetic resonance spectrometer for biomedical applications.

This article describes a systematic approach to decoupling automatic tuning control (ATC) and automatic matching control (AMC) systems in continuous-wave (CW) electron paramagnetic resonance (EPR) spectroscopy for animal experiments. This technique enables us to improve the stability of CW-EPR spectroscopy even if the animal is moving during the data acquisition of EPR spectra. The control systems are formulated to allow the behavior of interference between them to be investigated, since they are generally coupled due to the characteristics of the microwave resonator. The stability of the entire control system in a 1.1 GHz CW-EPR spectrometer is evaluated with the generalized Nyquist stability criterion. We compare the EPR spectra of a triarylmethyl (TAM) radical that is dosed in anesthetized mice in terms of the signal-to-noise ratio (SNR) to test the precompensator for decoupling the ATC and AMC systems. The experimental findings suggest that the present technique is useful for improving the SNR of EPR spectra in animal experiments. The SNR of the measured EPR spectra was improved by about 50% with the precompensator.

Algorithms↗
PubMed2004

An in-vivo experimental model for studying wound-healing after laser irradiation in the mouse foetus.

OBJECTIVE: The purpose of this study was to develop an experimental model to study wound-healing in the mouse foetus by inducing an injury with an argon laser. MATERIAL AND METHODS: ICR strain mouse dams were used in this study at day 14 of gestation. Laparotomy was performed on the dams under sodium pentobarbital anaesthesia, and foetuses were exposed from the uterus while wrapped in the amnion. Laser radiation was conducted through the amniotic membrane, and the beam was focused on to the naso-labial region. After laser irradiation, the foetus was returned to the abdominal cavity of the dam. Then the abdominal wall was closed, and an extrauterine pregnancy was maintained. Foetuses were sacrificed at intervals and wound healing was examined histologically. RESULTS: Immediately after laser irradiation, the foetal epithelium was detached and degeneration of the epithelium and subepithelial mesenchymal tissue were observed. Twenty-four hours after laser irradiation, normal epithelial cells surrounding the wound began to migrate along the margin of the degenerate tissue mass. By seventy-two hours after laser irradiation, the laser-induced wound had recovered, and scar formation was not observed. CONCLUSION: The application of an argon laser allowed to inflict a wound on a mouse foetus without damaging the amnion, and this experimental model appeared to be useful for studying the mechanism of foetal wound-healing.

Animals↗
PubMed2004

A new system for in vivo assessment of the degree of nonlinear generation using the second harmonic component in echo signals.

To assess the feasibility of a new system that uses the finite amplitude method and makes possible the assessment of the degree of nonlinear generation in vivo we adopted a method for agar-gel phantoms, using varying lipid concentrations and human livers in vivo. Instead of calculating the nonlinearity parameter B/A value, we evaluated the h value, which is determined by plotting the measured values of the ratio of the amplitude of the second harmonic component to that of the fundamental component. The agar-gel phantoms of various lipid concentrations and 20 cases of normal and 21 cases of fatty liver were studied in this investigation. There was an almost linear agreement between the h value and the lipid concentration of the phantom. The h value (mean +/- SD) was 0.25 +/- 0.03 in normal liver and 0.35 +/- 0.05 in fatty liver. A statistically significant difference was found between them (p < 0.0001). This new system was considered to be usable for evaluating h in vivo as an indicator of the lipid content of the liver.

Fatty Liver↗