Biomedical subjects

Y Gotoh

Publications and source records attributed to Y Gotoh.

At least 19 recordsLinked to original sources

PubMed2006

Transient encephalopathy with reversible white matter lesions in children.

We report on two children with acute encephalopathy showing mild clinical manifestations and reversible white matter lesions. In both patients, MRI revealed high intensities on T (2)-weighted imaging and marked reductions of water diffusion in the white matter of the bilateral centrum semiovale and the corpus callosum. These abnormalities disappeared along with the neurological symptoms within a week in both patients. These children represent a characteristic group of patients among childhood acute encephalopathy.

Acute Disease↗

PubMed2004

Ionization and solvation of HCl adsorbed on the D2O-ice surface.

The interaction of HCl with the D(2)O-ice surface has been investigated in the temperature range 15-200 K by utilizing time-of-flight secondary ion mass spectroscopy, temperature-programmed desorption, and x-ray photoelectron spectroscopy. The intensities of sputtered H(+)(D(2)O) and Cl(-) ions (the H(+) ions) are increased (decreased) markedly above 40 K due to the hydrogen bond formation between the HCl and D(2)O molecules. The HCl molecules which form ionic hydrates undergo H/D exchange at 110-140 K and a considerable fraction of them dissolves into the bulk above 140 K. The neutral hydrates of HCl should coexist as evidenced by the desorption of HCl above 170 K. They are incorporated completely in the D(2)O layer up to 140 K. The HCl molecules embedded in the thick D(2)O layer dissolve into the bulk, and the ionic hydrate tends to segregate to the surface above 150 K.

Journal Article↗

PubMed2004

Oxidative stress induces insulin resistance by activating the nuclear factor-kappa B pathway and disrupting normal subcellular distribution of phosphatidylinositol 3-kinase.

AIMS/HYPOTHESIS: Oxidative stress is associated with diabetes, hypertension and atherosclerosis. Insulin resistance is implicated in the development of these disorders. We tested the hypothesis that oxidative stress induces insulin resistance in rats, and endeavoured to identify mechanisms linking the two. METHODS: Buthionine sulfoximine (BSO), an inhibitor of glutathione synthase, was administered to Sprague-Dawley rats and 3T3-L1 adipocytes. Glucose metabolism and insulin signalling both in vivo and in 3T3-L1 adipocytes were examined. In 3T3-L1 adipocytes, the effects of overexpression of a dominant negative mutant of inhibitory kappa B (I kappa B), one role of which is to block oxidative-stress-induced nuclear factor (NF)-kappa B activation, were investigated. RESULTS: In rats given BSO for 2 weeks, the plasma lipid hydroperoxide level doubled, indicating increased oxidative stress. A hyperinsulinaemic-euglycaemic clamp study and a glucose transport assay using isolated muscle and adipocytes revealed insulin resistance in BSO-treated rats. BSO treatment also impaired insulin-induced glucose uptake and GLUT4 translocation in 3T3-L1 adipocytes. In BSO-treated rat muscle, adipose tissue and 3T3-L1 adipocytes, insulin-induced IRS-1 phosphorylation in the low-density microsome (LDM) fraction was specifically decreased, while that in whole cell lysates was not altered, and subsequent translocation of phosphatidylinositol (PI) 3-kinase from the cytosol and the LDM fraction was disrupted. BSO-induced impairments of insulin action and insulin signalling were reversed by overexpressing the dominant negative mutant of I kappa B, thereby suppressing NF-kappa B activation. CONCLUSIONS/INTERPRETATION: Oxidative stress induces insulin resistance by impairing IRS-1 phosphorylation and PI 3-kinase activation in the LDM fraction, and NF-kappa B activation is likely to be involved in this process.

3T3 Cells↗

PubMed2004

Interactions of D2O with methane and fluoromethane surfaces.

TOF-SIMS is used to investigate the interactions between D2O and hydrophobic molecules, such as CH4, CH3F, CH2F2, CHF3, and CF4, at cryogenic temperatures (15 K). By irradiation with a 1.5-keV He+ beam, the D(+)(D2O)n ions are ejected efficiently from the D2O nanoclusters physisorbed on the CF4 layer due to Coulomb explosion: the ion yields are by about two orders of magnitude higher than those from a thick D2O layer via the kinetic sputtering. The D(+)(D2O)n yields decrease on the CHnF(4-n) layer with increasing the number of the C-H group. This is because the Coulombic fission is quenched due to the delocalization of valence holes through the C-H...H-C and C-H...D2O contacts. A pure D2O film is hardly grown on the CH4 layer as a consequence of intermixing whereas the D2O molecules basically adsorb on the surfaces of fluoromethanes, suggesting the attractive (water-repellent) interactions in the C-H...D2O (C-F...D2O) contacts. The C-H...O bond behaves like a conventional O-H...O hydrogen bond as far as the collision-induced proton transfer reaction is concerned.

Journal Article↗

PubMed2004

A human dual-color enzyme-linked immunospot assay for simultaneous detection of interleukin 2- and interleukin 4-secreting cells.

The enzyme-linked immunospot (ELISPOT) assay is an efficient technique for the enumeration of single cells secreting antibodies and cytokines. For simultaneous differentiation of individual cells producing interleukin-2 (IL-2) and interleukin-4 (IL-4) at a single cell level in human peripheral blood mononuclear cells (PBMCs), a human dual-color ELISPOT assay has been optimized. In the present system, the red spots corresponding to IL-2-secreting cells (T helper type 1, Th1, cells) were developed with horseradish peroxidase and the amino ethyl carbazole (AEC)/H2O2. The blue spots corresponding to IL-4-secreting cells (T helper type 2, Th2, cells) were developed with an alkaline phosphatase and the Vector blue. The usefulness of the assay method was tested. With this system, we could detect the IL-2- and IL-4-secreting cells simultaneously in human PBMCs of a juvenile rheumatoid arthritis (JRA) patient. This procedure provides useful information on clinical immune disorders.

Adult↗

PubMed2004

Physical and functional interaction between BH3-only protein Hrk and mitochondrial pore-forming protein p32.

Bcl-2 homology domain (BH) 3-only proteins of the proapoptotic Bcl-2 subfamily play a key role as initiators of mitochondria-dependent apoptosis. To date, at least 10 mammalian BH3-only proteins have been identified, and it is now being realized that they have different roles and mechanisms of regulation in the transduction of apoptotic signals to mitochondria. Hrk/DP5 is one of the mammalian BH3-only proteins implicated in a variety of physiological and pathological apoptosis, yet the molecular mechanism involved in Hrk-mediated apoptosis remains poorly understood. In an attempt to identify cellular proteins participating in Hrk-mediated apoptosis, we have conducted yeast two-hybrid screening for Hrk-interacting proteins and isolated p32, a mitochondrial protein that has been shown to form a channel consisting of its homotrimer. In vitro binding, co-immunoprecipitation, as well as immunocytochemical analyses verified specific interaction and colocalization of Hrk and p32, both of which depended on the presence of the highly conserved C-terminal region of p32. Importantly, Hrk-induced apoptosis was suppressed by the expression of p32 mutants lacking the N-terminal mitochondrial signal sequence (p32(74-282)) and the conserved C-terminal region (p32 (1-221)), which are expected to inhibit binding of Hrk competitively to the endogenous p32 protein and to disrupt the channel function of p32, respectively. Furthermore, small interfering RNA-mediated knockdown of p32 conferred protection against Hrk-induced apoptosis. Altogether, these results suggest that p32 may be a key molecule that links Hrk to mitochondria and is critically involved in the regulation of Hrk-mediated apoptosis.

Amino Acid Sequence↗

PubMed2001

Akt mediates Rac/Cdc42-regulated cell motility in growth factor-stimulated cells and in invasive PTEN knockout cells.

Growth factors promote cell survival and cell motility, presumably through the activation of Akt and the Rac and Cdc42 GTPases, respectively. Because Akt is dispensable for Rac/Cdc42 regulation of actin reorganization, it has been assumed that Rac and Cdc42 stimulate cell motility independent of Akt in mammalian cells. However, in this study we demonstrate that Akt is essential for Rac/Cdc42-regulated cell motility in mammalian fibroblasts. A dominant-negative Akt inhibits cell motility stimulated by Rac/Cdc42 or by PDGF treatment, without affecting ruffling membrane-type actin reorganization. We have confirmed a previous report that Akt is activated by expression of Rac and Cdc42 and also observed colocalization of endogenous phosphorylated Akt with Rac and Cdc42 at the leading edge of fibroblasts. Importantly, expression of active Akt but not the closely related kinase SGK is sufficient for increasing cell motility. This effect of Akt is cell autonomous and not mediated by inhibition of GSK3. Finally, we found that dominant-negative Akt but not SGK reverses the increased cell motility phenotype of fibroblasts lacking the PTEN tumor suppressor gene. Taken together, these results suggest that Akt promotes cell motility downstream of Rac/Cdc42 in growth factor-stimulated cells and in invasive PTEN-deficient cells.

Animals↗

PubMed2001

Beta-amyloid induces neuronal apoptosis via a mechanism that involves the c-Jun N-terminal kinase pathway and the induction of Fas ligand.

Elevated levels of beta-Amyloid (Abeta) are present in the brains of individuals with either the sporadic or familial form of Alzheimer's disease (AD), and the deposition of Abeta within the senile plaques that are a hallmark of AD is thought to be a primary cause of the cognitive dysfunction that occurs in AD. Recent evidence suggests that Abeta induces neuronal apoptosis in the brain and in primary neuronal cultures, and that this Abeta-induced neuronal death may be responsible in part for the cognitive decline found in AD patients. In this study we have characterized one mechanism by which Abeta induces neuronal death. We found that in cortical neurons exposed to Abeta, activated c-Jun N-terminal kinase (JNK) is required for the phosphorylation and activation of the c-Jun transcription factor, which in turn stimulates the transcription of several key target genes, including the death inducer Fas ligand. The binding of Fas ligand to its receptor Fas then induces a cascade of events that lead to caspase activation and ultimately cell death. By analyzing the effects of mutations in each of the components of the JNK-c-Jun-Fas ligand-Fas pathway, we demonstrate that this pathway plays a critical role in mediating Abeta-induced death of cultured neurons. These findings raise the possibility that the JNK pathway may also contribute to Abeta-dependent death in AD patients.

Alzheimer Disease↗

PubMed2001

Caspase cleavage of MST1 promotes nuclear translocation and chromatin condensation.

MST1, mammalian STE20-like kinase 1, is a serine/threonine kinase that is cleaved and activated by caspases during apoptosis. MST1 is capable of inducing apoptotic morphological changes such as chromatin condensation upon overexpression. In this study, we show that MST1 contains two functional nuclear export signals (NESs) in the C-terminal domain, which is released from the N-terminal kinase domain upon caspase-mediated cleavage. Full-length MST1 is excluded from the nucleus and localized to the cytoplasm. However, either truncation of the C-terminal domain, point mutation of the two putative NESs, or treatment with leptomycin B, an inhibitor of the NES receptor, results in nuclear localization of MST1. Staurosporine treatment induces chromatin condensation, MST1 cleavage, and nuclear translocation. Staurosporine-induced chromatin condensation is partially inhibited by expressing a kinase-negative mutant of MST1, suggesting an important role of MST1 in this process. Significantly, MST1 is more efficient at inducing chromatin condensation when it is constitutively localized to the nucleus by mutation of its NESs. Moreover, inhibition of MST1 nuclear translocation by mutation of its cleavage sites reduces its ability to induce chromatin condensation. Taken together, these results suggest that truncation of the C-terminal domain of MST1 by caspases may result in translocation of MST1 into the nucleus, where it promotes chromatin condensation.

Active Transport, Cell Nucleus↗

PubMed2001

Calbindin-D28k and calretinin immunoreactive neurons in the olfactory bulb of the musk shrew, Suncus murinus.

The distribution, morphological features, and postnatal development of calbindin-D28k (CB) and calretinin (CR) immunoreactive neurons in the main olfactory bulb (MOB) of the musk shrew, Suncus murinus, were studied by immunostaining to determine the degree of colocalization of CB and CR, and the relationship of CB and CR to neuron development in the MOB of animals of the order Insectivora. In adults, CB-positive neurons were identified as periglomerular and perinidal cells in the periglomerular region, as superficial short-axon cells in the external plexiform layer, and as four types of interneurons (Cajal, horizontal, Golgi, and bitufted cells) in the mitral cell, internal plexiform, and granule cell layers. CR-positive neurons were identified as projection neurons (tufted and mitral cells) and interneurons (periglomerular, perinidal, and granule cells). On postnatal days 1 and 3, CB-positive neurons revealed numerous processes finely arborized near the somata, and were morphologically unidentifiable. At the same time, CR-positive neurons were identified as young periglomerular and granule cells, and as migrating bipolar cells extending leading processes with growth cones in each layer of the MOB and the subependymal layer between the anterior lateral ventricle and the center of the MOB. On postnatal day 28, mature CB-positive and CR-positive interneurons were distributed in their corresponding layers, whereas migrating CR-positive bipolar cells were rarely detected. No cells colocalized CB and CR. The results suggest that perinidal cells in the shrew MOB may develop postnatally, together with glomerular and granule cells. We suggest that CB is associated with mechanisms of the outgrowth of neuronal processes, whereas CR is involved in mechanisms of cell migration and outgrowth of neuronal processes, in some types of neurons in the developing stage of the shrew MOB.

Age Factors↗

PubMed2001

Akt inhibits the orphan nuclear receptor Nur77 and T-cell apoptosis.

Akt is a common mediator of cell survival in a variety of circumstances. Although some candidate Akt targets have been described, the function of Akt is not fully understood, particularly because of the cell type- and context-dependent apoptosis regulation. In this study, we demonstrate that one of the mechanisms by which Akt antagonizes apoptosis involves the inhibition of Nur77, a transcription factor implicated in T-cell receptor-mediated apoptosis. It has been suggested that Akt phosphorylates Nur77 directly, but whether Akt suppresses biological functions of Nur77 remains unknown. We found that Akt inhibited the DNA binding activity of Nur77 and stimulated its association with 14-3-3 in a phosphorylation site-dependent manner. Moreover, we found that expression of Akt suppressed Nur77-induced apoptosis in fibroblasts and activation-induced cell death of T-cell hybridomas. The inhibition of Nur77 by Akt suggests a mechanism that explains how T-cell receptor activation can promote survival in some instances even when Nur77 is induced. Collectively, these results may suggest that Akt is a negative regulator of Nur77 in T-cell apoptosis.

14-3-3 Proteins↗

PubMed2001

MKK6/3 and p38 MAPK pathway activation is not necessary for insulin-induced glucose uptake but regulates glucose transporter expression.

p38 mitogen-activated protein kinase (MAPK), which is situated downstream of MAPK kinase (MKK) 6 and MKK3, is activated by mitogenic or stress-inducing stimuli, as well as by insulin. To clarify the role of the MKK6/3-p38 MAPK pathway in the regulation of glucose transport, dominant negative p38 MAPK and MKK6 mutants and constitutively active MKK6 and MKK3 mutants were overexpressed in 3T3-L1 adipocytes and L6 myotubes using an adenovirus-mediated transfection procedure. Constitutively active MKK6/3 mutants up-regulated GLUT1 expression and down-regulated GLUT4 expression, thereby significantly increasing basal glucose transport but diminishing transport induced by insulin. Similar effects were elicited by chronic (24 h) exposure to tumor necrosis factor alpha, interleukin-1beta, or 200 mm sorbitol, all activate the MKK6/3-p38 MAPK pathway. SB203580, a specific p38 MAPK inhibitor, attenuated these effects, further confirming that both MMK6 and MMK3 act via p38 MAPK, whereas they had no effect on the increase in glucose transport induced by a constitutively active MAPK kinase 1 (MEK1) mutant or by myristoylated Akt. In addition, suppression of p38 MAPK activation by overexpression of a dominant negative p38 MAPK or MKK6 mutant did not diminish insulin-induced glucose uptake by 3T3-L1 adipocytes. It is thus apparent that activation of p38 MAPK is not essential for insulin-induced increases in glucose uptake. Rather, p38 MAPK activation leads to a marked down-regulation of insulin-induced glucose uptake via GLUT4, which may underlie cellular stress-induced insulin resistance caused by tumor necrosis factor alpha and other factors.

3T3 Cells↗

PubMed2001

Both phosphorylation and caspase-mediated cleavage contribute to regulation of the Ste20-like protein kinase Mst1 during CD95/Fas-induced apoptosis.

The serine/threonine kinase Mst1, a mammalian homolog of the budding yeast Ste20 kinase, is cleaved by caspase-mediated proteolysis in response to apoptotic stimuli such as ligation of CD95/Fas or treatment with staurosporine. Furthermore, overexpression of Mst1 induces morphological changes characteristic of apoptosis in human B lymphoma cells. Mst1 may therefore represent an important target for caspases during cell death which serves to amplify the apoptotic response. Here we report that Mst1 has two caspase cleavage sites, and we present evidence indicating that cleavage may occur in an ordered fashion and be mediated by distinct caspases. We also show that caspase-mediated cleavage alone is insufficient to activate Mst1, suggesting that full activation of Mst1 during apoptosis requires both phosphorylation and proteolysis. Another role of phosphorylation may be to influence the susceptibility of Mst1 to proteolysis. Autophosphorylation of Mst1 on a serine residue close to one of the caspase sites inhibited caspase-mediated cleavage in vitro. Finally, Mst1 appears to function upstream of the protein kinase MEKK1 in the SAPK pathway. In conclusion, Mst1 activity is regulated by both phosphorylation and proteolysis, suggesting that protein kinase and caspase pathways work in concert to regulate cell death.

Apoptosis↗

PubMed2001

Looping technique for transpapillary selective biopsy of the left hepatic duct.

PURPOSE: Because biopsy forceps tend to turn towards the right hepatic duct during endoscopic retrograde cholangiopancreatography (ERCP), selective access to the left hepatic duct is difficult. METHODS: In this study, we managed to insert biopsy forceps selectively into the left hepatic duct, by using a looping technique, in three patients. Biopsy forceps were inserted into the right hepatic duct by the conventional method. The elevator of the endoscope was kept down, and the shaft of the biopsy forceps was then advanced to the duodenal cavity until it formed a loop between the endoscope and the papilla. During the procedure, the tip of the forceps was kept at the hepatic hilus. RESULTS: In this condition, we were able to slowly rotate the tip of the forceps and direct the forceps towards the left. Sufficient material from the left hepatic duct was obtained in all patients. CONCLUSIONS: The looping technique was useful for selective access to the left hepatic duct.

Adult↗

PubMed2001

Cine viewing of abdominal CT.

A few studies have been reported that CT cine viewing on the CRT is superior to film-based viewing of CT images (Seltzer et al., Radiology 197 (1994) 119; Bonaldi et al., Am. J. Roentgenol. 170 (1998) 373; Tillich et al., Am. J. Roentgenol. 169 (1997) 1611). The purpose of our study is to know how to use cine viewing of abdominal CT. Thirty CT studies on the abdomen with both precontrast and postcontrast images were examined. The suitable rate of cine viewing ranged from 1 to 6 frames per second according to the size, the contrast and the complexity of the anatomical structures, and the slice thickness. For small or complex structures, checking each image might be required to know the full detail of them. Positional sorting among multiphase images, which is followed by consecutive display of a precontrast image, postcontrast early and late phase images at one position and so on, is useful to see the dynamic pattern of enhancement of the anatomical structures. However, there was no significant difference between cine viewing and film-based viewing concerning both the detectability of the anatomical structures and the conspicuity of enhancement of the liver and the pancreas, so that cine viewing might be an alternative to film-based viewing for CT diagnosis of the abdomen.

Abdomen↗

PubMed2001

Relationships among the physical parameters required to give a linear relation between slope and intercept of Fowler-Nordheim plots.

For field emitters with a given surface condition, but different apex radii, there exists an empirical linear relationship between the slopes and intercepts of their Fowler-Nordheim plots. This behavior is quite different from that predicted by the theory with the assumption of a smooth surface. We have already characterized the possible reasons for this discrepancy; one possibility is that the emission area rapidly increases with increase in the apex radius. The previous report (Jpn. J. Appl. Phys. Part II 32 (1993) L342), however, showed a curved relation between the slope and the intercept of Fowler-Nordheim plots. In the present article, we try to extract the relationship between the emission area and the apex radius needed in order to give a linear relationship between slope and intercept of the F-N plots. As an additional result of the present analysis, the change in work function of the field emitting surface is demonstrated.

Journal Article↗

PubMed2001

On the increase of the intensity ratio of doubly charged ions to singly charged ions for liquid gold and copper ion sources.

The reason for the increase of R21 which is defined by the intensity ratio of the doubly charged ions to the singly charged ions, was studied. Based on the conventional field evaporation theory, an increase in the electric field and/or the source temperature is considered to be attributable. We took the fact that R21 turned to decrease at the higher current regime into consideration and examined whether the change of R21 due to change in the field or temperature would finally meet the criterion for the decrease of R21. It was concluded that an increase of the source temperature may be a possible reason for the increase of R21.

Journal Article↗

PubMed2001

MST1-JNK promotes apoptosis via caspase-dependent and independent pathways.

BACKGROUND: MST1 is an upstream kinase of the JNK and p38 MAPK pathways whose expression induces apoptotic morphological changes such as nuclear condensation. During apoptosis, caspase cleavage of MST1 removes a C-terminal regulatory domain, increasing the kinase activity of the MST1 N-terminal domain. Downstream pathways of MST1 in the induction of apoptosis remain to be clarified. RESULTS: In this study, we found that the expression of MST1 resulted in caspase-3 activation. Therefore, MST1 is not only a target of caspases but also an activator of caspases. This caspase activation and apoptotic changes occur through JNK, since the co-expression of a dominant-negative mutant of JNK inhibited MST1-induced morphological changes as well as caspase activation. In contrast, neither a dominant-negative p38 nor the p38 inhibitor SB203580 inhibited them. MST1 induced nucleosomal DNA fragmentation, which was suppressed by caspase inhibitors or ICAD (Inhibitor of Caspase-Activated DNase). Surprisingly, however, other changes such as membrane blebbing and chromatin condensation were not inhibited by caspase inhibitors. CONCLUSION: These results suggest that MST1 most likely promotes two events through JNK activation; first, MST1 induces the activation of caspases, resulting in CAD-mediated DNA fragmentation, and second, MST1 induces chromatin condensation and membrane blebbing without utilizing downstream caspases.

Animals↗