Search PubMed⌕ Search

Biomedical subjects

K Sakaguchi

Publications and source records attributed to K Sakaguchi.

At least 163 records · Page 9Linked to original sources
PubMed1997

Preclinical Cushing's syndrome due to adrenocorticotropin-independent bilateral adrenocortical macronodular hyperplasia with concurrent excess of gluco- and mineralocorticoids.

A 48-year-old with bilateral adrenal incidentalomas was studied. Although the serum cortisol level was normal, autonomous cortisol secretion was shown by the loss of diurnal rhythm, no suppressibility by dexamethasone, and complete suppression of plasma adrenocorticotropin levels. Imaging analyses revealed bilateral adrenal masses, showing isotope uptake. He was diagnosed as preclinical Cushing's syndrome due to adrenocorticotropin-independent bilateral adrenocortical macronodular hyperplasia, confirmed histologically. Despite the undetectable plasma adrenocorticotropin level and renin activity, serum mineralocorticoids levels were not suppressed, which were decreased after subtotal adrenalectomy. It was suggested that hyperplastic adrenal glands could produce both gluco- and mineralocorticoids concomitantly.

Adrenal Cortex↗
PubMed1997

Effects of fenthion, isoxathion, dichlorvos and propaphos on the serum cholinesterase isoenzyme patterns of dogs.

The serum cholinesterase (ChE) isoenzyme patterns consist of 5 bands for normal beagle dogs. We examined the ChE isoenzyme patterns and ChE activities of 4 groups, each of which consisted of 4 dogs, given the organophosphate (OP) compounds fenthion (P = S type), isoxathion (P = S type), dichlorvos (P = O type) or propaphos (P = O type) po at 220, 75, 120 or 90 mg/kg respectively. Blood samples were collected for 30 d after the OP administrations. ChE isoenzyme bands 4 and 5 were inhibited 12 h after fenthion and isoxathion administration; dichlorvos and propaphos produced suppression of main bands 4 and 5 20 min after administration. This change was effective to differentiate the type of OP administered (P = S and P = O), while the main ChE isoenzyme bands of dog serum reflected inhibition of serum ChE activity.

Animals↗
PubMed1996

The oligomerization domain of p53: crystal structure of the trigonal form.

The structure of the oligomerization domain of the p53 tumor suppressor protein was determined in the trigonal crystal form, using a refined NMR structure as a model. A synthetic peptide comprising residues 319-360 of human p53 crystallized in the space group P3(1)21. There is one biologically relevant tetrameric domain in the crystallographic asymmetric unit. The structure was refined jointly with NMR data, only the third such case (the previous examples being IL-1beta (Shaanan, B., Gronenborn, A.M., Cohen, G.H., Gilliland, G.L., Veerapandian, B., Davies, D.R. and Clore, G.M. (1992) Science 257, 961-964 [1]) and BPTI (Schiffer, C., Huber, R., Wuthrich, K. and Van Gunsteren, W.F. (1994) J. Mol. Biol. 241, 588-599 [21)), to 2.5 A resolution with an R factor of 0.207. The distribution of tumor-derived mutations in the oligomerization region together with structural and biological data suggest a strategy for the design of antitumor therapeutics.

Biopolymers↗
PubMed1996

Transcriptional regulation of the Drosophila CycA gene by the DNA replication-related element (DRE) and DRE binding factor (DREF).

The Drosophila gene for cyclin A is expressed in dividing cells throughout development. This expression pattern is similar to those of genes related to DNA replication, suggesting involvement of some common control mechanism(s). In the upstream region (-71 to -64 with respect to the transcription initiation site) of the CycA gene, we found a sequence identical to the DNA replication-related element (DRE; 5'-TATCGATA), which is important for high level expression of replication-related genes such as those encoding DNA polymerase alpha and proliferating cell nuclear antigen. Transient expression assays with chloramphenicol acetyltransferase (CAT) were carried out to examine the function of the DRE sequence of the CycA gene. Deletion or base substitution mutations resulted in an extensive reduction in CAT expression. Furthermore, monoclonal antibodies against DRE binding factor (DREF) diminished or supershifted the complex of the DREF and DRE-containing fragment. The results indicate that the Drosophila CycA gene is under the control of a DRE/DREF system, as are DNA replication-related genes.

Animals↗
PubMed1996

The proteolytic environment involved in MHC class II-restricted antigen presentation can be modulated by the p41 form of invariant chain.

During biosynthesis, MHC class II associates with invariant chain which exists in two forms, p31 and p41. Both forms of invariant chain prevent peptide binding to class II, facilitate transport, and enhance class II localization to Ag-processing compartments. In spite of these shared functions, presentation of some Ags can be selectively enhanced by expression of p41. Here we show that p41 can function as a protease inhibitor: 1) the functional and biochemical consequences of p41 expression can be mimicked by inhibiting cysteine proteases in vivo; 2) the amount of intracellular active cysteine proteases is dramatically decreased in p41-positive cells; and 3) a polypeptide corresponding to the p41-unique region is a potent inhibitor of cathepsin L in vitro. These data suggest that p41 can enhance Ag presentation by reducing the proteolytic activity of the Ag-processing compartment, thus protecting a subset of antigenic epitopes from excessive degradation.

Animals↗
PubMed1996

Fatty acids selectively inhibit eukaryotic DNA polymerase activities in vitro.

The in vitro relationship between eukaryotic DNA polymerases and fatty acids was investigated. Some fatty acids strongly inhibited the activities of DNA polymerase alpha and/or beta in vitro. The kinetics of inhibition by linoleic acid showed that DNA polymerase alpha was non-competitively inhibited with respect to the DNA template and substrate (dTTP), while DNA polymerase beta was inhibited competitively with both DNA and substrate.

Animals↗
PubMed1996

FRAG1, a gene that potently activates fibroblast growth factor receptor by C-terminal fusion through chromosomal rearrangement.

A constitutively active form of fibroblast growth factor 2 (FGFR2) was identified in rat osteosarcoma (ROS) cells by an expression cloning strategy. Unlike other tyrosine kinase receptors activated by N-terminal truncation in tumors, this receptor, FGFR2-ROS, contains an altered C terminus generated from chromosomal rearrangement with a novel gene, designated FGFR activating gene 1 (FRAG1). While the removal of the C terminus slightly activates FGFR2, the presence of the FRAG1 sequence drastically stimulates the transforming activity and autophosphorylation of the receptor. FGFR2-ROS is expressed as a unusually large protein and is highly phosphorylated in NIH 3T3 transfectants. FRAG1 is ubiquitously expressed and encodes a predicted protein of 28 kDa lacking significant structural similarity to known proteins. Epitope-tagged FRAG1 protein showed a perinuclear localization by immunofluorescence staining. The highly activated state of FGFR2-ROS appears to be attributed to constitutive dimer formation and higher phosphorylation level as well as possibly altered subcellular localization. These results indicate a unique mechanism of receptor activation by a C terminus alteration through a chromosomal fusion with FRAG1.

3T3 Cells↗
PubMed1996

Autoreactive cytotoxic T lymphocytes in human immunodeficiency virus type 1-infected subjects.

A subtractive analysis of peptides eluted from major histocompatibility complex (MHC) class I human histocompatibility leukocyte antigen (HLA)-A2.1 molecules purified from either human immunodeficiency virus type-1 (HIV-1)-infected or uninfected cells was performed using micro high-performance liquid chromatography and mass spectrometry. Three peptides unique to infected cells were identified and found to derive from a single protein, human vinculin, a structural protein not known to be involved in viral pathogenesis. Molecular and cytofluorometric analyses revealed vinculin mRNA and vinculin protein overexpression in B and T lymphocytes from HIV-1-infected individuals. Vinculin peptide-specific CTL activity was readily elicited from peripheral blood lymphocytes of the majority of HLA-A2.1+, HIV+ patients tested. Our observations suggest that atypical vinculin expression and MHC class I-mediated presentation of vinculin-derived peptides accompany HIV infection of lymphoid cells in vivo, with a resultant induction of antivinculin CTL in a significant portion of HIV+ (HLA-A2.1+) individuals.

Amino Acid Sequence↗
PubMed1996

Characterization of DNA ligase from the fungus Coprinus cinereus.

DNA ligase was highly purified from the fungus Coprinus cinereus at the miotic recombination stage, pachytene. The pachytene DNA ligase showed three polypeptides with molecular masses of 88, 84 and 80 kDa, as estimated by the [32P]AMP-labeling assay. These three polypeptides were susceptible to reaction with an mAb against a 16-amino-acid sequence in human DNA ligase I, which is conserved in C-terminal regions of mammalian, vaccinia virus and yeast DNA ligases. Since rapidly purified preparations from fresh pachytene cells exhibited a single polypeptide of DNA ligase with a molecular mass of 88 kDa, the smaller polypeptides seemed to be limited-degradation products of the 88-kDa polypeptide during the isolation and purification procedures. K(m) values for ATP and (dT)20 hybridized with (dA)n were 1.5 microM and 90 nM, respectively. This enzyme was capable of joining (dT)20.(rA)n and (rA)12-18 (dT)n as well as (dT)20.(dA)n and able to ligate blunt-ended DNA in the presence of poly(ethylene glycol) 6000. DNA ligases were also partially purified from zygotene cells at the meiotic pairing stage and mitotic mycelium cells. In their molecular mass, immuno-reactivity, K(m) value and substrate specificity, they were indistinguishable from pachytene DNA ligase. These results suggest that the fungus C. cinereus at the pachytene stage contains DNA ligase with a molecular mass of 88 kDa as a main or a single species, which is quite similar to DNA ligases from the zygotene and mycelium cells in molecular and catalytic properties.

Amino Acid Sequence↗
PubMed1996

Interaction of Shc with adaptor protein adaptins.

The role of Shc as a substrate of receptors for growth factors and cytokines is well established. To gain further insight into the function of Shc in signal transduction, we used an affinity method to identify potential Shc-binding proteins. Incubation of bovine brain lysates with a glutathione S-transferase (GST)-Shc fusion protein immobilized on glutathione-Sepharose beads resulted in the binding of cellular proteins of approximately 115, 110, and 100 kDa as well as those of 50 and 17 kDa. Amino acid sequencing of tryptic peptides revealed that the 100-kDa protein was almost identical to beta-adaptin and that the 110- and 115-kDa proteins were almost identical to alphaA-adaptin. Using immunoblot analysis, anti-alpha-adaptin antibody recognized several proteins of 100 approximately 115 kDa, and anti-beta-adaptin antibody recognized a 100-kDa protein, suggesting that alphaA-, alphaC-, and beta-adaptins are bound to the GST-Shc fusion protein. Immunoblot analysis with anti-alpha-adaptin antibody revealed that alpha-adaptin was coimmunoprecipitated with Shc from PC12, KB, and COS cell lysates, suggesting a specific interaction of Shc and adaptins in intact cells. A binding study using mutant GST-Shc fusion proteins revealed that the collagen homologous region (amino acids 233-377) of Shc was required for adaptin binding. Conversely, the collagen homologous region of Shc inhibited the binding of adaptins to GST-Shc. In addition, adaptin was able to bind mutant fusion proteins containing amino acids 233-369, 233-355, 346-369, and 346-355 of Shc, but failed to bind a mutant containing amino acids 233-345, suggesting that amino acids 346-355 (RDLFDMKPFE) in the collagen homologous region of Shc are required for adaptin binding. Thus, this study indicates the specific interaction of Shc with alpha- and beta-adaptin components of plasma membrane adaptor proteins that are thought to be involved in receptor endocytosis.

Adaptor Protein Complex 2↗
PubMed1996

Involvement of prostaglandin endoperoxide H synthase-2 in osteoclast-like cell formation induced by interleukin-1 beta.

Interleukin-1 beta (IL-1 beta) stimulates osteoclast-like cell formation via prostaglandin E2 (PGE2) production. However, the regulatory mechanism for the production of PGE2 in bone cells is still unclear. Recently, it has been shown that two prostaglandin endoperoxide H synthase (PGHS) isozymes exist, termed PGSH-1 and PGHS-2. We report here that IL-1 beta induces PGE2 production in bone marrow culture induced by a PGHS-2-dependent mechanism. IL-1 beta stimulated the formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells (MNC) and the production of PGE2 in mouse bone marrow cultures. The dose response curves for the indomethacin inhibition of TRAP-positive MNC formation and PGE2 production were nearly identical. Cycloheximide (CHX) suppressed IL-1 beta-induced PGE2 production, suggesting that the production of PGE2 induced by IL-1 beta required de novo protein synthesis. Northern blot analysis determined that IL-1 beta induced PGHS-2 expression by 30 minutes and mRNA levels were maximal by 1-2 h. Cycloheximide potentiated the accumulation of PGHS-2 mRNA linearly up to 8 h. Dexamethasone, an inhibitor of the induction of PGHS-2, inhibited IL-1 beta-induced PGHS-2 mRNA expression and also suppressed IL-1 beta-stimulated formation of TRAP-positive MNC. Furthermore NS-398, as a selective PGHS-2 inhibitor, completely inhibited IL-1 beta-induced TRAP-positive MNC formation. Moreover, IL-1 beta-induced PGHS-2 mRNA expression and formation of TRAP-positive MNC were inhibited by calphostin C, a selective inhibitor of protein kinase C (PKC). These results indicate that IL-1 beta-induced formation of osteoclast-like cells requires PKC activation, induction of PGHS-2, and subsequent prostaglandin synthesis by this enzyme.

Animals↗
PubMed1996

Elimination of iomeprol by hemodialysis.

Iomeprol is commercially a relatively new type of contrast medium. There have been no reports concerning its dialysability and safety when administered to patients with end-stage renal failure undergoing regular therapeutic hemodialysis (HD). One hundred milliliters of iomeprol (350 mgl/ml) was injected intravenously at computed tomography in 10 patients suffering from chronic renal failure. HD was then performed within 30 min after the start of iomeprol injection. The duration of HD was 4 h and blood samples were taken at 15, 30 min, 1, 2 and 4 h after its start. Mean dialysance of urea was 176.0-182.9 ml/min, compared to 131.4-133.3 ml/min for iomeprol. Elimination rate of iomeprol was 821.4 +/- 4.6% 4 h after the start of HD. Two of the 10 patients had side effects. One of them complained of a heat sensation and the other of sneezing and rash. As 4-h HD effectively removed iomeprol from the body and there were no severe side effects, it is concluded that iomeprol is suitable for HD patients.

Aged↗
PubMed1996

Chemical synthesis of phosphorylated peptides of the carboxy-terminal domain of human p53 by a segment condensation method.

A segment condensation method was developed for the chemical synthesis of large (> 90 amino acid) phosphopeptides and was used to produce phosphorylated and non-phosphorylated derivatives of the C-terminal tetramerization and regulatory domains of human p53 (residues 303-393). Efficient condensation synthesis of the 91 residue p53 domain was achieved in two steps. The non-phosphorylated N-terminal segment p53(303-334) (1) and its derivative phosphorylated at serine 315 (1P315), and the non-phosphorylated middle segment p53(335-360) (2), were synthesized as partially protected peptide thioesters in the solid phase using Boc chemistry. The C-terminal segment p53(361-393) (3) and its derivative phosphorylated at serine 392 (3P392) were synthesized as partially protected peptides in the solid phase using Fmoc chemistry. Phosphoamino acid was incorporated into the N-terminal segment (1P315) at the residue corresponding to p53 serine 315 as Boc-Ser(PO3(Bzl)2)-OH during synthesis. Serine 392 in the C-terminal segment was selectively phosphorylated after synthesis by phosphitylation followed by oxidation. A derivative phosphorylated at serine 378 was synthesized in a one-step condensation of the unphosphorylated N-terminal segment (1) and the phosphorylated long C-terminal segment p53(335-393) (2-3P378). Yields of the ligated peptides after removal of the protecting groups and HPLC purification averaged 60% for the first condensation and 35% for the second condensation. All five p53 peptides exhibited monomer-tetramer association as determined by analytical ultracentrifugation. Circular dichroism spectroscopy revealed that phosphorylation at Ser315 increased the alpha-helical content, which was abolished when Ser392 also was phosphorylated, suggesting an interaction between N-terminal and C-terminal residues of the C-terminal domain of p53.

Amino Acid Sequence↗
PubMed1996

Phosphaturic mesenchymal tumor, mixed connective tissue variant (oncogenic osteomalacia).

A case of tumor-induced phosphaturic osteomalacia in a 54 year old man is reported. The patient was admitted because of progressive muscle spasms with pain and weakness in the bilateral thighs. Laboratory data showed hypophosphatemia, decreased tubular resorption of phosphate (TRP), a low 1,25-dihydroxyvitamin D level, and a high serum alkaline phosphatase level. Radiologic examinations revealed multiple lesions of osteomalacia in the ribs, and a small mass in the lower posterior mediastinum. After removal of the tumor, clinical symptoms disappeared and hypophosphatemia, decreased TRP, and the 1,25-dihydroxyvitamin D level were corrected. Microscopical examination revealed that the tumor was composed of mature adipose tissues, osseous tissues, and primitive stromal zones including osteoclast-like giant cells, non-mineralized woven bone, and various sized blood vessels. Patho-physiologic observations suggested that the tumor secreted some humoral substances inhibiting 25-hydroxyvitamin D-1 alpha-hydroxylase activity, renal phosphate resorption, and parathyroid hormone production.

Alkaline Phosphatase↗