Biomedical subjects

T Takao

Publications and source records attributed to T Takao.

At least 127 records · Page 7Linked to original sources

PubMed1995

Use of the derivatizing agent 4-aminobenzoic acid 2-(diethylamino)ethyl ester for high-sensitivity detection of oligosaccharides by electrospray ionization mass spectrometry.

A method for the high-sensitivity detection of oligosaccharides by electrospray ionization mass spectrometry (ESI-MS) is reported. The method involves the chemical derivatization with 4-aminobenzoic acid 2-(diethylamino)-ethyl ester (ABDEAE). This derivative, which contains a 2-(diethylamino)ethyl group, having a high proton affinity, enhances the ionization efficiency of analytes in the positive ESI mode. Experiments using maltohexaose as a model oligosaccharide revealed that derivatization with ABDEAE gave a remarkably large increase in molecular ion abundance. Using a mixture of acetonitrile, 2-methoxyethanol, 2-propanol, and water (1:1:1:1 v/v/v/v) as solvent for ESI, ABDEAE-derivatized maltohexaose could be detected at a level of 10 fmol. This represents a 5000-fold improvement in sensitivity over underivatized maltohexaose. ESI tandem mass spectrometry of the ABDEAE-derivatized maltohexaose provides structural information at the low-picomole level. In this spectrum, 1,5X' and 0,2A" series of sequence ions, arising from ring cleavage, were observed as the predominant ions.

Anions↗

PubMed1995

Corticotropin-releasing factor treatment upregulates interleukin-1 receptors in the mouse pituitary: reversal by dexamethasone.

Intraperitoneal injection of rat/human corticotropin-releasing factor (CRF) (40 micrograms/kg/0.2 ml of saline) resulted in a dramatic increase in specific iodine-125-labeled human interleukin-1 alpha ([125I]IL-1 alpha) binding in the male C57BL/6 mouse pituitary at 2 and 6 h after the injection although it did not affect [125I]IL-1 alpha binding in the mouse hippocampus, spleen and testis at any time after the injection. [125I]IL-1 alpha binding was unchanged at 2 h following dexamethasone (DEX) treatment (1 mg/kg/0.2 ml of 4% ethanol-saline) in the mouse pituitary and the hippocampus. In contrast, DEX inhibited CRF-induced upregulation of IL-1 receptors in the pituitary at 2 h after the injection. These data demonstrate complex interactions between CRF and DEX on IL-1 receptors during stress.

Adrenocorticotropic Hormone↗

PubMed1995

In vivo evidence for involvement of a 58 kDa component of nuclear pore-targeting complex in nuclear protein import.

We recently showed that a nuclear location signal (NLS)-containing karyophile forms a stable complex with cytoplasmic components for nuclear pore-targeting The complex, termed nuclear pore-targeting complex (PTAC), contained two essential proteins of 54 and 90 kDa, respectively, as estimated by electrophoresis. In this study, we found that the 54 kDa component of PTAC is the mouse homologue of Xenopus importin (m-importin). Cytoplasmic injection of the antibodies raised against recombinant m-importin showed an inhibitory effect on nuclear import of a karyophile in living mammalian cells. A portion of cytoplasmically injected antibodies migrated rapidly into the nucleus, indicating dynamic movement of this protein across the nuclear envelope. Moreover, the injected antibodies co-precipitated the karyophile, in an NLS-dependent manner, with endogenous m-importin in the cytoplasm. These results provide in vivo evidence that m-importin is involved in nuclear protein import through association with a NLS in the cytoplasm before nuclear pore binding.

Amino Acid Sequence↗

PubMed1995

The nuclear pore-targeting complex binds to nuclear pores after association with a karyophile.

We recently showed that a karyophilic protein forms a stable complex, termed nuclear pore-targeting complex (PTAC), with cytoplasmic components prior to nuclear pore-binding. In this study, we cloned a cDNA encoding a 97 kDa of PTAC (PTAC97). Recombinant PTAC97 completely reconstitutes the nuclear binding-step in conjunction with a 58 kDa component of PTAC (PTAC58) in the semi-intact cell-free transport assay. Biochemical analysis reveals that PTAC58 binds to a karyophilic protein, and PTAC97 is associated with PTAC58 in a 1:1 molar ratio. A complex of PTAC97 and PTAC58 targets nuclear pores, depending on the presence of a karyophile. These in vitro results suggest that the first step in nuclear import occurs through the targeting-complex formation of a karyophile with PTAC58 bound to PTAC97.

Amino Acid Sequence↗

PubMed1995

Role of heterogeneous N-terminal acylation of recoverin in rhodopsin phosphorylation.

Recoverin, a new member of the EF-hand superfamily, plays a critical role in the light/dark adaptation of retinal rods by regulating rhodopsin phosphorylation in a Ca(2+)-dependent manner. Recoverin is composed of four isoforms, each of which is modified at its N terminus by myristate (C14:0) or its structurally related fatty acid (C12:0, C14:2, or C14:1). Although the N-fatty acylation is implicated in protein-membrane and protein-protein interactions, the functional difference among the recoverin isoforms and the significance of the heterogeneous acylation have not been defined. Here we separated the heterogeneous recoverin into three fractions, C14:0-recoverin, C14:1-recoverin, and a mixture of C14:2- and C12:0- (C14:2/C12:0-) recoverin to evaluate the individual properties. Recoverin in every fraction bound Ca2+ as assessed by fluorescence spectroscopy and inhibited the light-dependent rhodopsin phosphorylation in the same range of free Ca2+ concentration (0.3-0.8 microM). However, the magnitude of the inhibition at higher Ca2+ concentration was different among the isoforms and ranked in the same order of the hydrophobicity of the N-fatty acyl groups: C14:0 > C14:1 > C14:2/C12:0. These results indicate that the diverged hydrophobicity of the recoverin N terminus plays an important role in the interaction with the membranes and/or its target protein but not with Ca2+.

Acylation↗

PubMed1995

O-glycosylation of the Thr70 residue of cell-adhesive lysozyme in yeast.

The cell-adhesive protein Cys-RGD4 has been constructed using a yeast expression system by inserting the sequence Cys-Arg-Gly-Asp-Ser-Cys (CRGDSC) between Val74 and Asn75 of human lysozyme [Yamada, T., Uyeda, A., Kidera, A. & Kikuchi, M. (1994b) Biochemistry 33, 11678-11683]. The Cys74a, Arg74b, Gly74c, Asp74d, Ser74e, Cys74f-lysozyme mutant, purified from the yeast culture supernatant contained glycosylated variants, in addition to the unglycosylated form. Peptide mapping analyses suggested that the glycosylation occurred at the Thr70 residue in the Cys-RGD4 molecule. Electrospray ionization mass spectrometric analysis demonstrated the presence of two hexose residues in the major variant, and one, three, four, or five hexose residues in the minor variants. All of these hexose residues were identified as mannose by analysis of the oligosaccharide mixture obtained by mild alkaline treatment of the variants. No other glycosylation was observed, although the Cys-RGD4 molecule possesses a total of 12 threonine and serine residues. In addition, the Thr70 residue is not glycosylated in either native lysozyme or the Arg-Gly-Asp-Ser (RGDS)-inserted mutant, RGD4 [Yamada, T., Matsushima, M., Inaka, K., Ohkubo, T., Uyeda, A., Maeda, T., Titani, K., Sekiguchi, K. & Kikuchi, M. (1993) J. Biol. Chem. 268, 10588-10592]. Thus, this O-glycosylation seems to be specific for both the mutant lysozyme molecule and the site of the threonine residue. Structural analyses of these lysozymes by X-ray crystallography suggest that the conformation of the serine-containing or threonine-containing region can affect the specificity of yeast O-glycosylation.

Amino Acid Sequence↗

PubMed1995

Characterization of a highly toxic, large molecular size heat-stable enterotoxin produced by a clinical isolate of Yersinia enterocolitica.

A novel heat-stable enterotoxin (ST) designated as Y-STc was purified to homogeneity from the culture supernatant of a pathogenic strain of Yersinia enterocolitica serotype O3 and its amino acid sequence was determined. The mature Y-STc was found to consist of 53 amino acid residues, which includes the putative pro-sequence. The molecular weight of Y-STc was 5638 and constituted the largest molecular size in the family of currently known STs. The minimum effective dose of purified Y-STc in the suckling mouse assay was 0.6 ng (0.0 pmol), indicating that, despite the long sequence, Y-STc is the most toxic in the ST family.

Amino Acid Sequence↗

PubMed1995

Primary structure of a potent endogenous dopa-containing inhibitor of phenol oxidase from Musca domestica.

The complete amino acid sequence of a low molecular weight peptide from the hemolymph of the housefly Musca domestica L., which had been determined to competitively inhibit phenol oxidase (PO; monophenol, dihydroxy-phenylalanine:oxygen oxidoreductase; EC 1.14.18.1) in the nM range, was unambiguously established by employing both automatic Edman degradation and mass spectrometry. The physiologically active peptide, which was designated phenol oxidase inhibitor (POI), has an observed molecular weight of 4213.1 +/- 0.2 by electrospray ionization mass spectrometry. The relatively short and structurally dense peptide contained 38 amino acid residues rich in cysteine and lysine. Comparison of the observed and calculated molecular mass indicates that apparently all six cysteine residues form disulfide bridges. Interestingly, sequence analyses of both the intact and protease-digested S-pyridylethylated POI showed that one of the two tyrosine residues (Tyr-32) is hydroxylated to a 3,4-dihydroxyphenylalanine (dopa) residue. This agreed with the increase of 16 mass units observed in mass spectrometric measurements. This was further verified by submission of free L-dopa to the sequencer, which gave a retention time consistent with the atypical peak observed at the Edman cycle of the peptide containing dopa. This study demonstrates the existence of a biologically active, dopa-containing peptide among the insects. Since the POI activity was most prominent in aged pupae, especially pharate adults, the POI may play an important role in smoothing the way of adult emergence through hindering excessive melanization, as well as hardening, of cuticular proteins under the epicuticle.

Amino Acid Sequence↗

PubMed1995

Modulation of interleukin-1 receptors in the brain-endocrine-immune axis by stress and infection.

We summarize data from some of our recent studies on in vitro and in vivo modulation of interleukin-1 (IL-1) receptors in the mouse brain-endocrine-immune axis by stress and infection. Ether-laparotomy stress in mice resulted in a selective increase in pituitary IL-1 receptors and a significant decrease in pituitary receptors for corticotropin-releasing factor (CRF), a major regulator of the endocrine response to stress. Intraperitoneal injection of rat/human CRF mimicked the effects of stress and resulted in a dramatic increase in [125I]IL-1alpha binding in the pituitary; [125I]IL-1alpha binding in the hippocampus, spleen, and testis was unaffected by stress or CRF treatment. Glucocorticoid treatment with dexamethasone alone did not alter [125I]IL-1alpha binding but significantly inhibited CRF-induced upregulation of IL-1 receptors in the pituitary. The intracellular mechanism(s) involved in stress and CRF-induced upregulation of IL-1 receptors in the pituitary gland were examined by evaluating the effects of treatment of AtT-20 mouse pituitary corticotroph cells with a variety of neuroendocrine mediators of stress. CRF, forskolin, and isoproterenol (beta2 adrenergic receptor agonist) produced dose-dependent increases in cAMP production and [125I]IL-1alpha binding. In contrast, somatostatin and dexamethasone significantly inhibited CRF-stimulated increase of cAMP production and [125I]IL-1alpha binding, suggesting a primary role for cAMP in the regulation of pituitary IL-1 receptors. Next, we investigated the modulation of IL-1beta levels and IL-1 receptors following infection of mice with the endotoxin, lipopolysaccharide (LPS). Acute administration of low doses of endotoxin (30 mu g LPS/mouse) dramatically increased IL-1beta levels and reciprocally decreased [125I]IL-1alpha binding in peripheral tissues (pituitary, testis, liver, and spleen) but not in brain (hippocampus). This effect appeared to be dose related since higher doses of endotoxin (300 mu g LPS/mouse) significantly decreased [125I]IL-1alpha binding in both peripheral tissues and brain. Endotoxin induced modulation of the IL-1 system was also dependent on the treatment regimen since two low-dose LPS injections (at 0 and 12 h) increased IL-1beta concentrations and decreased [125I]IL-1alpha binding in both central and peripheral tissues. These data provide further support for a role for IL-1 in coordinating brain-endocrine-immune responses to stress and infection.

Anesthetics, Inhalation↗

PubMed1995

Nitric oxide plays no role in ACTH release induced by interleukin-1 beta, corticotropin-releasing hormone, arginine vasopressin and phorbol myristate acetate in rat pituitary cell cultures.

To investigate the role of nitric oxide (NO) in pituitary ACTH secretion, the effect of a nitric oxide synthase (NOS) inhibitor, N omega-Nitro-L-arginine (Nitro-Arg), on ACTH secretion induced by interleukin (IL)-1 beta, corticotropin releasing hormone (CRH), arginine vasopressin (AVP) and phorbol myristate acetate (PMA) was examined in rat anterior pituitary cell cultures. Nitro-Arg did not affect IL-1 beta-induced ACTH release in pituitary cell cultures incubated with Dulbecco modified Eagle's medium, Basal Medium Eagle or Krebs Ringer bicarbonate-glucose buffer. It did not affect CRH-, AVP- or PMA-induced ACTH release either. These results suggest that NO does not play an important role in ACTH release at the pituitary level.

Adrenocorticotropic Hormone↗

PubMed1995

[A case of Churg-Strauss syndrome in which MPO-ANCA (antibodies to myeloperoxidase) appeared to reflect the disease activity].

A 42-year-old housewife was admitted to our hospital because of an asthma attack, fever, severe eosinophilia, mononeuritis-multiplex, arthralgia, skin-eruptions, transient pulmonary infiltration, and other symptoms. Chung-Strauss syndrome was diagnosed. When there were signs and symptoms of vasculitis, chest radiography revealed that pulmonary infiltrates had decreased, but her chest CT showed ground glass opacities of both lung fields and fine granular shadows. Differential cell count of bronchoalveolar lavage fluid revealed a high level of eosinophils, and transbronchial lung biopsy specimens showed vasculitis and marked infiltration of eosinophils. When signs and symptoms of vasculitis were prominent, antineutrophil cytoplasmic antibodies (ANCA) in the serum were examined. PR3-ANCA (antibodies to serine-protease, nearly equal to c-ANCA), which is specific to Wegener's granulomatosis, was negative, but MPO-ANCA (antibodies to myeloperoxidase, nearly equal to p-ANCA) was positive and the level was very high (478 EU/ml). After corticosteroid therapy, her symptoms markedly improved, and MPO-ANCA became negative. MPO-ANCA appeared to reflect the disease activity in this case.

Adult↗

PubMed1995

[A case of hypersensitivity pneumonitis probably caused by a humidifier in winter].

A 64-year-old man was admitted complaining of cough, hemoptysis, dyspnea, and fever. His chest X-ray film on admission showed reticulo-granular shadows in both lung fields. Ausculation of his chest revealed fine crackles in both lower zones. After admission, he was treated with antibiotics, but his chest-radiographic appearance worsened temporarily, and sputum cytology results were repeatedly positive. Diagnosis was difficult. Differential cell count of the bronchoalveolar lavage fluid showed lymphocytosis, with a high CD 4/8 ratio. Transbronchial lung biopsy specimens revealed Masson bodies and alveolitis. With antibiotic therapy alone, his condition improved, and he was discharged. Five and a half hours later, his symptoms worsened and he was readmitted. His chest X-ray film on the second admission was almost the same as that on the first admission. His symptoms became less severe, and his condition improved without treatment. Hypersensitivity pneumonitis was diagnosed. Because the onset was in February, this was not considered to be a case of so-called summer type hypersensitivity pneumonitis a provocation test was done with water from the humidifier he had been using at home. The white blood cell count increased and PaO2 decreased significantly, so the result of the provocation test was thought to be positive. Therefore, so-called humidifier lung was strongly suspected. The results of sputum cytology on the first admission were probably falsely positive.

Alveolitis, Extrinsic Allergic↗

PubMed1995

[Five cases of drug-induced pneumonitis due to Sho-saiko-to or interferon-alpha or both].

We encountered five cases of drug-induced pneumonitis due to Sho-saiko-to or interferon-alpha or both. In all 5 cases the underlying disease was chronic hepatitis or liver cirrhosis caused by hepatitis C virus. Interferon-alpha alone was administered in one case, Sho-saiko-to alone was administered in two cases, and both were administered in two cases. Bronchoalveolar lavage was done in 4 cases. In three cases, lymphocytosis and abnormally low CD4/8 ratios were found on examination of bronchoalveolar lavage fluid. In the only case in which interferon-alpha alone was given the percentage of neutrophils in bronchoalveolar lavage fluid was abnormally high, and the adult respiratory distress syndrome developed. Lymphocyte stimulation tests were done in four cases, and in all four cases the only positive results were against Sho-saiko-to or against interferon-alpha. The frequency of drug-induced pneumonitis among patients with chronic hepatitis or liver cirrhosis was 0.7% in those given only Sho-saiko-to, 0.5% in those given only interferon-alpha, and 4.0% in those given both interferon-alpha and Sho-saiko-to. Therefore, pneumonitis due to Sho-saiko-to and to interferon-alpha is more likely to occur if these two drugs are given simultaneously.

Aged↗

PubMed1995

[Experience with the Groshong port in a home TPN patient--a case report].

We experienced a home TPN patient using Groshong port. A 45-year-old woman, recurrent pancreatic head cancer, was treated with home TPN. The Groshong port was inserted via external jugular vein and stayed at home for 8 months with home TPN. Lipid emulsion was administered twice a week. After administration of lipid emulsion, 20 ml of saline was injected and catheter lock was performed with only saline. Heparin was not used for catheter lock. After 8 months' home TPN, patient died. We could not find any precipitate in the catheter and reservoir. A unique feature of Groshong port is a pressure-sensitive two-way valve at the intravascular end. The valve of this port opens outward during fluid infusion or bolus injection. During blood withdrawal, the valve opens inward and then closed after the procedure. Because the valve is designed to remain closed between -7 and 80 mmHg, it will not open spontaneously under normal central venous pressure conditions. This valve eliminates the need for heparin flusing. Catheter obstruction during prolonged TPN is a serious complication. Compound of heparin, lipid emulsion and electrolytes, especially calcium is now becoming one of the most important causes of catheter obstruction and catheter removal. We used the Groshong port to prevent catheter obstruction. After 8 months' use of this port, no precipitate was found in this port. We conclude that the Groshong port is useful for home TPN catheter because it can prevent catheter obstruction due to heparin-lipid-electrolytes compound.

Female↗

PubMed1994

Purification and characterization of a novel superantigen produced by a clinical isolate of Yersinia pseudotuberculosis.

A superantigen designated as Yersinia pseudotuberculosis-derived mitogen (YPM) was purified in an equal manner from both the culture supernatant and cell lysate of a clinical isolate (KUR-1) of Y. pseudotuberculosis serotype 4b. A significant proliferative response of human peripheral blood mononuclear cells to purified YPM was detectable even at a concentration of 1 pg/ml. The N-terminal sequence of YPM which included 23 amino acid residues was determined, by automated Edman degradation, as Thr-Asp-Tyr-Asp-Asn-Thr-Leu-Asn-Ser-Ile-Pro-Ser-Leu-Arg-Ile-Pro-Asn-Il e-Ala-Thr-Tyr-Thr-Gly-. This sequence differed from not only all the, hitherto, reported superantigens but also known proteins. While molecular weights of known bacterial superantigens are more than 22,000, electrospray ionization mass spectrometry showed that the molecular weight of YPM was 14524.4. These results indicate that YPM comprises a novel superantigen with substantial structural differences from other bacterial superantigens produced by Gram-positive cocci.

Amino Acid Sequence↗

PubMed1994

Characterization of interactions between transducin alpha/beta gamma-subunits and lipid membranes.

The gamma-subunits of heterotrimeric guanine nucleotide-binding regulatory proteins (G-proteins) are isoprenylated and alpha-carboxyl methylated at their COOH-terminal cysteine residues. These modifications are necessary for membrane attachment of the beta gamma complex, but a requirement of an additional factor has been proposed for the stable binding. We explored a possible contribution of the blocked amino terminus of beta-subunits of bovine photoreceptor G-protein, transducin (T alpha/T beta gamma = Gt alpha/beta 1 gamma 1), and of three beta gamma complexes (beta 1 gamma 2, beta 1 gamma 3, and beta 1 gamma 7) purified from bovine brains. Structural analyses revealed that every beta 1-subunit has an N-acetylated serine, which is unlikely to contribute to the membrane association. Since neither protease nor heat treatment of photoreceptor membranes affected the membrane binding of T beta gamma, it seems unlikely that rhodopsin (or other membrane proteins) serves as an anchor protein for accepting T beta gamma. In fact, T beta gamma bound to phospholipid large unilamellar vesicles (LUVs), of which the polar head groups strongly influenced the binding: T beta gamma alone showed 2-fold higher binding for negatively charged phosphatidylserine-LUVs than for neutral phosphatidylcholine (PC)-LUVs, while the affinity of T alpha/T beta gamma complex for the phosphatidylserine-LUVs was lower than that for the PC-LUVs. These results indicate that 1) an ionic interaction between T beta gamma and membrane surface plays an important role in the stable membrane association, and 2) the domain(s) of T beta gamma responsible for the association would be different between trimeric and dissociated states. We also found that synthetic peptides corresponding to the COOH-terminal region of T gamma inhibited T alpha-T beta gamma interaction only when the peptides were isoprenylated. This suggests that the isoprenyl moiety is located at the contact site between the subunits, not at the membrane-binding domain, when T beta gamma is complexed with T alpha.

Adenosine Diphosphate Ribose↗

PubMed1994

Reciprocal modulation of corticotropin-releasing factor and interleukin-1 receptors following ether-laparotomy stress in the mouse.

Ether-laparotomy stress resulted in a dramatic decrease in specific iodine-125-labeled ovine CRF binding ([125I]oCRF) in the pituitary at 6 h after the onset of the stress although it did not affect [125I]oCRF binding in the pituitary at 2 h after the stress. [125I]oCRF binding was unchanged in the frontal cortex after the stress. In contrast, [125I]interleukin-1 (IL-1)alpha binding was significantly increased in the pituitary at 2 h after the stress and tended to be higher than non-stressed levels at 6 h after the stress but was not statistically significant. Ether-laparotomy stress did not affect [125I]IL-1 alpha binding in hippocampus, spleen and testis at any time after the stress. Plasma adrenocorticotropic hormone (ACTH) and corticosterone were increased at 2 h after the stress. These data demonstrate complex interactions between CRF and IL-1 receptors on HPA axis during stress.

Adrenocorticotropic Hormone↗

PubMed1994

Cyclic AMP-dependent modulation of interleukin-1 receptors in the mouse AtT-20 pituitary tumor cell line.

Previous studies have demonstrated an upregulation of interleukin-1 (IL-1) receptors following treatment of mouse AtT-20 pituitary tumor cells with corticotropin-releasing factor (CRF). In the present study, we determined the modulation of IL-1 receptors and adenylate cyclase activity in AtT-20 cultures following treatment with CRF, isoproterenol, forskolin, somatostatin and dexamethasone. CRF, isoproterenol and forskolin dose-dependently increased cAMP production and [125I]IL-1 alpha binding. In contrast, somatostatin and dexamethasone significantly inhibited CRF-stimulated cAMP production and decreased both basal and CRF-mediated increases in [125I]IL-1 alpha binding. Parallel modulation of IL-1 receptors by agents that stimulate (CRF, isoproterenol and forskolin) or inhibit (somatostatin) cAMP production in AtT-20 cells suggest the importance of this second messenger in regulating IL-1 receptors.

Adrenocorticotropic Hormone↗