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Biomedical subjects

S Nagata

Publications and source records attributed to S Nagata.

At least 19 recordsLinked to original sources
PubMed1992

Constitutive and inducible factors bind to regulatory element 3 in the promoter of the gene encoding mouse granulocyte colony-stimulating factor.

The expression of the mouse gene (G-CSF) encoding granulocyte colony-stimulating factor is controlled by at least three regulatory elements, GPE1, GPE2 and GPE3 (G-CSF promoter elements). A set of 30-mer oligodeoxyribonucleotides (oligos) scanning the GPE3 region (-104 to -51) of the G-CSF promoter was synthesized, and the tetramer of each oligo was inserted upstream from the cat gene with the simian virus 40 enhancer element. By introducing these hybrid genes into human squamous carcinoma CHU-2 and mouse macrophage BAM3 cells, the enhancer core element of the GPE3 was localized to the region from -98 to -79 in the promoter. A nuclear factor which specifically binds to the core element of the GPE3 was constitutively detected in human CHU-2 cells, whereas the expression of a similar, but distinctly different, factor was significantly induced in BAM3 cells by lipopolysaccharide. The results suggest that these nuclear factors play important roles in the constitutive expression of G-CSF in CHU-2 cells and its inducible expression in macrophages.

Animals↗
PubMed1992

[Echocardiography-guided transseptal left atrial puncture: a safe approach to percutaneous transvenous mitral commissurotomy].

Transseptal left atrial puncture has generally been carried out under fluoroscopic control. In our experience, cardiac tamponade occurred in 3 of 83 cases during transseptal puncture for percutaneous transvenous mitral commissurotomy (PTMC) and percutaneous transvenous aortic valvuloplasty (PTAV). We tried to perform the puncture under simultaneous fluoroscopic and 2DE guidance to decrease the rate of complications. As a result, no complication has occurred in 55 cases. Furthermore, 2DE indicated that directing the puncture needle towards 4 or 5 o'clock was inappropriate, but that 3 o'clock was an appropriate direction for puncture of the fossa ovalis of a left atrium dilated because of mitral stenosis. In conclusion, transseptal left atrial puncture can safely and easily be carried out under a combination of fluoroscopic and 2DE guidance, as this procedure displays the correct position and direction of the needle tip for interatrial septal puncture.

Adolescent↗
PubMed1992

Rational design of potent antagonists to the human growth hormone receptor.

A hybrid receptor was constructed that contained the extracellular binding domain of the human growth hormone (hGH) receptor linked to the transmembrane and intracellular domains of the murine granulocyte colony-stimulating factor receptor. Addition of hGH to a myeloid leukemia cell line (FDC-P1) that expressed the hybrid receptor caused proliferation of these cells. The mechanism for signal transduction of the hybrid receptor required dimerization because monoclonal antibodies to the hGH receptor were agonists whereas their monovalent fragments were not. Receptor dimerization occurs sequentially--a receptor binds to site 1 on hGH, and then a second receptor molecule binds to site 2 on hGH. On the basis of this sequential mechanism, which may occur in many other cytokine receptors, inactive hGH analogs were designed that were potent antagonists to hGH-induced cell proliferation. Such antagonists could be useful for treating clinical conditions of hGH excess, such as acromegaly.

Antibodies, Monoclonal↗
PubMed1992

The rat peptidylarginine deiminase-encoding gene: structural analysis and the 5'-flanking sequence.

Genomic clones of the rat peptidylarginine deiminase (PAD)-encoding gene (PAD) were isolated, and the gene organization was analyzed by restriction mapping and nucleotide sequencing. The PAD spans more than 50 kb and contains 16 exons and 15 introns. The lengths of the introns from 0.5 kb to more than 16.5 kb. A 1.7-kb sequence in the 5'-flanking region was determined. S1 nuclease mapping revealed two putative cap sites 79 and 81 bp upstream from the N-terminal ATG codon of PAD, which had been determined by amino acid sequence analysis. This ATG was confirmed to be the translation start site, since no other ATG codon was found in the open reading frame downstream from the cap sites. The 5'-flanking sequence contains four potential SP1-binding sites, a putative Pit-1/GHF-1-binding site, four short sequences either identical or homologous to the sequences in the promoter regions of rat or human growth hormone encoding genes, as well as a sequence similar to an estrogen-responsive element. However, neither a typical TATAA box, nor CCAAT box is present. These results provide important clues for elucidating the mechanism of female-specific and/or sex cycle-dependent gene expression.

Amino Acid Sequence↗
PubMed1992

mRNA capping enzyme. Isolation and characterization of the gene encoding mRNA guanylytransferase subunit from Saccharomyces cerevisiae.

The highly purified yeast mRNA capping enzyme is composed of two separate chains of 52 (alpha) and 80 kDa (beta), responsible for the activities of mRNA guanylyltransferase and RNA 5'-triphosphatase, respectively (Itoh, N., Yamada, H., Kaziro, Y., and Mizumoto, K. (1987) J. Biol. Chem. 262, 1989-1995). The gene encoding the mRNA guanylyltransferase subunit (alpha subunit), CEG1, has been isolated by immunological screening of a yeast genomic expression library in lambda gt11 with polyclonal antibodies directed against purified yeast capping enzyme. The identity of CEG1 was confirmed by epitope selection and by expressing the gene in Escherichia coli to give a catalytically active mRNA guanylyltransferase. The gene is present in one copy per haploid genome, and encodes a polypeptide of 459 amino acid residues. From its primary structure as well as its mRNA size, it was concluded that the alpha and the beta subunits of yeast mRNA capping enzyme are encoded by two separate genes, not as a fused protein. CEG1 is located on the chromosome VII by a pulse-field gel electrophoresis. Gene disruption experiment indicated that CEG1 is essential for the growth of yeast. We have also found another open reading frame (ORF2) which lies in close proximity to CEG1 in our clones and encodes a 450 amino acid-polypeptide of yet unknown function.

Amino Acid Sequence↗
PubMed1992

Lymphoproliferation disorder in mice explained by defects in Fas antigen that mediates apoptosis.

Fas antigen is a cell-surface protein that mediates apoptosis. It is expressed in various tissues including the thymus and has structural homology with a number of cell-surface receptors, including tumour necrosis factor receptor and nerve growth factor receptor. Mice carrying the lymphoproliferation (lpr) mutation have defects in the Fas antigen gene. The lpr mice develop lymphadenopathy and suffer from a systemic lupus erythematosus-like autoimmune disease, indicating an important role for Fas antigen in the negative selection of autoreactive T cells in the thymus.

Amino Acid Sequence↗
PubMed1992

cDNA clones encoding leucine-zipper proteins which interact with G-CSF gene promoter element 1-binding protein.

The gene expression of granulocyte colony-stimulating factor (G-CSF) is induced by lipopolysaccharide (LPS). GPE1, a cis-controlling element of the G-CSF gene, functions as an LPS-responsive element. GPE1-binding protein (GPE1-BP), a leucine-zipper protein, did not independently activate G-CSF gene expression. Protein blot analysis with biotinylated GPE1-BP revealed that there were nuclear proteins that interact specifically with GPE1-BP. Three leucine-zipper proteins were isolated from mouse cDNA expression libraries by this method: NF-IL6, ATF4, and a novel ATF4-related ATFx. The interactions of these proteins with GPE1-BP may play key roles in G-CSF gene expression.

Amino Acid Sequence↗
PubMed1992

The cDNA structure, expression, and chromosomal assignment of the mouse Fas antigen.

The cell surface Fas antigen is a membrane-associated polypeptide which can mediate apoptosis. cDNA clones encoding the Fas antigen were isolated from a cDNA library constructed with mRNA from the mouse macrophage cell line BAM3. The nucleotide sequence and the deduced amino acid sequence of the mouse Fas antigen were 58.5 and 49.3% identical, respectively, to the corresponding sequences of human Fas antigen cDNA. The mouse Fas antigen consists of 306 amino acids with a calculated Mr of 34,971 and contains a single transmembrane domain which divides the molecule into extracellular and cytoplasmic domains. A 2.1-kb mRNA coding for the Fas antigen was detected in the mouse thymus, heart, liver, and ovary but not in brain and spleen. The expression of the Fas antigen gene in mouse fibroblast L929 and macrophage BAM3 cell lines was significantly induced by treatment with IFN-gamma but not by IFN-alpha/beta. Interspecific backcross analysis indicated that the gene coding for the Fas antigen is in the distal region of mouse chromosome 19.

Amino Acid Sequence↗
PubMed1992

Chromosomal gene organization of the human granulocyte colony-stimulating factor receptor.

The human chromosomal gene for the granulocyte CSF (G-CSF) receptor was molecularly cloned from a human gene library. The gene is about 16.5 kb long, and present in a single copy per haploid human genome. The human G-CSF receptor gene consists of 17 exons, and the sequences of exons are completely identical to those of cDNAs isolated from human U-937 myeloid leukemia or placenta cDNA libraries. The G-CSF receptor can be subdivided into several regions: an Ig-like domain, a cytokine receptor homologous domain, three fibronectin type III domains, a transmembrane domain, and a cytoplasmic region. Exons 3-17 code for the G-CSF receptor protein, and each subdomain of the receptor is encoded by a set of exons. Primer extension analysis of the G-CSF receptor mRNA identified major and minor transcription start sites. There is no canonical "TATA" box upstream of the CAP site. About 110 nucleotides upstream of the transcription initiation site of the gene, there is an element of 18 nucleotides that is homologous to the sequences found in the promoter of human myeloperoxidase and neutrophil elastase genes.

Amino Acid Sequence↗
PubMed1992

Increased peptidylarginine deiminase expression during induction of prolactin biosynthesis in a growth-hormone-producing rat pituitary cell line, MtT/S.

Insulin and type I insulin-like growth factor (IGF-I) suppressed growth hormone (GH) expression followed by the induction of prolactin (PRL) biosynthesis in MtT/S cells cultured with normal sera. Insulin also increased the peptidylarginine deiminase activity in a dose-dependent manner. The increase was detectable at 1 ng/ml and reached a maximum (about 16-fold higher than the control) at 1 micrograms/ml. IGF-I showed similar but less prominent effects. The enzyme activity started to increase by 15 hr after the addition of insulin (500 ng/ml), and reached a plateau level at 48 hr. There were concurrent increases in the enzyme mRNA level, enzyme biosynthesis, and enzyme protein contents detected by Northern blot hybridization, [35S]-amino-acid incorporation, and Western immunoblot analysis, respectively. Two-color immunofluorescence staining at 1 day after the insulin addition detected a small number of peptidylarginine-deiminase-positive cells (about 1% of the total cells) which were also GH-positive. The enzyme-positive cells increased to 12% on day 2 and to 24-26% on days 4-6. PRL-positive cells first appeared in the enzyme-positive cell population on day 2, and PRL-positive, enzyme-negative cells appeared later. These results suggest that peptidylarginine deiminase expression increases in association with the hormone switching in MtT/S cells. When the cells were cultured in a steroid-depleted medium, insulin failed to increase the enzyme activity. The insulin action could be specifically restored by estrogen, indicating estrogen-insulin synergism in regulation of the enzyme expression.

Animals↗
PubMed1992

Identification of epitopes associated with different biological activities on the glycoprotein of vesicular stomatitis virus by use of monoclonal antibodies.

Thirteen monoclonal antibodies (MAbs) to the glycoprotein (G) of vesicular stomatitis virus (VSV) serotype Indiana were prepared and examined for their effects on various biological activities of VSV, including in vitro infection, hemagglutination, adsorption to cells, and mediation of cell fusion. Competitive binding assays with these MAbs revealed the presence of at least seven distinct antigenic determinants (epitopes) on the G protein. In some cases, overlappings among epitopes to various degrees were observed as partial inhibition or binding enhancement. The MAbs to all the epitopes but one (epitopes 1-6) reacted with the denatured G protein in a Western immunoblot analysis. Four of the epitopes (epitopes 2, 4, 5, and 7) were involved in neutralization and two (epitopes 1 and 2) in hemagglutination inhibition. None of the MAbs inhibited the adsorption of radiolabeled VSV to BHK-21 cells; the MAbs to epitope 2 slightly enhanced the virus adsorption. All neutralization epitopes except epitope 2 (epitopes 4, 5, and 7) were associated with inhibition of VSV-mediated cell fusion. These results show a direct spatial relationship between the epitopes recognized by the MAbs and functional sites on G protein and further insights into the structure and function of G protein.

Adsorption↗
PubMed1992

Direct secretion from left atrium and pulmonary extraction of human atrial natriuretic peptide.

To evaluate direct secretion from the left atrium and pulmonary extraction of human atrial natriuretic peptide (hANP), we measured plasma hANP levels in the pulmonary artery, pulmonary vein, and left atrium in patients with either mitral stenosis or atrial septal defect. Left atrial pressure in patients with mitral stenosis was significantly higher than that in patients with atrial septal defect (7.5 +/- 1.0 mm Hg vs 3.1 +/- 0.5 mm Hg, p less than 0.01). The significant increase in the hANP level in the left atrium was recognized only in patients with mitral stenosis (149 +/- 33 pg/ml in the left atrium vs 130 +/- 28 pg/ml in the pulmonary vein, p less than 0.05). The plasma hANP level in the pulmonary vein was significantly lower than that in the pulmonary artery in both patients with mitral stenosis and those with atrial septal defect, which suggests that hANP is extracted in the lung. We conclude that hANP is secreted not only through the coronary sinus but also directly from the left atrium, stimulated by high left atrial pressure, and that circulating hANP is partially extracted in the pulmonary circulation.

Adult↗
PubMed1992

Hemifacial spasm due to tumor, aneurysm, or arteriovenous malformation.

The authors report eight cases of so-called symptomatic hemifacial spasm. They had gross pathological lesions such as a tumor (one epidermoid, one neurinoma, and two meningiomas), vascular malformation (one medullary venous malformation and two arteriovenous malformations), and aneurysm. In all four cases with a tumor, no artery compressed the facial nerve at the root exit zone. In three of the four cases, the hemifacial spasm disappeared after removal of the tumor in contact with the facial nerve. Compression or encasement of the facial nerve by the tumor was the pathogenesis of the hemifacial spasm in these three cases. The remaining case with tumor (tentorial meningioma) did not have a mass or vessel that directly compressed the facial nerve at the root exit zone. However, the hemifacial spasm disappeared after the removal of the tumor. In a case with a medullary venous malformation with arterial component, an engorged draining vein compressed the root exit zone of the facial nerve. In the remaining three vascular cases--two cases of arteriovenous malformation and a case of saccular aneurysm--enlarged feeding arteries and an aneurysm directly compressed the root exit zone of the facial nerve. Not only arterial or venous but also mass compression can cause hemifacial spasm in some symptomatic cases. Surgical decompression of the facial nerve from the causative organic lesion is the primary choice of treatment.

Adult↗
PubMed1992

Clinical application of transpulmonary contrast-enhanced Doppler technique in the assessment of severity of aortic stenosis.

OBJECTIVE: The aim of this study was to demonstrate the clinical usefulness of the transpulmonary contrast-enhanced Doppler technique by using it to assess the severity of aortic stenosis. BACKGROUND: Sonicated albumin microbubbles can pass through the pulmonary circulation after peripheral venous injection and have been reported to enhance Doppler signals from the left side of the heart. Therefore, their use to determine aortic flow velocity would facilitate the assessment of the severity of aortic stenosis. METHODS: Twenty-two patients with aortic stenosis and seven normal volunteers were examined. Aortic flow velocity was recorded with continuous wave Doppler technique from an apical window before and after injection of 2 ml of sonicated albumin. RESULTS: In 10 patients with aortic stenosis, the aortic velocity envelope was too indistinct to determine the peak velocity before sonicated albumin was injected. After injection, the aortic flow Doppler signal was enhanced in 9 of the 10 patients and the velocity envelope became clear enough to measure the peak velocity, enabling calculation of the transaortic pressure gradient. In the remaining 12 patients with aortic stenosis and in all 7 normal volunteers, the velocity envelope was clear before injection and became much clearer after injection. The calculated transaortic pressure gradient showed a good agreement with catheterization measurements (y = 1.1x-6.5, r = 0.88, p less than 0.001, SEE = 16 mm Hg, n = 13). Duration of Doppler signal enhancement was measured as the time during which the envelope was clearer than before injection throughout the ejection period. The duration was significantly shorter in patients with aortic stenosis than in normal volunteers (16 +/- 5 vs. 52 +/- 32 s, p less than 0.01). There was a significant correlation between left ventricular systolic pressure measured by catheterization and the duration of signal enhancement (r = -0.69), suggesting that albumin microbubbles were fragile at high pressure. CONCLUSIONS: The transpulmonary contrast-enhanced Doppler technique using sonicated albumin is useful for assessing the severity of aortic stenosis even in patients with poor Doppler recordings, although the duration of signal enhancement might be affected by left ventricular systolic pressure.

Aortic Valve Stenosis↗
PubMed1992

Functional expression and tissue distribution of a novel receptor for vasoactive intestinal polypeptide.

Vasoactive intestinal polypeptide (VIP), a 28 amino acid peptide hormone, plays many physiological roles in the peripheral and central nerve systems. A functional cDNA clone of the VIP receptor was isolated from a rat lung cDNA library by cross-hybridization with the secretin receptor cDNA. VIP bound the cloned VIP receptor expressed in mouse COP cells and stimulated adenylate cyclase through the cloned receptor. The rat VIP receptor consists of 459 amino acids with a calculated Mr of 52,054 and contains seven transmembrane segments. It is structurally related to the secretin, calcitonin, and parathyroid hormone receptors, suggesting that they constitute a new subfamily of the Gs protein-coupled receptors. VIP receptor mRNA was detected in various rat tissues including liver, lung, intestines, and brain. In situ hybridization revealed that VIP receptor mRNA is widely distributed in neuronal cells of the adult rat brain, with a relatively high expression in the cerebral cortex and hippocampus.

Amino Acid Sequence↗
PubMed1992

Hyperproduction of human interferon gamma by rat cells maintained in low-serum medium using the fibronectin gene promoter.

An expression vector, pF1900M, which expresses a cloned gene at a high level in quiescent mammalian cells was constructed using the rat fibronectin (FN) promoter. Human interferon gamma (HuIFN-gamma) cDNA inserted downstream of the FN promoter in pF1900M was introduced into rat 3Y1 cells and several IFN-producing cell lines were established. These cells secreted a low level of IFN when they were growing but secreted at a high level after they had reached confluence. The level was further increased when the confluent cells were maintained in low-serum medium and a cell line, I7, produced 4 x 10(5) IU/ml of IFN, comparable to that produced by genetically engineered Escherichia coli in 2 days. The IFN-producing ability of I7 cells could be maintained by successive replacements of low-serum medium for at least 2 weeks. HuIFN-gamma secreted into the medium had a molecular weight range of 22,000 to 25,000, similar to that of IFN-gamma produced by human lymphocytes. The N-linked glycosylation of HuIFN-gamma seemed to occur properly, since treatment of the IFN with N-glycanase resulted in a reduction of molecular weight to 17,000, which corresponds to that calculated from the deduced amino acid sequence of HuIFN-gamma.

Animals↗