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R Inoue

Publications and source records attributed to R Inoue.

At least 109 records · Page 6Linked to original sources
PubMed1997

Increase in negative supercoiling of plasmid DNA in Escherichia coli exposed to cold shock.

Negative supercoiling of plasmid DNA in Escherichia coli cells can decrease transiently when exposed to heat shock. The effect of cold shock on DNA supercoiling was examined, and analysis by agarose gel electrophoresis in the presence of chloroquine revealed that negative supercoiling of plasmid DNA in cells increased when cells were exposed to cold shock. This increase was transient and was nil when the cells were pretreated with nalidixic acid, an inhibitor of DNA gyrase. In a mutant deficient in expression of HU protein, the increase in negative supercoiling of DNA by cold shock is less apparent than in wild-type cells. It is proposed that DNA gyrase and HU protein have a role in the DNA supercoiling reaction seen with cold shock.

Bacterial Proteins↗
PubMed1997

Reduced expression of the interferon-gamma messenger RNA in IgG2 deficiency.

The specific defect that causes IgG2 deficiency, which is one of the primary immunodeficiencies, is unknown. Recently, it was shown that interferon-gamma (IFN-gamma) induces synthesis of human germline C gamma 2 transcripts. In the authors' previous study and the present one, peripheral blood lymphocytes (PBLs) of all five tested patients with IgG2 deficiency failed to produce enough IFN-gamma when stimulated with phytohaemagglutinin or concanavalin A although they produced a sufficient amount of interleukin-2 (IL-2). The low level of IgG2 production in pokeweed mitogen-stimulated PBLs of four tested patients was improved by the addition of recombinant IFN-gamma. In this study, the amount of IFN-gamma messenger RNA showed various degrees of reduction in all five tested patients. Sequence analysis of the IFN-gamma coding regions and flanking regions revealed neither a point mutation nor a deletion for any of the patients. Thus the results suggest that the reduced expression of IFN-gamma messenger RNA may play an important role in the IgG2 deficiency of these patients.

Antigens, CD↗
PubMed1997

Identification of autophosphorylation sites in c-Yes purified from rat liver plasma membranes.

c-Yes was purified 322-fold from a rat liver plasma membrane fraction to a single 60-kDa band on SDS-PAGE. The purified protein contained essentially no phosphotyrosine residues and was autophosphorylated with Mg2+. ATP exclusively at tyrosine residues with a concomitant increase in the protein-tyrosine kinase activity. The autophosphorylated c-Yes was extensively digested by trypsin and the resultant two major phosphopeptides, peptides I and II, were purified by HPLC on a reversed-phase C-18 column. The amino acid sequence of peptide I was determined to be LIEDNEYTAR, which is identical with the sequence from Leu-418 through Arg-427 of mouse c-Yes, indicating that one of the autophosphorylation sites corresponds to Tyr-424 of the mouse c-Yes. After partial determination of the N-terminal sequence of 10 amino acid residues of peptide II, the 230 bp sequence of rat cDNA that encodes the N-terminal 76 amino acid residues of c-Yes covering peptide II, was determined. From the predicted amino acid sequence, the sequence of peptide II was assumed to be from Tyr-16 through Lys-46, YTPENPTEPVNTSAGHYGVEHATAATTSSTK. The purified c-Yes phosphorylated the tyrosine residue of synthetic peptides covering Tyr-32 and its surrounding sequence but did not phosphorylate peptides covering Tyr-16 and its surrounding sequence, suggesting that the other autophosphorylation site is Tyr-32.

Amino Acid Sequence↗
PubMed1997

Tissue and subcellular distributions, and characterization of rat brain protein phosphatase 2A containing a 72-kDa delta/B" subunit.

A 74-kDa delta/B" subunit was isolated by heparin-Sepharose column chromatography from human erythrocyte protein phosphatase 2A (PP2A) consisting of a 34-kDa catalytic subunit (alpha/C) and 63- and 74-kDa regulatory subunits (beta/A and delta/B") in a ratio of 1:1:1. The purified delta/B" was used as an immunogen in mice, to prepare specific antisera against delta/B". Immunoblot analyses with the antisera detected an immunoreactive 72-kDa protein in the cytosol from various rat tissues including erythrocytes, brain, lung, testis, adrenal gland, heart, spleen, kidney, and liver. The 72-kDa protein was highly abundant in brain and was distributed evenly in cerebral cortex, cerebellum, and brain stem. The 72-kDa protein was also detected in mitochondria and microsome fractions. An immunoreactive 68-kDa protein was detected mainly in nuclear and microsome fractions. The 72-kDa protein from rat brain cytosol copurified with phosphorylated H2B histone phosphatase activity during successive chromatographies on DEAE-Toyopearl, AH-Sepharose, Sephadex G-150, H1 histone-Toyopearl, TSK DEAE-5PW, protamine-Toyopearl, and TSK G3000SW columns. The purified enzyme migrated as a single protein band on nondenaturing PAGE and as three protein bands of 34, 63, and 72 kDa in a ratio of 1:1:1 on SDS-PAGE. The molecular weight of the enzyme was estimated to be 170,000 from the s20,W value of 7.2 +/- 0.3 S and the Stokes radius of 5.5 +/- 0.1 nm. The rat brain enzyme was classified as PP2A, based on the following properties; (1) an IC50 for okadaic acid of 10(-9) M; (2) its preferential dephosphorylation of the a subunit of phosphorylase kinase; (3) its insensitivity to protein inhibitor 2; and (4) its heterotrimeric subunit structure. The Km value and the molecular activity of the enzyme for phosphorylated H2B histone were 72.3 +/- 0.3 microM and 192 +/- 2 mol Pi released/min/mol enzyme, respectively, and were comparable to those of human erythrocyte PP2A (alpha1 beta1 delta1/ CAB"). The 72-kDa subunit in the purified rat brain PP2A was phosphorylated in vitro by cAMP-dependent protein kinase.

Animals↗
PubMed1997

Preferential potentiation by hypotonic cell swelling of muscarinic cation current in guinea pig ileum.

The effects of hypotonic cell swelling (HCS) on muscarinic receptor-activated cationic current in guinea pig ileal smooth muscle were investigated by the whole cell patch-clamp technique. With nystatin-perforated recording, reduced external tonicity from 312 to 262 mosM caused cell swelling but hardly affected the membrane currents activated by depolarization, such as outward-rectifying K and voltage-dependent Ca currents. In contrast, the inward current evoked by carbachol at -60 mV was greatly increased (approximately 50%) by the same extent of hypotonicity. This effect is likely to occur through potentiation of nonselective cation channels coupled to the muscarinic receptor (mNSCCs) and probably does not involve elevated intracellular Ca2+ concentration ([Ca2+]i), since neither removal of external Ca2+ nor [Ca2+]i buffering with 10 mM 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid significantly affected the results. Furthermore, the time course and degree of this potentiation closely matched those of video-microscopically monitored HCS. These results support the view that mechanosensitive modulation may be a powerful mechanism to regulate mNSCCs activity in gut smooth muscle, together with membrane potential and [Ca2+]i.

Animals↗
PubMed1997

Identification of Dermatophagoides farinae-2-derived peptides and class II HLA molecules recognized by T cells from atopic individuals.

Der f2, the group 2 allergen of Dermatophagoides farinae, is one of the major inhalation allergens in Japan. Using the mixture of a panel of overlapping synthetic peptides that spread over the entire Der f2 molecule, we found that polyclonal Der f2-specific short-term T cell lines prepared from peripheral blood mononuclear cells of 5 individuals allergic to Der f who carry most of the common HLA haplotypes seen in the Japanese population can respond mainly to 7 different peptides. Distribution of the T cell epitopes on Der f2 was not identical with that on Der p2. Five of 7 peptides stimulated T cells of more than 2 donors, regardless of HLA types. Inhibition patterns by anti-HLA class II mAbs were heterogenous in proliferative responses of each cell line, three were mainly inhibited by anti-HLA-DR mAb, and the others were inhibited by anti-HLA-DQ mAb. One of these T cell lines, SM, of which the proliferative response was partially inhibited by anti-HLA-DQ mAb, was cloned. Indeed, the T cell clone SM4.6 was restricted by DQ6 molecules encoded for by an HLA-DRB1*1502-DRB5*0102-DQA1*0103-DQB1*0601 haplotype. These results indicate that patients' T cells recognize Der f2 in association with a variety of HLA-DR or HLA-DQ as antigen-presenting molecules. Thus, although some peptides do have a more potent T cell stimulatory activity than others, the TCR ligands formed with the Der f2 molecule are highly heterogeneous, a factor also noted in Der f1-specific T cell lines in our previous study.

Allergens↗
PubMed1997

Two Japanese siblings with Bloom syndrome gene mutation and B-cell lymphoma.

Bloom syndrome (BS) is a rare autosomal recessive genetic disorder characterized by lupus-like erythematous telangiectasia of the face, sun sensitivity, infertility and stunted growth. Upper respiratory tract and gastrointestinal infections are commonly associated with the decreased immunoglobulin levels found in BS patients. Chromosomal abnormalities are hallmarks of the disorder, and high frequencies of sister chromatid exchanges and quadriradial configurations in lymphocytes and fibroblasts are virtually diagnostic. Recently, the causative gene for BS (BLM) has been identified. We encountered and defined a family with a nonsense mutation in BLM. The brother and sister were homozygous for the mutation and both developed B-cell malignant lymphoma in their twenties. These findings indicate the importance of prenatal diagnosis and the detection of BS carriers based on molecular genetic analysis.

Adenosine Triphosphatases↗
PubMed1997

Establishment of a new cell line from a patient with hairy cell leukemia-Japanese variant.

A cell line, JHC-2, was established from the peripheral blood of a patient with hairy cell leukemia (HCL)-Japanese variant. The JHC-2 cells have cytologic features similar to those of the original tumor cells. They displayed hairy cytoplasmic projections by phase contrast and scanning electron microscopy. The tartrate-resistant acid phosphatase reaction was weakly positive. The immunophenotype of the JHC-2 cells was CD5-, CD10-, CD11c+/-, CD19+, CD21+, CD23+, CD24-, CD25+/-, CD38- and FMC-7+. The expression of surface immunoglobulin (IgG, kappa) and the configuration of Ig gene rearrangements in the JHC-2 cells were identical to those in the original leukemic cells, and the JHC-2 cells displayed trisomy 9 on cytogenetic examination. Southern blot analysis for the Epstein-Barr virus (EBV) genome showed that the JHC-2 cells contained the EBV genome, although the freshly isolated leukemic cells did not. These results indicate that the JHC-2 cell line is an EBV spontaneously transformed B cell line originating from HCL cells.

Acid Phosphatase↗
PubMed1997

[Successful use of propofol for anesthesia for plasty of eyelids in epidermolysis bullosa].

A 21-year-old male with recessive dystrophic epidermolysis, required anesthesia for plastic surgery of eyelids. We took special care for the attachment of monitoring equipment and placing an intravenous cannula. Anesthesia was induced by intravenous propofol 1 mg.kg-1 and maintained with local injection of 1.0% lidocaine and continuous infusion of propofol 3-4 mg.kg-1.hr-1. Tracheal intubation was not performed and oxygen was given through a nasal cannula and spontaneous breathing was maintained throughout the anesthetic management. Respiratory condition was monitored by measuring Spo2 and end-tidal CO2 by capnography obtained from the nasal cannula. Operation was performed successfully and he was discharged without any major complications.

Adult↗
PubMed1996

Microsatellite instability in B-cell lymphoma originating from Bloom syndrome.

Bloom syndrome (BS) is a rare autosomal recessive genetic disorder characterized by lupus-like erythematous telangiectasias of the face, sun sensitivity, stunted growth infertility and immunodeficiency. In addition, BS patients are highly predisposed to cancers. Although recently the causative gene of BS (BLM) was identified as a DNA helicase homologue, the function of BLM in DNA replication has not been elucidated. In this study, p53 mutation and microsatellite instability in B-cell lymphomas originating from 2 sibling BS patients were investigated. In the originally developed tumor of both patients, no p53 mutation was detected. In one patient, however, after treatment by ionizing radiation the B-cell lymphoma recurred, showing a 9-bp deletion in exon 7. In lymphoma cells and an EB-virus-transformed cell line from BS lymphocytes of this patient, microsatellite instability was also detected from the reduced length of microsatellite DNA markers, although in the other patient microsatellite instability was not detected. Thus, 2 B-cell lymphomas, despite having the same BLM mutation, showed different phenotypes in terms of p53 mutation and microsatellite instability.

Adult↗
PubMed1996

VLA-5 in the plasma cell line, FR4ds, acts as a common regulator of VLA-4 and VLA-6 in spreading induced by fibronectin and laminin.

VLA-5 recognizes the GRGDSP sequence of fibronectin (FN) in the extracellular matrix (ECM). We examined the role of beta1 integrin in the spreading of the human plasma cell line, FR4ds, induced by FN and laminin (LN). We first examined the role of VLA-5 in the spreading induced by FN. Anti-alpha4 antibody induced 46.4% inhibition, whereas anti-alpha5 had no effect. A combination of anti-alpha4 and anti-alpha5 enhanced the inhibition of spreading significantly. Complementary inhibition was also demonstrated using the GRGDSP peptide plus anti-alpha4 and the GRGDNP peptide of LN plus anti-alpha4. The results suggested that VLA-5 is a regulator of VLA-4 and that it is involved in the recognition of GRGDNP. We then examined the role of VLA-5 in the spreading induced by LN. Anti-alpha6 induced 53.1% inhibition. Anti-alpha5 alone had no effect. A combination of alpha5 and anti-alpha6, however, significantly enhanced the inhibition of spreading. The combination of GRGDSP plus anti-alpha6 and GRGDNP plus anti-alpha6 resulted in complete inhibition. These results suggested that VLA-5 participates in the recognition of LN cooperatively with VLA-6 and that VLA-5 is a common regulator of VLA-4 and the LN receptor, VLA-6.

Cell Movement↗
PubMed1996

Rare activation of the human c-Ha-ras transgene of mice in hemangioendothelial sarcomas and liver tumors induced by Glu-P-1.

A transgenic mouse (Tg), having the human c-Ha-ras proto-oncogene, has been demonstrated to develop hemangioendothelial sarcomas (HESs) which are associated with the transgene mutation, but not to develop liver tumors. In this study, we examined the effects of 2-amino-6-methyldipyrido [1,2-a:3',2'-d] imidazole (Glu-P-1), a food-borne carcinogen, which has been demonstrated to induce HESs and liver tumors in CDF1 mice, on Tg mice. Chronic administration of 0.05% Glu-P-1 in the diet induced HESs in Tg (7/17), but not in 18 non-transgenic mice (N-Tg). With basal diet, two out of 17 Tg but none of 22 N-Tg, developed HESs. In contrast, Glu-P-1 administration induced liver tumors, both in Tg and in N-Tg; 16/17 in Tg and 13/18 in N-Tg. The incidence of hepatocellular carcinomas in Tg was higher than that in N-Tg (8/17 versus 3/18). With basal diet, only one out of 17 Tg and none of 22 N-Tg developed liver tumors. The Ha-ras mutation in tumors developed by the groups administered Glu-P-1, was examined. No mutations were detected in the transgene and mouse c-Ha-ras genes in all three HESs examined. In contrast, when 29 liver tumors taken from Tg were examined, two mutations of the transgene were detected in two HCCs. No mouse c-Ha-ras gene mutations were detected in any of the 47 liver tumors examined, which had developed in Tg and N-Tg mice. These results suggest that the transgene plays a role in the development of HESs induced by Glu-P-1, but not as a result of its mutation. Further, the transgene plays no significant role in the development of liver tumors induced by Glu-P-1, but does play a role in the malignant conversion of some liver tumors, as a result of its mutation.

Animals↗
PubMed1996

Electrophysiological characterization of a motilin agonist, GM611, on rabbit duodenal smooth muscle.

Effects of motilin and a newly synthesized erythromycin derivative, GM611, on membrane potential and currents of rabbit duodenal smooth muscle have been investigated by intracellular potential recording and whole cell patch-clamp technique and compared with results from contractile experiments. Motilin and GM611 (0.01-100 nM) dose dependently produced slowly sustained depolarizations (half-maximal effective dose = 0.15 and 3.9 nM for motilin and GM611, respectively) but exhibited biphasic effects on spike activities superimposed on slow waves. With small depolarizations, the number of spike discharges increased, whereas larger depolarizations markedly reduced spike amplitude. Motilin-induced (or GM611-induced) depolarization appeared to be associated with the activation of monovalent cation-selective channels, and the reduction in the spike amplitude appeared mainly to be associated with inhibition of voltage-dependent Ca2+ channels. Furthermore, data from patch-clamp experiments suggested that Ca2+ release occurred from heparin-sensitive internal stores upon stimulation of motilin receptors by these agonists. Possible implications of these electrophysiological effects in motilin- or GM611-induced tonic and phasic contractions have been discussed.

Animals↗
PubMed1996

Fenamates potentiate the alpha 1-adrenoceptor-activated nonselective cation channels in rabbit portal vein smooth muscle.

The agonistic action of fenamates on the alpha 1-adrenoceptor-activated cationic current (Icat) in rabbit portal vein smooth muscle was investigated with the whole-cell patch clamp technique. At -50 mV, the fenamates (100-500 microM) increased Icat dose-dependently, up to several fold. This increase was not accompanied by changes in the reversal potential and strongly inhibited by 500 microM Cd2+ or 10 mM procaine. The enhancing effect of fenamates was also observed on the cationic current activated by intracellularly applied GTP gamma S. These results suggest that fenamates may be useful as a new class of activator for receptor-operated cation channels in smooth muscle.

Adrenergic alpha-Agonists↗
PubMed1996

Effect of ebselen on bovine and rat nitric oxide synthase activity is modified by thiols.

The inhibition of nitric oxide synthase (NOS) by ebselen, 2-phenyl-1,2-benzisoselenazole-3(2H)-one, was reversed by the addition of 10(-5) M dithiothreitol, suggesting that ebselen reacts with a critical thiol group of NOS in the inhibitory mechanism. In the presence of 10(-4) to 10(-3)M dithiothreitol, ebselen dose-dependently enhanced NOS activity, implicating another interaction of ebselen with NOS under these conditions. Thus, the effect of ebselen on the NOS activity is modified by thiols.

Animals↗
PubMed1995

Lysophosphatidylcholine inhibits endothelium-dependent hyperpolarization and N omega-nitro-L-arginine/indomethacin-resistant endothelium-dependent relaxation in the porcine coronary artery.

BACKGROUND: Oxidized LDL and lysophosphatidylcholine (LPC) have been reported to inhibit the endothelium-dependent relaxation (EDR) mediated by nitric oxide. Recently, a new vasorelaxing factor, endothelium-derived hyperpolarizing factor (EDHF), which hyperpolarizes and relaxes the porcine coronary artery in the presence of N omega-nitro-L-arginine (NNA) and indomethacin (IM), has been reported. We examined whether LPC also inhibits both the EDHF-mediated relaxation and membrane hyperpolarization of the porcine coronary artery. METHODS AND RESULTS: EDHF was evaluated as the bradykinin- or A23187-induced relaxation of the porcine coronary artery contracted by prostaglandin F2 alpha in the presence of NNA and IM. We also directly measured the membrane potential of the porcine coronary artery. The effects of LPC on both relaxation and membrane hyperpolarization were investigated. At concentrations of 0 to 20 mumol/L, LPC dose-dependently inhibited the NNA/IM-resistant EDR induced by bradykinin and A23187, and the relaxation was reversible after the absorption of LPC with albumin. LPC also inhibited the bradykinin- and A23187-induced hyperpolarization of the porcine coronary artery. CONCLUSIONS: In the present study, LPC was found to inhibit not only nitric oxide-mediated but also EDHF-mediated relaxation of the porcine coronary artery. Our findings suggest a new regulatory mechanism in the atherosclerotic coronary artery.

Animals↗