Biomedical subjects

H Ohmori

Publications and source records attributed to H Ohmori.

At least 181 records · Page 10Linked to original sources

PubMed1993

Two phosphatases for 6-phospholysine and 3-phosphohistidine from rat brain.

6-Phospholysine phosphatase (30 kDa) and 3-phosphohistidine/6-phospholysine phosphatase (150 kDa) were purified partially from rat brain cytosol. The former hydrolyzed 6-phospholysine to inorganic phosphate and lysine, and the Km value was 0.83 mM. The latter hydrolyzed 6-phospholysine or 3-phosphohistidine to inorganic phosphate and lysine or histidine, and the Km values were 0.78 and 0.37 mM for 6-phospholysine and 3-phosphohistidine, respectively. Moreover, the latter enzyme preparation dephosphorylated AMP, GMP, and p-nitrophenyl phosphate. Substrate specificities of both enzymes were distinct from those of N omega-phosphoarginine phosphatase (Kuba, M., Ohmori, H., and Kumon, A. (1992) Eur. J. Biochem. 208, 747-752) and phosphoamidase (EC 3.9.1.1), indicating that there are phosphatases corresponding to N-phosphorylated basic amino acids. The possibility that these enzymes work in vivo as protein phosphatases to regulate the level of N-phosphorylated proteins is discussed.

Animals↗

PubMed1993

The genomic diversity among equine herpesvirus-1 strains isolated in Japan.

The DNAs from nine Japanese field isolates of equine herpesvirus-1 (EHV-1) were analyzed by digestion with the restriction endonuclease Bam HI and Southern hybridization. Comparing restriction profiles among the EHV-1 strains, there was no considerable difference between isolates before and after vaccine application, but some minor variations in the mobility of Bam HI fragments were observed. To identify these variable fragments, all genomic DNA sequences of the Japanese prototype of EHV-1 have been cloned as Bam HI restriction fragments into the plasmid pUC-18. Physical maps of the virus DNA were constructed by a combination of Southern blot analysis and double enzyme digestion of the cloned fragments. By using these cloned fragments as probes in Southern blot analysis, the areas of heterogeneity observed among the field EHV-1 isolates were located in both terminals of UL, the center of UL, IR, US and TR regions of the genome.

Animals↗

PubMed1993

Suppression of IgE antibody response in mice by a naphthalene derivative, TEI-6472.

In the present report, we investigated suppressive effects of a naphthalene derivative, (7E)-N-(2-carboxyphenyl)-8-(2-naphthyl)-5,6-trans-5,6-methano-7- octenamide L-lysine salt (TEI-6472), on in vitro and in vivo antigen-specific IgE response. Anti-trinitrophenyl (TNP) IgE response induced in vitro in TNP-keyhole limpet hemocyanin (KLH)-primed murine spleen cells was suppressed by about 60% in the presence of 10(-6) M TEI-6472. On the other hand, anti-TNP IgG1 and IgM response were not significantly suppressed under the same conditions. Proliferative responses of BALB/c spleen cells stimulated by lipopolysaccharide, concanavalin A or allogeneic spleen cells were not inhibited by TEI-6472 at 10(-6)-10(-5) M. Interleukin 4 production from helper T-cell clone, D10.G4.1 was suppressed only slightly (less than 20%) at 10(-6) M TEI-6472. This compound was also effective in suppressing secondary anti-TNP IgE response in mice that were immunized twice with TNP-KLH and alum. When 10-20 mg/kg/day TEI-6472 was administered s.c. for 5 consecutive days starting from one day before the first and the second immunization, secondary anti-TNP IgE response was inhibited most strongly (40-45%). Anti-TNP IgG1 response was also inhibited but to a smaller extent (20-24%), while anti-TNP IgM response was suppressed only slightly (0-15%). These results suggest that, under appropriate conditions, TEI-6472 can suppress IgE responses more preferentially both in vitro and in vivo.

Animals↗

PubMed1993

Acetylcholine increases intracellular Ca2+ concentration and hyperpolarizes the guinea-pig outer hair cell.

Extracellularly applied acetylcholine (ACh) induced outward currents in isolated outer hair cells of a guinea-pig cochlea. The ACh induced current was carried by K+ ions. The current amplitude was ACh dose dependent with a KD of 12 microM. The ACh induced outward current was reversibly blocked by extracellularly applied atropine (1 microM), d-tubocurarine (d-TC, 1 microM), apamin (1 microM) and strychnine (0.1-10 microM). D-TC (10 microM) not only blocked the ACh induced outward current, but also reduced the amplitude of depolarization induced outward current. ACh induced a rise of intracellular Ca2+ concentration ([Ca2+]i). D-TC (10 microM) reduced but did not totally block the increase of [Ca2+]i. In a low Ca2+ (0.1 mM) extracellular medium, the amplitude of ACh induced current was reduced rapidly and was recovered gradually to the normal level after the extracellular Ca2+ concentration was resumed. It is probable that ACh hyperpolarizes the guinea-pig outer hair cell membrane by activation of a Ca(2+)-activated K+ conductance.

Acetylcholine↗

PubMed1993

Efferent synapse mechanisms in chick hair cells.

Electrical stimulation of the efferent fiber to the hair cell is known to suppress peripheral auditory function. In isolated hair cells from the chick, acetylcholine (ACh) generated membrane hyperpolarization when a puff was applied at concentrations from 1 microM to 5 mM. Ionophoretic application of ACh generated a membrane hyperpolarization with a rapid time course. This is due to the increase of K conductance and is accompanied by the rise of the [Ca]i.

Acetylcholine↗

PubMed1993

Isolation of N omega-phosphoarginine hydrolase from rat liver and its physical properties.

N omega-Phosphoarginine hydrolase from rat liver cytosol was purified to apparent homogeneity on SDS-PAGE, by employing column chromatographies on Sephadex G-75, DEAE-cellulose, QAE-Toyopearl, and glutathione-2-pyridyl-disulfide-Superose. One milligram protein of the final preparation released 4 mumol/min of inorganic phosphate from N omega-phosphoarginine. The molecular mass on SDS-PAGE, the Stokes' radius and the sedimentation coefficient were estimated to be 17.3 kDa, 1.63 nm, and 2.0 s, respectively, indicating that this enzyme consists of a single peptide. The stability of the enzyme to heat depended on the buffers employed and treatment of the enzyme preparation in 50 mM Tris-HCl, pH 7.0 at 50 degrees C, reduced the hydrolytic activity with a decay constant of 0.099 per min.

Animals↗

PubMed1993

Suppression of the slow K+ current by cholinergic agonists in cultured chick cochlear ganglion neurones.

1. Effects of cholinergic agonists on the cultured chick cochlear ganglion (CG) neurone were examined using the whole-cell patch-clamp method. 2. Acetylcholine (ACh, 0.1-100 microM) and its non-hydrolysable form, carbamylcholine (CCh, 0.1-300 microM), suppressed the outward current. The CCh-sensitive current was activated at membrane potentials more positive than -70 mV. 3. The CCh-sensitive current slowly activated after step depolarization with a time constant from 20 to 150 ms. The activation time constant decreased monotonically with depolarization of the membrane. 4. The reversal potential of CCh-sensitive current changed as a function of the external K+ concentration (-79, -65 and -44 mV in 5, 10 and 25 mM, respectively) and was approximately equal to the potassium equilibrium potential (-89, -71 and -48 mV in 5, 10 and 25 mM, respectively). The CCh-sensitive current is concluded to be K+ selective. 5. The CCh-sensitive current showed a sigmoid log dose vs. response relationship with an apparent dissociation constant (KD) of 1.4 microM and a Hill coefficient of 1.0. When ACh was applied, an apparent KD of 1.8 microM and a Hill coefficient of 1.0 was measured. 6. The suppression of K+ current by CCh was blocked by atropine (3 microM) and pirenzepine (3 microM), suggesting that the current is mediated by an M1 muscarinic receptor. 7. The CCh suppression of the K+ current was enhanced by GTP-gamma-S (0.1 mM), suggesting that a GTP-binding protein is involved. 8. The CCh suppression of the K+ current was mimicked by protein kinase C activators, 1-oleoyl-2-acetyl-sn-glycerol (OAG, 100 microM), phorbol dibutyrate (PDBu, 2 microM) and phorbol 12-myristate 13-acetate (PMA, 1 microM). The protein kinase inhibitor, staurosporine (0.2 microM) applied internally blocked the CCh suppression of the K+ current which suggests an involvement of protein kinase C.

Animals↗

PubMed1993

Antigen-specific but not polyclonal IgE response in murine B cells cocultured with Th2 clone is refractory to suppression by IL4-depletion.

We established culture conditions allowing the induction of antitrinitrophenyl (TNP) IgE response by the coculture of unprimed C3H B cells with conalbumin (CA)-specific type 2 helper T cell clone, D10.G4.1 in the presence of TNP-CA. With the help of this culture system, the role of Th2 cells and interleukin 4 (IL4) derived from them in the induction of antigen-specific IgE response was analyzed. A maximum level of anti-TNP IgE was secreted at 0.5-1 micrograms/ml TNP-CA. When the antigen was increased to 100 micrograms/ml, anti-TNP IgE production was abolished, while antigen-nonspecific (polyclonal) IgE was produced concomitantly. Anti-TNP IgE synthesis was not significantly suppressed in the presence of monoclonal anti-IL4 (11B11), nor was the response further enhanced by the addition of recombinant IL4. In contrast, polyclonal IgE response was abolished by 11B11. In order to deplete endogenous IL4 more strictly, we employed an antisense DNA for IL4 mRNA that can effectively inhibit IL4 production from D10 cells. Even in the presence of both 11B11 and the antisense DNA, it was found that anti-TNP IgE response was not suppressed significantly. These results suggest that antigen-specific IgE response induced in B cells cocultured with established Th2 cells does not depend on IL4 in contrast to polyclonal IgE response.

Animals↗

PubMed1993

[Tissue concentration of intravesically instilled (2"R)-4'-o-tetrahydropyranyl-adriamycin or adriamycin in superficial bladder cancer].

Twenty-one patients with superficial bladder cancer entered an analysis of single dose (2'R)-4'-O-tetrahydropyranyladriamycin (THP) or adriamycin (ADM) administration. The patients in each group that have been or not have been treated previously with anti-cancer drugs were randomized into two groups, one was given THP and the other ADM. Thirty-mg of THP or ADM dissolved in 30 ml of physiologic saline was instilled into the bladder, and retained for 1 hour. After 1 hour retention of the drugs, tumor tissues and normal mucosas were removed by punch biopsy forceps transurethrally. The tissue concentrations of THP and ADM were estimated by high performance liquid chromatography. The tissue concentrations of THP and ADM in the tumors were significantly greater (p < 0.05) than those in the normal bladder mucosas. The tissue concentration of THP in the tumors were greater than that of ADM. The tissue concentrations of THP and ADM in the tumor of patients who have been treated with anti-cancer drugs previously were less than those of patients who have not. This results demonstrated that prior therapy with anti-cancer drug may cause a resistance for intravesical instillation chemotherapy. However in patients with prior therapy, the tissue concentration of THP in the tumors were greater than that of ADM. Based on these findings, THP has been shown to be were effective as an intravesical instillating agent especially, in cases with prior chemotherapy.

Administration, Intravesical↗

PubMed1993

[Transrectal hyperthermia for the treatment of chronic prostatitis].

A total of 36 cases with chronic non-bacterial and non-chlamydial prostatitis or prostatodynia underwent 5 weekly, 1-hour sessions of transrectal microwave hyperthermia (43 degrees C) to the prostate. All patients had a long history of the condition and failed to respond to a variety of conventional treatments. The efficacy of the treatment was evaluated by the effects on subjective symptoms and/or on white blood cells in expressed prostatic secretions. Concerning the overall clinical efficacy, excellent results were obtained in 11 (30.6%), good in 8 (22.2%), fair in 8 (22.2%) and poor in 9 (25.2%). Although minor complications were noted in 5 cases (anal pain; 2, hematospermia; 1, hematuria; 1), all cases received full sessions. These results indicated the usefulness of the hyperthermia for this benign condition, which has so far responded poorly to the conventional therapy.

Adult↗

PubMed1992

Characterization of N omega-phosphoarginine hydrolase from rat liver.

N omega-Phosphoarginine hydrolase from rat liver hydrolyzed N omega-phosphoarginine into arginine and inorganic phosphate, whereas it did not release inorganic phosphate from 19 other phosphorylated compounds containing a N-P bond, an O-P bond or a C-P bond. In addition, it was not able to transfer the phosphoryl moiety from N omega-phosphoarginine to ADP. These results indicated that this enzyme was distinct from both phosphoamidase and arginine kinase. Its properties were as follows: thiol compounds were essential for its activity; it was stimulated by 1.5-2-fold in the presence of 0.001% Lubrol, Tween 20, poly(oxyethylene) 9-lauryl ether and Nonidet P-40, while 0.004% sodium lauryl sulfate inhibited the activity completely; concentrations of sodium molybdate and sodium vanadate necessary for 50% inhibition were 7 microM and 12 microM, respectively; some proteins stimulated the activity, while lysophosphatidic acid, lysophosphatidylinositol, and phosphatidic acid suppressed the activity even in the presence of poly(oxyethylene) 9-lauryl ether.

Animals↗

PubMed1992

Mapping of antigenic sites to monoclonal antibodies on the primary structure of the F1-ATPase beta subunit from Escherichia coli: concealed amino-terminal region of the subunit in the F1.

To analyze relationships between the ternary and primary structures of the beta subunit of Escherichia coli F1 ATPase, we prepared two monoclonal antibodies beta 12 and beta 31 against the beta peptide. These antibodies bind to the beta subunit but do not bind to the F1 ATPase, resulting in no inhibition of the ATPase activities. Several different portions of the beta subunit peptide were prepared by constructing expression plasmids carrying the corresponding DNA segment of the beta subunit gene amplified by the polymerase chain reaction. Western blotting analysis using these peptides revealed that the antibodies bound to a peptide of 104 amino acid residues from the amino terminal end, which is outside the previously estimated catalytic domain between residues 140 and 350. These results indicated that the amino terminal portion of the maximal 104 residues is not exposed to the surface of the F1 ATPase. The binding spectrum of the antibodies to the subunit from various species including Vibrio alginolyticus and thermophilic bacterium PS3 indicated possible epitope sequences within the 104 residues. The ternary structure of the beta subunit, in terms of cleavage sites by endopeptidases, was analyzed using the antibodies. A 43-kDa peptide without binding ability to beta 12 and beta 31 appeared upon cleavage by lysyl endopeptidase. The results suggested that lysyl residues from around 70 to 100 from the amino terminus are exposed to the surface of the beta subunit.

Amino Acid Sequence↗

PubMed1992

Immobilized ferredoxins for affinity chromatography of ferredoxin-dependent enzymes.

An immobilized ferredoxin more stable than the conventional immobilized spinach ferrodoxin was prepared by reacting CNBr-Sepharose with ferredoxins isolated from barley and Synechococcus vulcanus, a thermophilic blue-green alga. The dissociation constants of immobilized ferredoxin from spinach, barley and S. vulcanus for spinach ferredoxin-NADP reductase were 0.922, 2.505 and 5.209 microM, respectively, whereas those for barley ferredoxin-NADP reductase were 1.159, 0.579 and 2.851 microM, respectively. The order of stability was S. vulcanus greater than barley greater than spinach. The immobilized ferredoxin was applied to the simultaneous detection of ferredoxin-dependent enzymes in spinach chloroplasts. Over 20 polypeptides were detected. Synechococcus ferredoxin could also be immobilized on a Toyopearl gel and repeatedly used in an automated high-performance liquid chromatographic system.

Chemical Phenomena↗

PubMed1992

Establishment of an enzyme release assay for cytotoxic T lymphocyte activity.

In the present report, we describe the establishment of a cell line that can be used as the target for measuring the activity of cytotoxic T lymphocytes (CTL) by an enzyme release assay. We transfected P3/NS1-Ag4-1 (NS-1), a myeloma cell line derived from BALB/c mice with Escherichia coli beta-galactosidase (beta-Gal) gene, and isolated a stable transformant designated as NS-1/Z that expressed a high level of the enzyme activity intracellularly. The effector cells showing cytotoxicity against NS-1/Z were induced when the spleen cells of AKR or C3H mice were cultured with mitomycin C-treated BALB/c spleen cells for 4 days. When 2 x 10(4) NS-1/Z cells were incubated with varying numbers of effector cells, beta-Gal activity was released from the target cells depending on the number of effector cells and the time of incubation for up to 8 h. A highly sensitive enzyme assay was performed by using a fluorescent substrate, 4-methylumbelliferyl-beta-D-galactoside. The cytotoxicity was specific for H-2 haplotype of the stimulator cells, and was abolished by treating the effector cells with anti-Lyt 2 plus complement. The sensitivity of the enzyme release assay was comparable to that of 51Cr release assay. These results indicate that NS-1/Z can be used as a target cell line for the non-radioactive measurement of CTL activity.

Animals↗

PubMed1992

Enhancement of DNA transfection efficiency by heat treatment of cultured mammalian cells.

The expression of genes introduced into various mammalian cell lines was enhanced by raising the temperature of the cells to 42 degrees C for a few hours after DNA transfection. This heat treatment resulted in an up to 10-fold increase in the frequency of the cells that transiently expressed a foreign gene such as that of beta-galactosidase, whereas it had only a limited enhancing effect on the development of stable transformants. By immunotitration analysis, it was confirmed that the enhanced expression of beta-galactosidase activity correlated well with the increase of the enzyme protein. This procedure may have an applicability for augmenting the frequency of transient gene expression in many cell types.

Animals↗

PubMed1992

Intravesical instillation of adriamycin in the presence or absence of verapamil for the treatment of superficial bladder cancer: preliminary report of a collaborative study.

A case-controlled collaborative study on the intravesical administration of Adriamycin in the presence or absence of verapamil, a calcium-channel blocker, as chemotherapy of superficial bladder cancer was carried out at two universities, Okayama and Kagoshima, and their affiliated hospitals. Although little is known about the expression of P-glycoprotein in superficial bladder cancer, it may be a cause of multidrug resistance (MDR). Verapamil was used as an inhibitor of P-glycoprotein. Arm A consisted of Adriamycin given at 50 mg/50 ml saline, and arm B constituted Adriamycin given at 50 mg/40 ml saline plus 5 ampules (10 ml) of injectable verapamil. The drugs were instilled into the bladder for 3 consecutive days in each of 3 consecutive weeks for a total of 9 instillations. No significant difference in antitumor effects was observed between arm A and arm B. Recurrent tumors responded better than did primary tumors to both arm-A and arm-B treatments (P = 0.012). In both treatment arms, significant differences (P = 0.031) in the response rate were found between tumors with diameters of less than 1 cm and those measuring 1-3 cm in diameter. Although the number of evaluable patients was limited, recurrent subjects who had previously received Adriamycin instillations responded in both treatment arms.

Administration, Intravesical↗

PubMed1992

Enhancement of antigen-induced interleukin 4 and IgE production by specific IgG1 in murine lymphocytes.

Conalbumin (CA)-specific type 2 helper T cell (Th2) clone, D10G4.1 (D10) produces IL4 when stimulated with varying doses of TNP-CA in the presence of mitomycin C-treated C3H spleen cells or purified B cells as antigen-presenting cells (APC). The production of IL4 was assessed by bioassay and by expression of IL4 mRNA. IL4 production reached maximum at 100 micrograms/ml of TNP-CA, whereas 1 microgram/ml of the antigen induced less than 10% of the maximum level of IL4. This lower level of IL4 production was augmented to the maximum level when monoclonal anti-TNP IgG1 was added to the culture at 0.5-1 microgram/ml. Anti-TNP IgE, but not anti-TNP IgM, was also effective, though IgE was 1/10 as effective as IgG1. IgG1 with an irrelevant specificity and F(ab')2 of anti-TNP IgG1 did not show augmenting effects. Moreover, the enhancement by anti-TNP IgG1 was completely abolished by monoclonal antibody against murine Fc gamma RII, 2.4G2. These results suggest that a low dose of the antigen complexed with IgG1 is focused on APC by means of Fc gamma RII, processed, and presented efficiently to the Th2 clone. On the other hand, the co-culture of D10 with normal C3H B cells in the presence of 1-100 micrograms/ml TNP-CA resulted in polyclonal IgE production. Anti-TNP IgG1 markedly augmented the lower level of IgE production induced by a suboptimal dose of the antigen (1 microgram/ml). This augmentation was shown to be dependent on endogenous IL4 because the enhancement was abolished by monoclonal anti-IL4 (11B11).

Animals↗

PubMed1992

Differences of L-myc polymorphic patterns of neuroblastoma in patients under 1 year versus older ages: a preliminary report.

The age of the patient at the onset of symptoms or at diagnosis is generally accepted as one of the most important prognostic factors of neuroblastomas (NBs). Children less than 1 year of age have a better survival rate than older patients, but the reason for this is unknown. Forty-eight unselected NB patients were divided into two groups: less than 1 year (younger NB patient) and over 1 year (older NB patient) of age at diagnosis. Two of 12 younger NB patients and 18 of 36 older NB patients had N-myc amplification in their tumors. To elucidate further the possible genetic difference between younger and older NB patients, studies of restriction fragment length polymorphism (RFLP) of the L-myc gene was carried out in these two groups. The L-myc locus showed 2-allele polymorphism, allele L(10 kb) and S (6.6 kb), after digestion with EcoRI. Patients homozygous for L-band have been reported as individuals having less metastatic potential in some cancers. The allele frequencies of L and S in neuroblastomas of younger NB patients were 0.50 and 0.50, while those of older NB patients were 0.35 and 0.65, respectively. Although we did not determine L-myc RFLP in normal tissue of individual patients, we expect that the distribution of allele L and S is partly affected by possible allelic loss involving the L-myc region. However, the L-myc RFLP patterns in younger NB patients were the same as those of normal individuals and significantly differed from those of older NB patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors↗