Evaluation of DNA probe test for rapid diagnosis of Mycoplasma pneumoniae infections.
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Adolescent↗
Biomedical subjects
N Miwa
Publications and source records attributed to N Miwa.
[Neonatal brain-derived carcinostatic factor (NBCF)--cytocidal action to neuroblastoma cells and molecular characters as a glycoprotein].
Neuronal cells as many as 40-70% in several regions of the brain are destined to suffer from programmed cell death at the terminal stage of the ontogenesis. At the stage the mouse brain secretes some species of cell growth-inhibitory factors including a novel glycoprotein (62KD, pI9.1) which inhibits DNA synthesis and proliferation of tumor cells preferentially over those of normal cells. The factor was designated as NBCF (neonatal brain-derived carcinostatic factor) because it is not produced at stages other than neonatal and prenatal stages or from other organs. The activity is exhibited through a protenic body of NBCF as an active principle, whereas the glycan moiety contributes assumedly to secretion and/or activity stabilization. NBCF is cytocidal more markedly to neuroblastoma cells than a diversity of other tumor cells; the susceptibility to NBCF becomes marked when neuroblastoma cells are undifferentiated.
Animals↗
Late-phase accumulation of inositol phosphates stimulated by prostaglandins D2 and F2 alpha in neuroblastoma x glioma hybrid NG108-15 cells.
The accumulation of inositol phosphates (IPs) in response to prostaglandins (PGs) was studied in NG108-15 cells preincubated with myo-[3H]inositol. As a positive control, bradykinin caused accumulation of IPs transiently at an early phase (within 1 min) and continuously during a late phase (15-60 min) of incubation in the cells. PGD2 and PGF2 alpha did not significantly cause the accumulation of IPs at an early phase but significantly stimulated inositol bisphosphate (IP2) and inositol monophosphate (IP) formation at late phase of incubation. The maximum stimulation was obtained at greater than 10(-7) M concentrations of these PGs, the levels being three-and twofold for IP2 and IP1, respectively. 9 alpha, 11 beta-PGF2 has a slight effect but PGE2 and the metabolites of PGD2 and PGF2 alpha have no effect up to 10(-6)M. The effects of PGD2 and PGF2 alpha were not additive, but the effect of each PG was additive to that of bradykinin at a late phase of incubation. Inositol 1-monophosphate was mainly identified in the stimulation by 10(-5) M PGD2 and 10(-5) M PGF2 alpha, whereas both inositol 1-monophosphate and inositol 4-monophosphate were produced in the stimulation by 10(5) M bradykinin. Depletion of extracellular Ca2+ diminished the stimulatory effect of PGD2 and PGF2 alpha and late-phase effect of bradykinin, but simple Ca2+ influx into the cells by high K+, ionomycin, or A23187 failed to cause such late-phase effects. These results suggest that PGD2 and PGF2 alpha specifically stimulate hydrolysis of inositol phospholipids.
Animals↗
[A waterborne outbreak of typhoid fever associated with a small drinking water supply system in Fuji city].
Fifteen cases of typhoid fever occurred in "HARADA" district Fuji City in Shizuoka Prefecture during the period of 1983 to 1985. Epidemiological and bacteriological investigations were carried out on the samples from a small water supply system, and drains of apartment houses in 1985. Water from a small river in the neighboring water supply system, and faecal specimens from people living in close proximity were investigated. The results obtained were as follows: 1) Salmonella typhi were isolated from 3/3 patients (1 pupil and 2 employees) in 1983, 3/5 patients (3 employees) in 1984, and from 4/7 patients (1 kindergartner, 1 pupil and 2 junior high school boys) in 1985. Phage type of these isolates were the same type "D1". 2) In the bacteriological survey on environment, S. typhi (phage type D1) were isolated from water of the water source of "HARADA" water supply in Fuji City with membrane filter methods, and from the sand source "HARADA" water supply with enrichment culture methods. Also, with an examination that tested 3,670 inhabitants, it was found that a carrier (phage type D1) was living near the "HARADA" water supply. 3) In the three selective media for the isolation of S. typhi, Bismuth sulfite agar (Difco) was the most sensitive, and detected 13 samples (61.9%) out of 21, from water source and sewage.
Adolescent↗
Effect of acylated derivatives of ascorbate on ornithine decarboxylase induction in Ehrlich ascites tumor cells.
We studied the effects of 6-0-acylated derivatives of ascorbate on ornithine decarboxylase (ODC, E.C.4.1.1.17) activity and the ODC-mRNA level in Ehrlich ascites cells. These derivatives are known to inhibit cell growth. When cells grown to the stationary phase were diluted and stimulated with fresh medium containing 10% dialyzed fetal calf serum, the ODC activity increased rapidly and transiently with a peak at 4 hr. Ascorbate or 2,6-dipalmitoyl ascorbate had little or no effect on the induction of ODC, but 6-0-palmitoyl ascorbate (6P) suppressed the enzyme activity in a time- and dose-dependent way. 6-0-Stearoyl ascorbate had a similar effect. The amount of ODC-mRNA formed after growth stimulation was not affected by 6P. These results indicate that the treatment of Ehrlich ascites tumor cells with 6P caused dose-dependent suppression of ODC induction post-transcriptionally.
Acetylation↗
Altered production of the active oxygen species is involved in enhanced cytotoxic action of acylated derivatives of ascorbate to tumor cells.
Our previous study shows that 6-O-acyl derivatives of L-ascorbic acid inhibits more markedly cell growth of mouse Ehrlich carcinoma than ascorbic acid. The present study shows that 6-O-palmitoyl ascorbic acid but not ascorbic acid prolongs the lifespan of mice into which tumors such as Meth A fibrosarcoma, MM46 mammary carcinoma, Ehrlich carcinoma and sarcoma 180 are implanted. The potentiated cytotoxicity of 6-O-palmitoyl ascorbic acid is not due to an increase in duration time of the cytotoxic action, because 6-O-palmitoyl ascorbic acid is gradually inactivated during contact with tumor cells and exhibits a similar action time curve to that of ascorbic acid as shown by clonal growth assay. Cytotoxicity of 6-O-palmitoyl ascorbic acid is markedly diminished by combined addition of catalase and superoxide dismutase (SOD), as shown by dye exclusion assay, whereas the cytotoxicity was slightly reduced by either enzyme alone but not by the specifically inactivated or heat-denatured enzymes. In contrast, cytotoxicity of ascorbic acid is abolished by catalyse but not SOD. Autooxidation of 6-O-palmitoyl ascorbic acid was not inhibited by catalase plus SOD. The results indicate that cytotoxicity of 6-O-palmitoyl ascorbic acid is attributed at least partly to both hydrogen peroxide (H2O2) and superoxide (O2-.) generated at the early stage. Cytotoxicity of 6-O-palmitoyl ascorbic acid is also appreciably attenuated by singlet oxygen (1O2) scavengers such as hydroquinone, 1,4-diazobicyclo-2,2,2-octane or sodium azide, but not by hydroxyl radical scavengers including butylated hydroxytoluene, D-mannitol, benzoic acid and ethanol. Thus, in contrast to cytotoxicity of ascorbic acid mediated entirely by H2O2 initially generated, acylated ascorbic acid produces a diversity of active oxygen species including H2O2, O2-. and other species secondarily generated via disproportion, which may be additively involved in the enhanced cytotoxic action.
Animals↗
Prostaglandin induces Ca2+ influx and cyclic GMP formation in mouse neuroblastoma X rat glioma hybrid NG108-15 cells in culture.
Various prostaglandins (PGs) (10 nM-30 microM) were added to NG108-15 cells in culture, and changes in the levels of intracellular cyclic GMP and Ca2+ were investigated. Exposure of the cells to PGF2 alpha, PGD2, and PGE2 (10 microM) transiently increased the cyclic GMP content 7.5-, 3.9-, and 3.1-fold, respectively. Furthermore, the increased levels of cyclic GMP correlated well with the rise in cytosolic free Ca2+ concentrations induced by the PGs. Other PGs (10 microM), including metabolites and synthetic analogs, which had no effect on intracellular Ca2+, failed to increase the cyclic GMP content in the cells. When extracellular Ca2+ was depleted from the culture medium, the PG-induced increase in cyclic GMP level was almost completely abolished. In addition, treatment of the cells with quin 2 tetraacetoxymethyl ester dose-dependently inhibited the PG-induced cyclic GMP formation. The increase in cyclic GMP content caused by treatment of the cells with a high K+ level (50 mM) was completely blocked by voltage-dependent Ca2+ entry blockers, such as verapamil (10 microM), nifedipine (1 microM), and diltiazem (100 microM); however, the PG (10 microM)-induced increase in cyclic GMP content was not affected by such Ca2+ entry blockers. These findings indicate that PG-induced cyclic GMP formation may require the rise in intracellular Ca2+ level and that the voltage-dependent Ca2+ channels may not be involved in the PG-induced rise in Ca2+ content.
Aminoquinolines↗
Photosensitivity induced by griseofulvin.
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Aged↗
Production of tumor growth-inhibiting protein by the neonatal mouse brain: dependence on intracellular glutamine metabolism.
We previously reported that the mouse brain at the neonatal stage but not at the adult stage secreted a carcinostatic factor of 62,000 Da, termed NBCF, which inhibited clonal growth and DNA synthesis of malignant cells preferentially over those of normal cells. In the present study, NBCF production by the neonatal mouse brain in culture was investigated. Addition of L-glutamine to the culture medium markedly promoted NBCF production in a concentration-dependent manner. The production seemed to be specific to glutamine, since no promotive effect was exerted by L-glutamic acid, its analogue DL-alpha-aminoadipic acid, L-aspargine, or L-aspartic acid or by other amino acids or vitamins. NBCF production was markedly reduced in culture medium either devoid of L-glutamine or containing 6-diazo-5-oxo-L-norleucine, a glutamine antagonist, or L-methionine sulfoximine, an inhibitor of glutamine synthetase. Thus NBCF production was promoted by extracellular supply, intracellular synthesis, and utilization of L-glutamine but was not affected by its deamidated form or homologue amino acids. On the other hand, NBCF production was completely repressed by addition of cycloheximide to the culture medium. The repressive effect was also exerted by actinomycin D although not completely, whereas cytosine beta-D-arabinofuranoside did not repress NBCF production. These results indicated that NBCF production by cultivation was independent of DNA replication but dependent mostly on a transcription stage and its following stages and partly on a translation stage from the preexisting mRNA to the protein.
Animals↗
Potentiated susceptibility of ascites tumor to acyl derivatives of ascorbate caused by balanced hydrophobicity in the molecule.
Orders of susceptibility of Ehrlich ascites tumor to L-ascorbic acid (Asc), its 6-stearoyl (6S), 6-palmitoyl (6P) and 2,6-dipalmitoyl (DP) derivatives were assessed in vitro and in vivo: 6P (a 50% growth inhibitory concentration (IC50) for cultured cells, 12 microM; an increased life-span of treated mice, 283%) greater than 6S (61 microM; 240%) much greater than Asc (430 microM; 122%) greater than or equal to DP (greater than 200 microM; 89%), indicating that the enhanced susceptibility was due to acyl moiety substituted at C6-hydroxyl group of Asc, but was retracted by further substitution at C2-hydroxyl group. Equimolar mixture of Asc and palmitic acid, stearic acid or their methyl esters was much less cytotoxic than 6P or 6S. Thus the enhanced susceptibility was not primarily due to an additive cytotoxic effect of ascorbyl and acyl moieties, but to a balanced hydrophobicity introduced into the molecule by a poorly cytotoxic acyl moiety.
Animals↗
Binding of [3H]-nitrendipine and [3H]-diltiazem to rat myocardial sarcolemma.
The binding properties of two major and chemically distinct calcium antagonists, [3H]-nitrendipine and [3H]-diltiazem, were investigated in highly purified rat cardiac sarcolemma. In the case of [3H]-nitrendipine, there appeared a single set of high affinity binding sites with a B max of approximately 0.9 pmol X mg-1 protein and a KD of approximately 0.15 nmol/l. Because of the extremely high value obtained for KD (29 mumol/l), the specificity of [3H]-diltiazem binding seemed questionable. The specific binding of 0.1 nmol/l [3H]-nitrendipine to cardiac sarcolemma was inhibited by nitrendipine and nifedipine (1 mumol/l), only partly inhibited by verapamil (1 mumol/l), and was enhanced by diltiazem (0.1-10 mumol/l). The stimulation of [3H]-nitrendipine binding by diltiazem was associated with an increase in the number of binding sites, Bmax, but with no change in the KD or the Hill coefficient. An enantiomer of diltiazem (1-cis) neither stimulated nor inhibited the [3H]-nitrendipine binding. These results strongly suggest that major prototype calcium antagonists have distinct and variously interacting sites of action in the rat cardiac sarcolemma.
Animals↗
Adriamycin and altered membrane functions in rat hearts.
To evaluate the potential roles of alterations of membrane functions and resulting calcium overload in the pathogenesis of acute adriamycin cardiotoxicity, we observed sarcolemmal, sarcoplasmic reticular and mitochondrial functions in isolated hearts perfused by the Langendorff technique and exposed to adriamycin. Myocardial tissue calcium content was increased to 120 and 130% of the control level after 30 and 60 min perfusion with adriamycin (50 micrograms/ml), respectively. Sarcolemmal ouabain-sensitive Na+,K+-ATPase activity was decreased by 46% after 30 min perfusion, compared with the control. Mitochondrial calcium uptake was also depressed but sarcoplasmic reticular calcium uptake and binding remained unaltered. Mitochondrial respiratory activity was depressed after 60 min perfusion, when glutamate was used as a substrate, thereby indicating that adriamycin had an inhibitory effect on the NADH-dehydrogenase system. Thus, among membrane functions directly or indirectly regulating calcium movement, the inhibition of enzyme systems susceptible to lipid peroxidation may play important roles in calcium overload induced by adriamycin.
Animals↗
Specific binding of [3H] mepyramine to histamine H1-receptors in the sarcolemma from porcine aorta and coronary artery.
We studied the subcellular distribution and the properties of [3H] mepyramine binding in the porcine vascular smooth muscle. A close correlation was observed between the specific binding activity of [3H] mepyramine and the extent of the enrichment of sarcolemmal marker enzyme in 4 subfractions obtained by sucrose density gradient centrifugation of the aortic microsome. In the binding isotherm in the sarcolemmal fractions from the aorta and coronary artery, there was no difference in the Kd value, but the Bmax of the coronary artery was significantly lower than that of the aorta. Thus, there is a single type of high affinity mepyramine binding site in the porcine vascular smooth muscle sarcolemma and the number of H1-receptors of the coronary artery may be smaller than that of the aorta.
Aminopyridines↗
Inhibition of tumor cell growth by protein factors derived from the developing mouse brain.
It was recently reported (Endoh et al. 1981, Exp Cell Biol 49:272-277) that conditioned medium of neonatal mouse brain (CM-NB) inhibited the growth of mouse neuroblastoma cells. In this work we fractionated CM-NB by size exclusion high performance liquid chromatography, and separated two active principles (28,000 and 62,000 daltons) Each or a combination of the 28,000 and 62,000 dalton fractions showed a differential inhibitory effect on DNA synthesis or clonal growth of the three human lung cell lines: the normal diploid fibroblast WI38 cells were less susceptible than their simian virus 40-transformed VA13 cells and carcinoma A549 cells. This preferential growth-inhibition of malignant cells was also observed for rat fibroblast 3Y1 and its simian virus 40-transformed W3Y cells, and for two other normal and five other malignant cell lines. The growth-inhibitory activity of CM-NB or the 28,000 and 62,000 dalton fractions was lost by pronase, trypsin, tetrahydrofuran, acetonitrile, or dithiothreitol in the presence of guanidine, and also labile to heat, vigorous agitation, or freeze-thawing. The activity was also found in the conditioned medium of prenatal mouse brain, but not in either the conditioned medium of the adult brain and of the secondary culture of the neonatal brain, or in the homogenate and rinsing fluid of the neonatal brain. Thus the mouse brain at the terminal stage of ontogenesis liberates proteinaceous factors, which exhibit a preferential growth-inhibition of tumor or transformed cells and act on malignant cells of human and rodent origin.
Animals↗
Accumulation of autolysosomes after receptor-mediated introduction of Ep459-asialofetuin conjugate into lysosomes in rat hepatocytes.
Administration of Ep459-asialofetuin conjugate (Ep459-AF) and pepstatin-asialofetuin conjugate (Ps-AF) to rats effectively inhibited lysosomal BANA hydrolase and cathepsin D in the liver, respectively, at a very low dose. Ep459-AF treatment also led to an accumulation of autolysosomes in rat liver. There was a close correlation between the accumulation of autolysosomes and the inhibition of BANA hydrolase activity. However, as opposed to the inhibition of thiol proteases, the inhibition of cathespin D did not cause accumulation of autolysosomes in the rat liver. These results suggest that autophagy in rat hepatocytes is a common occurrence under normal physiological conditions and that thiol proteases are digestive enzymes essential for the autolysomes.
Aminopeptidases↗
Conformational changes in human urokinase induced by a specific reduction of disulfide bond in Cys-194-Cys-222 associated with exhibition of enzymatic activity.
The disulfide bond in Cys-194-Cys-222 of the isolated B chain (UK X B) of human urinary urokinase was selectively carboxamidomethylated (R X CAM-UK X B) after specific reduction with dithiothreitol in the presence of the competitive inhibitor N-alpha-benzoyl-L-argininamide (BzArgNH2) and 0.3 M guanidine with inactivation. There were no differences between UK X B and R X CAM-UK X B in CD spectra of 200-250 nm and infrared spectra in the amide I and II regions, whereas two out of five negative CD bands of 250-310 nm were slightly reduced in R X CAM-UK X B. R X CAM-UK X B was more susceptible than UK X B to guanidine denaturation, H-2H exchange in peptide protons and tyrosine-residue ionization. These results indicate that Cys-194-Cys-222 contributes to conformational stabilization against perturbants, but not to static conformation in the resting state, except for a slight disruption in the amino-acid side-chains. BzArgNH2 and 0.3 M guanidine, corresponding to the threshold point toward denaturing transition, enhanced more markedly all the five CD bands of 250-310 nm in UK X B than in R X CAM-UK X B, but did not alter CD spectra of 200-250 nm. The deletion of either BzArgNH2 or guanidine did not induce the inactivation or the cleavage of Cys-194-Cys-222 upon the reduction of UK X B, nor did it induce a marked CD enhancement in the near-ultraviolet upon the addition to UK X B. The results suggest that the BzArgNH2-UK X B interaction enhances the solvent accessibility to the disulfide bond with aid of critical guanidine-denaturation.
Arginine↗
A specific disulfide bond associated with the activity of human urokinase. Its topological identification and reductive cleavage followed by kinetic changes in enzymatic reaction and affinity labeling.
A single SH group in the B chain (33 kDa), generated by the specific reduction of the single interchain SS bond of human urinary urokinase, was alkylated (UK X B) with iodoacetamide to prevent a spontaneous SH-SS interchange. An SS bond in UK X B was exclusively alkylated with iodoacetamide (R X CAM-UK X B) after reduction with dithiothreitol in 0.3 M guanidine X HCl in the presence of the competitive inhibitor N alpha-benzoyl-L- argininamide with concomitant loss of 65-68% of the esterolytic activity towards N-acetyl-glycyl-L-lysine methyl ester. This specific SS bond was located at Cys194 - Cys222 whose SS loop contained the active-site Ser198 , as determined by amino acid analyses and identification of the N and C termini of the tryptic digest. Transformation of UK X B into R X CAM-UK X B induced no shift of the optimal pH in the bell-shaped pH/activity profile; pH values for 50% activity were similar (pH 9.7) for 10-min alkalization of the enzyme but different between UK X B (pH 9.4) and R X CAM-UK X B (pH 8.8) for 18-h alkalization. An unaltered Km value and a decline by 64% in kcat in the esterolytic activity indicate that the pretransition Michaelis complex is formed without degeneration of the primary substrate-binding site, but the catalytic pathway thereafter has deteriorated. In affinity labeling with dansyl chloride or N alpha-tosyl-L-lysine chloromethylketone, which interrupted the catalysis at the latest at a stage involving the abortive acyl intermediate, the second-order rate constant for UK X B was lowered to 28% or 35% for R X CAM-UK X B, respectively, but the labeling yields were similar. The results indicate that indispensable structural elements, such as the catalytic triad and oxyanion hole, are maintained but a local conformation, which is necessary for efficient transition to the acyl intermediate and/or for resistance against alkaline inactivation, is destabilized with Cys194 - Cys222 scission.
Affinity Labels↗
Natural infection in non-human primates with adult T-cell leukemia virus or a closely related agent.
A total of 703 sera from 10 species of monkeys were examined for the presence of antibodies to adult T-cell leukemia (ATL)-associated antigens (ATLA). ATLA represent core protein(s) of ATL virus (ATLV) and ATLV-determined polypeptides. Anti-ATLA antibodies were found in all seven macaque species tested but not in three non-macaque species. The frequencies of seropositive macaques ranged from 10 to 50%. In three macaque species (Japanese monkeys, rhesus monkeys, and crab-eating monkeys) in which sufficient numbers of animals were studied, more females than males were anti-ATLA positive and the antibody-positive rate increased with age. In Japanese monkeys, over 70% were seropositive after the age of 10 years. A family study of Japanese monkeys suggested maternal transmission of ATLV or a closely related agent. These seroepidemiological features are consistent with those of Japanese in the ATL-endemic area. Three of 10 rhesus monkeys were found to be anti-ATLA-positive 13 days after arrival in Japan, suggesting that they had been infected with ATLV in Indonesia where they were captured. Thus, ATLV appears to be prevalent among several macaque species as well as in man, and its pathogenetic role in these non-human primates should be explored in relation to ATL in man.
Animals↗