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Biomedical subjects

Y Ono

Publications and source records attributed to Y Ono.

At least 253 records · Page 14Linked to original sources
PubMed1999

Mutational analysis of the regulatory mechanism of PKN: the regulatory region of PKN contains an arachidonic acid-sensitive autoinhibitory domain.

PKN is a fatty acid- and Rho GTPase-activated protein kinase whose catalytic domain in the carboxyl terminus is homologous to those of protein kinase C (PKC) family members. The amino terminal region of PKN is suggested to function as a regulatory domain, since tryptic cleavage or the binding of Rho GTPase to this region results in protein kinase activation of PKN. The structural basis for the regulation of PKN was investigated by analyzing the activity of a series of deletion/site-directed mutants expressed in insect cells. The amino-terminally truncated form of PKN (residue 455-942) showed low basal activity similar to that of the wild-type enzyme, and was arachidonic acid-dependent. However, further deletion (residue 511-942) resulted in a marked increase in the basal activity and a decrease in the arachidonic acid dependency. A (His)(6)-tagged protein comprising residues 455-511 of PKN (designated His-Ialpha) inhibited the kinase activity of the catalytic fragment of PKN in a concentration-dependent manner in competition with substrate (K(i) = 0.6+/-0.2 microM). His-Ialpha also inhibited the activity of the catalytic fragment of PRK2, an isoform of PKN, but had no inhibitory effect on protein kinase A or protein kinase Cdelta. The IC(50) value obtained in the presence of 40 microM arachidonic acid was two orders of magnitude greater than that in the absence of the modifier. These results indicate that this protein fragment functions as a specific inhibitor of PKN and PRK2, and that arachidonic acid relieves the catalytic activity of wild-type PKN from autoinhibition by residues 455-511 of PKN. Autophosphorylation of wild-type PKN increased the protein kinase activity, however, substitution of Thr64, Ser374, or Thr531 in the regulatory region of PKN with alanine, abolished this effect. Substitution of Thr774 in the activation loop of the catalytic domain of PKN with alanine completely abolished the protein kinase activity. These results suggest that these phosphorylation sites are also important in the regulation of the PKN kinase activity. Potential differences in the mechanism of activation between the catalytic regions of PKN and PRK2 are also discussed.

Animals↗
PubMed1999

Activation of protein kinase B induced by H(2)O(2) and heat shock through distinct mechanisms dependent and independent of phosphatidylinositol 3-kinase.

Protein kinase B (PKB) is a downstream target of phosphatidylinositol (PI) 3-kinase in the signaling pathway of growth factors, and is activated by cellular stress such as H(2)O(2) and heat shock. To study the mechanism of the stress-induced activation of PKB, PI 3-kinase products were measured in stress-stimulated cells. Both PI 3,4-bisphosphate and PI 3,4, 5-trisphosphate increased in H(2)O(2)-treated cells, and the elevation of these phospholipids and activation of PKB were concurrently blocked by wortmannin, a potent inhibitor of PI 3-kinase. In heat-shocked cells, the level of PI 3,4-bisphosphate did not change while that of PI 3,4,5-trisphosphate increased slightly, and an association between PKB molecules was observed. Two active PKB fractions, presumably monomeric and oligomeric forms, were resolved from heat-shocked cells by gel filtration column chromatography. Activation of the former was suppressed by pretreatment with wortmannin, whereas the generation and activation of the latter were not blocked by the PI 3-kinase inhibitor. Only the monomeric form, but not the oligomeric form, was recovered from H(2)O(2)-treated cells, and its activation was prevented by wortmannin. These results indicate that PKB is activated by two distinct mechanisms that are dependent and independent of PI 3-kinase in stress-stimulated cells.

Animals↗
PubMed1999

L-arginine reverses severe nephrosclerosis in aged spontaneously hypertensive rats.

OBJECTIVE: Acute and prolonged effects of L-arginine on systemic and renal hemodynamics and on renal pathological changes were examined in 85-week-old spontaneously hypertensive rats (SHR). RESULTS: After 3 weeks of L-arginine administration (n = 9; 2 g/l in drinking water), mean arterial pressure remained unchanged, although the cardiac index increased (187 +/- 26 versus 263 +/- 15 ml/min per kg; P < 0.05) and total peripheral resistance decreased (1.15 +/- 0.18 versus 0.67 +/- 0.06 AU; P < 0.05); the glomerular filtration rate increased (0.41 +/- 0.07 versus 0.79 +/- 0.07 ml/min; P < 0.01). Control untreated, aged SHR (n = 10) demonstrated severe nephrosclerosis histologically, but those treated with L-arginine demonstrated a markedly reduced glomerular injury score (164 +/- 22 versus 83 +/- 9; P < 0.005), and their urinary protein excretion (39 +/- 5 versus 19 +/- 5 mg/100 g body weight per day; P < 0.05) and serum creatinine concentration (1.4 +/- 0.1 versus 0.9 +/- 0.1 mg/dl; P < 0.05) diminished. Intravenous L-arginine (300 mg/kg body weight) given to untreated SHR reduced mean arterial pressure, increased the cardiac index (+98 versus +1%; P < 0.05) and decreased total peripheral resistance (+56 versus +13%, P < 0.005); however, these variables remained unchanged after 3 weeks of L-arginine treatment. CONCLUSIONS: Three weeks of treatment with L-arginine improved systemic hemodynamics, renal function and renal histologic changes in aged SHR with naturally occurring nephrosclerosis. These data provide an important insight into the pathophysiology of nephrosclerosis in hypertension and with aging, which is seen clinically.

Animals↗
PubMed1999

[Extended-spectrum beta-lactamase (ESBL) produced by Escherichia coli and Klebsiella pneumoniae isolated from Teikyou University Hospital--the first report].

We studied the cefotaxime (CTX)-resistant (MIC > or = 32 micrograms/ml) clinical isolates of E. coli and K. pneumoniae in Teikyo University Hospital from 1990 to 1996. The incidence of CTX-resistant isolates was 0.4% (6/1,282) in E. coli and 0.6% (7/1,044) in K. pneumoniae, in 1990. In 1995, the incidence of CTX-resistance increased to 1.7% (50/2,910) in E. coli (p = 0.0013) and 7.2% (144/1,996) in K. pneumoniae (p < 0.0001). These species have been detected in the stool (86 isolates), urine (59 isolates), sputum (15 isolates), pus (15 isolates), throat (10 isolates) and others (12 isolates) in 1995. MIC50 of ampicillin (ABPC), ABPC with clavlanic acid (CVA) 5 micrograms/ml, piperacillin (PIPC), PIPC with CVA 5 micrograms/ml, ceftazidime, CTX, ceftizoxime, cefpodoxime, cefepime, aztreonam, cefmetazole, latamoxef, and imipenem used against 33 isolates (11 isolates of E. coli, 22 isolates of K. pneumoniae), which were detected in 1996-1997, was > 512 micrograms/ml, 8 micrograms/ml, > 512 micrograms/ml, 8 micrograms/ml, 4 micrograms/ml, > 512 micrograms/ml, 16 micrograms/ml, > 512 micrograms/ml, 256 micrograms/ml, 32 micrograms/ml, 2 micrograms/ml, 0.25 microgram/ml and 0.25 microgram/ml, respectively. This susceptibility pattern were very similar to the Toho-1 type beta-lactamases producing strains.

Cefotaxime↗
PubMed1999

Epstein-barr virus (EBV) nuclear protein 2-induced disruption of EBV latency in the Burkitt's lymphoma cell line Akata: analysis by tetracycline-regulated expression.

The Burkitt's lymphoma (BL) cell line Akata retains the latency I program of Epstein-Barr virus (EBV) gene expression and cross-linking of its surface immunoglobulin G (IgG) by antibodies results in activation of viral replication. When EBV nuclear antigen 2 (EBNA2) was artificially expressed by a constitutive expression vector, the Cp EBNA promoter remained inactive and accordingly the latency III program was not induced. In contrast, expression of LMP2A and activity of the Fp lytic promoter were activated. Consistent with this Fp activity, the rate of spontaneous activation of the EBV replicative cycle was increased significantly, suggesting the possibility that EBNA2 can induce EBV replication. The efficiency of anti-IgG-induced activation of the viral replication was reduced in Akata cells expressing EBNA2. To obtain more direct evidence for EBNA2-induced activation of the EBV replicative cycle, this protein was next expressed by a tetracycline-regulated expression system. EBNA2 was undetectable with low doses (<0.5 microgram/ml) of tetracycline, while its expression was rapidly induced after removal of the antibiotic. This induced expression of EBNA2 was immediately followed by expression of EBV replicative cycle proteins in up to 50% of the cells, as shown by indirect immunofluorescence and immunoblot analysis. These results suggest an unexpected potential of EBNA2 to disrupt EBV latency and to activate viral replication.

Burkitt Lymphoma↗
PubMed1999

Expression and functional characteristics of calpain 3 isoforms generated through tissue-specific transcriptional and posttranscriptional events.

Calpain 3 is a nonlysosomal cysteine protease whose biological functions remain unknown. We previously demonstrated that this protease is altered in limb girdle muscular dystrophy type 2A patients. Preliminary observations suggested that its gene is subjected to alternative splicing. In this paper, we characterize transcriptional and posttranscriptional events leading to alterations involving the NS, IS1, and IS2 regions and/or the calcium binding domains of the mouse calpain 3 gene (capn3). These events can be divided into three groups: (i) splicing of exons that preserve the translation frame, (ii) inclusion of two distinct intronic sequences between exons 16 and 17 that disrupt the frame and would lead, if translated, to a truncated protein lacking domain IV, and (iii) use of an alternative first exon specific to lens tissue. In addition, expression of these isoforms seems to be regulated. Investigation of the proteolytic activities and titin binding abilities of the translation products of some of these isoforms clearly indicated that removal of these different protein segments affects differentially the biochemical properties examined. In particular, removal of exon 6 impaired the autolytic but not fodrinolytic activity and loss of exon 16 led to an increased titin binding and a loss of fodrinolytic activity. These results are likely to impact our understanding of the pathophysiology of calpainopathies and the development of therapeutic strategies.

Alternative Splicing↗
PubMed1999

Apoptosis, coronary arterial remodeling, and myocardial infarction after nitric oxide inhibition in SHR.

This study was designed to investigate the relationship between apoptosis (programmed cell death) and coronary arterial remodeling in spontaneously hypertensive rats (SHR) following prolonged nitric oxide synthesis inhibition. In addition, we evaluated whether the development of coronary arterial smooth muscular cell apoptosis was related to hemodynamics or to vascular hypertrophy. Three groups of 20-week-old male SHR were investigated: controls, and two groups that received two doses of N(G)-nitro-L arginine (L-NAME, 50 mg/L and 80 mg/L) each for 3 weeks. Mean arterial pressure and total peripheral resistance index increased whereas cardiac index diminished with L-NAME. Pathohistological study demonstrated increased pericardiac fibrosis and coronary arterial injury score in the L-NAME group in a dose-dependent manner. The high dose of L-NAME (Group 3) produced myocardial infarction in 78% of the rats. The wall:lumen ratio of epicardial coronary arteries was greater in L-NAME treated SHR (0.23+/-0.02 versus 0.16+/-0.02; P<0.05) and was associated with markedly increased apoptosis (15.3+/-6 versus 1. 9+/-1; P<0.05) without smooth muscle cell proliferation (PCNA positive cells). Apoptosis occurred predominantly in hypertrophic coronary arterial smooth muscular cells; myocardial infarction and ventricular fibrosis were exacerbated by impaired hemodynamics induced by L-NAME. These data suggest that coronary endothelial dysfunction and myocardial ischemic disease induced by L-NAME were responsible for apoptosis of coronary arterial smooth muscle cells, myocardial fibrosis, and infarction, all pathological findings that are consistent with what may be found in clinical hypertensive heart disease.

Animals↗
PubMed1999

Effects of knockout of the protein kinase C beta gene on glucose transport and glucose homeostasis.

The beta-isoform of protein kinase C (PKC) has paradoxically been suggested to be important for both insulin action and insulin resistance as well as for contributing to the pathogenesis of diabetic complications. Presently, we evaluated the effects of knockout of the PKCbeta gene on overall glucose homeostasis and insulin regulation of glucose transport. To evaluate subtle differences in glucose homeostasis in vivo, knockout mice were extensively backcrossed in C57BL/6 mice to diminish genetic differences other than the absence of the PKCbeta gene. PKCbeta-/- knockout offspring obtained through this backcrossing had 10% lower blood glucose levels than those observed in PKCbeta+/+ wild-type offspring in both the fasting state and 30 min after i.p. injection of glucose despite having similar or slightly lower serum insulin levels. Also, compared with commercially obtained C57BL/6-129/SV hybrid control mice, serum glucose levels were similar, and serum insulin levels were similar or slightly lower, in C57BL/6-129/SV hybrid PKCbeta knockout mice in fasting and fed states and after i.p. glucose administration. In keeping with a tendency for slightly lower serum glucose and/or insulin levels in PKCbeta knockout mice, insulin-stimulated 2-deoxyglucose (2-DOG) uptake was enhanced by 50-100% in isolated adipocytes; basal and insulin-stimulated epitope-tagged GLUT4 translocations in adipocytes were increased by 41% and 27%, respectively; and basal 2-DOG uptake was mildly increased by 20-25% in soleus muscles incubated in vitro. The reason for increased 2-DOG uptake and/or GLUT4 translocation in these tissues was uncertain, as there were no significant alterations in phosphatidylinositol 3-kinase activity or activation or in levels of GLUT1 or GLUT4 glucose transporters or other PKC isoforms. On the other hand, increases in 2-DOG uptake may have been partly caused by the loss of PKCbeta1, rather than PKCbeta2, as transient expression of PKCbeta1 selectively inhibited insulin-stimulated translocation of epitope-tagged GLUT4 in adipocytes prepared from PKCbeta knockout mice. Our findings suggest that 1) PKCbeta is not required for insulin-stimulated glucose transport; 2) overall glucose homeostasis in vivo is mildly enhanced by knockout of the PKCbeta gene; 3) glucose transport is increased in some tissues in PKCbeta knockout mice; and 4) increased glucose transport may be partly due to loss of PKCbeta1, which negatively modulates insulin-stimulated GLUT4 translocation.

Adipocytes↗
PubMed1999

Expression of adrenomedullin in feto-placental circulation of human normotensive pregnant women and pregnancy-induced hypertensive women.

The adrenomedullin (AM) peptide and the expression of AM messenger RNA (mRNA) from feto-maternal tissues of 22 normotensive pregnant women and from 7 women with pregnancy-induced hypertension (PIH) during third-trimester were examined to clarify the pathophysiological features of PIH. Samples of the placenta, uterine muscle, umbilical artery, and fetal membranes were obtained from each patients under informed consent. The AM peptide was measured by a radioimmunoassay and the AM mRNA level was analyzed by Northern hybridization. The total immunoreactive AM (fmol/mg wet tissue) was significantly increased in the fetal membranes (1.95+/-0.20 vs. 3.03+/-0.44) and the umbilical artery (0.11+/-0.01 vs. 0.15+/-0.02) of the patients with PIH. On the other hand, the AM mRNA level was higher in the umbilical artery, and lower in the fetal membranes in the patients with PIH. The present results thus suggest that the changes in the expression of AM in fetal membrane and umbilical artery in PIH may play an important role in the fetal and maternal circulation.

Adrenomedullin↗
PubMed1999

Simultaneous determination of baicalin, wogonoside, baicalein, wogonin, berberine, coptisine, palmatine, jateorrhizine and glycyrrhizin in Kampo medicines by ion-pair high-performance liquid chromatography.

An ion-pair high-performance liquid chromatographic method for the simultaneous determination of four flavonoids, namely baicalin, wogonoside, baicalein and wogonin, and four berberine-type alkaloids, namely berberine, coptisine, palmatine and jateorrhizine, and glycyrrhizin in Kampo medicines is described. The analysis can be accomplished within 30 min with a Wakosil-II 5C18 HG column by linear gradient elution using a mobile phase containing aqueous phosphoric acid, sodium dodecyl sulfate and acetonitrile at a flow-rate of 1.0 ml x min(-1), a thermostatic oven at 45 degrees C, and detection at 265 nm. The method was applied to quantifying these components in three Kampo decoctions: Oren-gedoku-to, San'o-shashin-to and Hange-shashin-to. The decoctions were diluted with 65% methanol at the final stage because a large quantity of precipitate, mainly from baicalin and berberine, was formed. The within-day relative standard deviations were less than 2.02% (n=10). The recoveries of these compounds were 90.3-102%. The detection limits of these compounds were 0.02-1.96 microM per injection (5 microl).

Alkaloids↗
PubMed1999

Soft-tissue interposition after closed reduction in developmental dysplasia of the hip. The long-term effect on acetabular development and avascular necrosis.

We reviewed 98 children (133 hips) with developmental dysplasia of the hip who underwent arthrography immediately after closed reduction by overhead traction. We followed the patients to skeletal maturity to investigate whether soft-tissue interposition influences acetabular development and avascular necrosis over the long term. The shape of the limbus and the thickness of the soft-tissue interposition at the acetabular floor, as shown on arthrograms at the time of reduction, were not directly related to the final radiological results or to the incidence of avascular necrosis. Even if marked soft-tissue interposition was found on the initial arthrogram, spontaneous disappearance was noted in 71% up to the age of five years. The final radiological results showed no difference between those in which the interposition disappeared and those with none at the time of closed reduction. However, the requirement for secondary surgery at the age of five years was significantly higher in those with more than 3.5 mm of soft-tissue interposition. In the no-disappearance group (group C) further operation was necessary in 100% and the results were significantly worse at maturity according to Severin's classification. We suggest that the indications for open reduction should not be based solely on the arthrographic findings at the time of closed reduction.

Adolescent↗
PubMed1999

Immunohistochemical characteristics of estrogen receptor alpha positive cells in glandular epithelium of the rat seminal vesicle.

Epithelial cells of the rat seminal vesicle stained positively for nuclear estrogen receptor alpha (ER alpha). We studied these cells using immunohistochemical means. We demonstrated in a previous study that some glandular epithelial cells of the seminal vesicles of immature castrated rats treated with estrogen for 1-2 weeks had multilayer features. The present study shows that these glandular epithelial cells are nuclear ER and basal cell-specific cytokeratin (34betaE12) positive. These findings suggested characteristics of basal cells. Moreover, we demonstrated that these cells express transforming growth factor beta1 (TGFbeta1) as a result of castration and estrogen treatment. Our findings indicate that glandular epithelial cells with multilayer features, which stained positively for nuclear ER alpha have basal cell features and may play an important role in the expression of TGFbeta1 through an epithelial-stromal interaction.

Animals↗
PubMed1999

Environmental and possible genetic contributions to character dimensions of personality.

The relationship between dimensions of personality characteristics and the perceived rearing attitude of parents in the Japanese population were investigated. The scores on a measure of perceived parental attitude of 153 normal female students, measured on the Parker Parental Bonding Instrument, were correlated with personality features from the Japanese version of the Cloninger Temperament and Character Inventory. Self-directedness, especially the subclasses of Responsibility vs Blaming and Congruent Second Nature vs Incongruent Habits, was significantly related to high scores on Maternal Care and low scores on Maternal Overprotection. The subscale of Self-acceptance vs Self-striving correlated only with low scores on Maternal Overprotection. Paternal Care was only related to the total scale scores on Self-directedness. Results suggest that some personality traits may be related to the perceived attitudes of parents, especially of the mother, during childhood.

Adolescent↗
PubMed1999

[Clinical study of radical retropubic prostatectomy for prostate cancer].

PURPOSE: To evaluate the results of radical retropubic prostatectomy in patients treated at a single institution. MATERIALS AND METHODS: Between April 1985 and July 1997, 76 patients with prostate cancer underwent radical retropubic prostatectomy, including 73 receiving pelvic lymphadenectomy. The median age and follow-up time were 68 years old and 44 months, respectively. The pathological stage was pT0 in 6 patients, pT2 in 29, pT3 in 39, pT4 in 2, and pN+ in 22. RESULTS: The surgical margin was positive in 10% of the pT2 patients and 61% of the pT3 patients. Twelve patients had recurrence. Recurrence was shown by biological failure in 4 patients and clinical failure in 8. The disease-free 5-year survival rates (Kaplan-Meier) were 100% in pT0 patients, 87% in pT2, 72% in pT3, 50% in pT4, 77% in pN-, 75% in pN+, 73% for a positive surgical-margin, and 83% for a negative surgical-margin. There were no statistical differences between any of these factors. However, the disease-free survival rate in pT3 patients with poorly differentiated adenocarcinoma (PDA) who received postoperative radiotherapy combined with hormonal therapy was significantly superior to that in patients with the same characteristics who received hormonal therapy (100% vs 27%; p = 0.011). The cause-specific 5-year survival rates were 100% in pT0, 100% in pT2, 92% in pT3, 50% in pT4, 94% in pN-, 93% in pN+, 93% for a positive surgical-margin, 98% for a negative surgical-margin, 100% in the aforementioned pT3 patients with PDA and postoperative radiotherapy combined with hormonal therapy and 86% in pT3 patients with PDA and postoperative hormonal therapy. There were no statistical differences between any of these factors. CONCLUSIONS: Our results suggest that radical prostatectomy is available for both organ-confined and non organ-confined advanced prostate cancer. Postoperative radiotherapy combined with hormonal therapy is especially useful for patients in pT3 with PDA.

Adenocarcinoma↗