Designed prostaglandins with neurotrophic activities.
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Animals↗
Biomedical subjects
T Satoh
Publications and source records attributed to T Satoh.
Studies on the interactions between drugs and estrogen: analytical method for prediction system of gynecomastia induced by drugs on the inhibitory metabolism of estradiol using Escherichia coli coexpressing human CYP3A4 with human NADPH-cytochrome P450 reductase.
To establish a prediction system for drug-induced gynecomastia in clinical fields, a model reaction system was developed to explain numerically this side effect. The principle is based on the assumption that 50% inhibition concentration (IC(50)) of drugs on the in vitro metabolism of estradiol (E2) to its major product 2-hydroxyestradiol (2-OH-E2) can be regarded as the index for achieving this purpose. By using human cytochrome P450s coexpressed with human NADPH-cytochrome P450 reductase in Escherichia coli as the enzyme, the reaction was examined. Among the nine enzymes (CYP1A1, 1A2, 2A6, 2C8, 2C9, 2C19, 2D6, 2E1, and 3A4) tested, CYP3A4 having a V(max)/K(m) (ml/min/nmol P450) value of 0.32 for production of 2-OH-E2 was shown to be the most suitable enzyme as the reagent. The inhibitory effects of ketoconazole, cyclosporin A, and cimetidine toward the 2-hydroxylation of E2 catalyzed by CYP3A4 were obtained, and their IC(50) values were 7 nM, 64 nM, and 290 microM, respectively. The present results suggest that IC(50) values thus obtained can be substituted as the prediction index for gynecomastia induced by drugs, considering the patients' individual information.
Catechols↗
CYP3A is responsible for N-dealkylation of haloperidol and bromperidol and oxidation of their reduced forms by human liver microsomes.
We studied the biotransformation of haloperidol, bromperidol and their reduced forms by human liver microsomes. Nifedipine oxidation (CYP3A) activity correlated significantly with N-dealkylation rates of haloperidol and bromperidol and oxidation rates of their reduced forms, while neither ethoxyresorufin O-deethylation (CYP1A2) activity nor dextromethorphan O-deethylation (CYP2D6) activity did. In chemical and immunoinhibition studies, only troleandomycin and anti-CYP3A4 serum inhibited both formation rates of 4-fluorobenzoylpropionic acid, a metabolite of haloperidol and bromperidol, and back oxidation rates. Among 10 recombinant isoforms examined, only CYP3A4 showed catalytic activity. The Vmax and Km values of N-dealkylation of bromperidol and reoxidation of reduced bromperidol were similar to those of haloperidol and reduced haloperidol, respectively. The present study indicates that CYP3A plays a major role in N-dealkylation of and oxidation back to bromperidol as well as haloperidol and suggests that modification of in vivo CYP3A activity by inhibition or induction may affect the pharmacokinetics and therapeutic effects of haloperidol and bromperidol.
Aryl Hydrocarbon Hydroxylases↗
Frequent hypermethylation of CpG islands and loss of expression of the 14-3-3 sigma gene in human hepatocellular carcinoma.
The 14-3-3 sigma gene has been implicated in G2/M cell cycle arrest by p53. Frequent inactivation of the 14-3-3 sigma gene by hypermethylation of CpG islands has recently been reported in human breast carcinoma. The aim of this study was to examine the methylation status of CpG islands of the 14-3-3 sigma gene in hepatocellular carcinoma (HCC). The methylation status of the 14-3-3 sigma gene was evaluated in four normal liver tissues and 19 paired specimens of carcinoma and adjacent non-tumorous liver tissues using bisulfite-single strand conformation polymorphism (bisulfite-SSCP), a combination of sodium bisulfite modification and fluorescence-based polymerase chain reaction (PCR)-SSCP. The 14-3-3 sigma protein expression was examined by immunohistochemical staining. Hypermethylation of CpG islands of the 14-3-3 sigma gene was detected in 89% (17/19) of the HCC tissues but not in any of the four normal liver tissues. All of the 14 methylation-positive HCC samples analysed by immunohistochemistry showed loss of 14-3-3 sigma expression, while both of the methylation-negative HCC samples retained the expression, and a significant correlation was found between methylation and loss of expression. Lower levels of methylation were detected in adjacent non-tumorous liver tissues (6/16 in cirrhotic tissues and 1/3 in chronic hepatitis tissues), but the 14-3-3 sigma expression was retained in all of these tissues. In a methylation-positive HCC cell line, HLE, 5-aza-2'-deoxycytidine (5-aza-dC)-induced demethylation of CpG islands led to reactivation of gene expression, indicating that hypermethylation plays a causal role in inactivation of the 14-3-3 sigma gene in HCC. Hypermethylation and the resulting loss of expression of the 14-3-3 sigma gene corresponds to one of the most common abnormalities reported to date in HCC, suggesting their crucial role in the development and/or progression of HCC.
14-3-3 Proteins↗
Regulation of a novel human phospholipase C, PLCepsilon, through membrane targeting by Ras.
Phosphoinositide-specific phospholipase C (PI-PLC) plays a pivotal role in regulation of intracellular signal transduction from various receptor molecules. More than 10 members of human PI-PLC isoforms have been identified and classified into three classes beta, gamma, and delta, which are regulated by distinct mechanisms. Here we report identification of a novel class of human PI-PLC, named PLCepsilon, which is characterized by the presence of a Ras-associating domain at its C terminus and a CDC25-like domain at its N terminus. The Ras-associating domain of PLCepsilon specifically binds to the GTP-bound forms of Ha-Ras and Rap1A. The dissociation constant for Ha-Ras is estimated to be approximately 40 nm, comparable with those of other Ras effectors. Co-expression of an activated Ha-Ras mutant with PLCepsilon induces its translocation from the cytosol to the plasma membrane. Upon stimulation with epidermal growth factor, similar translocation of ectopically expressed PLCepsilon is observed, which is inhibited by co-expression of dominant-negative Ha-Ras. Furthermore, using a liposome-based reconstitution assay, it is shown that the phosphatidylinositol 4,5-bisphosphate-hydrolyzing activity of PLCepsilon is stimulated in vitro by Ha-Ras in a GTP-dependent manner. These results indicate that Ras directly regulates phosphoinositide breakdown through membrane targeting of PLCepsilon.
Amino Acid Sequence↗
Designed cyclopentenone prostaglandin derivatives as neurite outgrowth-promoting compounds for CAD cells, a rat catecholaminergic neuronal cell line of the central nervous system.
Here we reported the effects of neurite outgrowth-promoting prostaglandins (NEPP's) on neurons of the central nervous system (CNS). Serum deprivation promoted neurite outgrowth from CAD cells, a CNS-derived cathecholaminergic neuronal cell line. NEPP's (0.05-0.2 microM) accelerated the neurite outgrowth from CAD cells in serum-free medium but didn't in serum-containing medium. Through the study of structure-function relationship with the NEPP's 1-10, NEPP 10 (13,14-dihydro-15-epi-Delta(7)-prostaglandin A(1) (methyl ester) revealed the best compound, exhibiting potent neurite outgrowth-promoting activity with minimal cytotoxicity, suggesting that it is the best compound for drug development.
Animals↗
Stimulation of Ras guanine nucleotide exchange activity of Ras-GRF1/CDC25(Mm) upon tyrosine phosphorylation by the Cdc42-regulated kinase ACK1.
Ras-GRF1 is a brain-specific guanine nucleotide exchange factor (GEF) for Ras, whose activity is regulated in response to Ca(2+) influx and G protein-coupled receptor signals. In addition, Ras-GRF1 acts as a GEF for Rac when tyrosine-phosphorylated following G protein-coupled receptor stimulation. However, the mechanisms underlying the regulation of Ras-GRF1 functions remain incompletely understood. We show here that activated ACK1, a nonreceptor tyrosine kinase that belongs to the focal adhesion kinase family, causes tyrosine phosphorylation of Ras-GRF1. On the other hand, kinase-deficient ACK1 exerted no effect. GEF activity of Ras-GRF1 toward Ha-Ras, as defined by in vitro GDP binding and release assays, was augmented after tyrosine phosphorylation by ACK1. In contrast, GEF activity toward Rac1 remained latent, implying that ACK1 does not represent a tyrosine kinase that acts downstream of G protein-coupled receptors. Consistent with enhanced Ras-GEF activity, accumulation of the GTP-bound form of Ras within the cell was shown through the use of Ras-binding domain pull-down assays. Furthermore, Ras-dependent activation of ERK2 by Ras-GRF1 was enhanced following co-expression of activated ACK1. These results implicate ACK1 as an upstream modulator of Ras-GRF1 and suggest a signaling cascade consisting of Cdc42, ACK1, Ras-GRF1, and Ras in neuronal cells.
Cell Line↗
Palladium-catalyzed arylation of cyclopentadienes
Cyclopentadiene and metallocenes, typically zirconocene dichloride, are suitable substrates for multiple arylations with aryl bromides in palladium-catalyzed reactions. Thus, various aryl bromides bearing either an electron-donating or an electron-withdrawing substituent can react with these substrates to afford the corresponding 1,2,3,4,5-pentaaryl-1,3-cyclopentadienes in a single preparative step. Derivatives of cyclopentadiene, including di- and trisubstituted cyclopentadienes, and indene are arylated in a similar fashion.
Journal Article↗
MgcRacGAP is involved in the control of growth and differentiation of hematopoietic cells.
In a search for key molecules that prevent murine M1 leukemia cells from undergoing interleukin (IL)-6-induced differentiation into macrophages, we isolated an antisense complementary DNA (cDNA) that encodes full-length mouse MgcRac-GTPase-activating protein (GAP) through functional cloning. Forced expression of this antisense cDNA profoundly inhibited IL-6-induced differentiation of M1 cells into macrophage lineages. We also isolated a full-length human MgcRacGAP cDNA, which encodes an additional N-terminal polypeptide of 105 amino acid residues compared with the previously published human MgcRacGAP. In human HL-60 leukemic cells, overexpression of the full-length form of human MgcRacGAP alone induced growth suppression and macrophage differentiation associated with hypervacuolization and de novo expression of the myelomonocytic marker CD14. Analyses using a GAP-inactive mutant and 2 deletion mutants of MgcRacGAP indicated that the GAP activity was dispensable, but the myosin-like domain and the cysteine-rich domain were indispensable for growth suppression and macrophage differentiation. The present results indicated that MgcRacGAP plays key roles in controlling growth and differentiation of hematopoietic cells through mechanisms other than regulating Rac GTPase activity.
Amino Acid Sequence↗
Plasma brain natriuretic peptide as a prognostic indicator in patients with primary pulmonary hypertension.
BACKGROUND: Plasma brain natriuretic peptide (BNP) level increases in proportion to the degree of right ventricular dysfunction in pulmonary hypertension. We sought to assess the prognostic significance of plasma BNP in patients with primary pulmonary hypertension (PPH). METHODS AND RESULTS: Plasma BNP was measured in 60 patients with PPH at diagnostic catheterization, together with atrial natriuretic peptide, norepinephrine, and epinephrine. Measurements were repeated in 53 patients after a mean follow-up period of 3 months. Forty-nine of the patients received intravenous or oral prostacyclin. During a mean follow-up period of 24 months, 18 patients died of cardiopulmonary causes. According to multivariate analysis, baseline plasma BNP was an independent predictor of mortality. Patients with a supramedian level of baseline BNP (>/=150 pg/mL) had a significantly lower survival rate than those with an inframedian level, according to Kaplan-Meier survival curves (P<0.05). Plasma BNP in survivors decreased significantly during the follow-up (217+/-38 to 149+/-30 pg/mL, P<0. 05), whereas that in nonsurvivors increased (365+/-77 to 544+/-68 pg/mL, P<0.05). Thus, survival was strikingly worse for patients with a supramedian value of follow-up BNP (>/=180 pg/mL) than for those with an inframedian value (P<0.0001). CONCLUSIONS: A high level of plasma BNP, and in particular, a further increase in plasma BNP during follow-up, may have a strong, independent association with increased mortality rates in patients with PPH.
Adolescent↗
All eight stereoisomeric D-glyconic-delta-lactams: synthesis, conformational analysis, and evaluation as glycosidase inhibitors.
An efficient and general synthetic route to all eight stereoisomeric D-glycono-delta-lactams has been developed. The strategy involves, as a key step, a stereodivergent delta-lactam formation with configurational retention or inversion at C-4 of a starting gamma-lactone to lead to two epimers of delta-lactam from one parent gamma-lactone. Conformations of eight glycono-delta-lactams were examined by X-ray crystallographic analysis and molecular modeling. Analyses of conformation and glycosidase-inhibition provide useful information for the design of new glycosidase inhibitors.
Animals↗
Regulation of cyclophosphamide-induced eosinophilia in contact sensitivity: functional roles of interleukin-5-producing CD4(+) lymphocytes.
Blood and tissue eosinophilia is obtained when mice pretreated with cyclophosphamide (CY) and sensitized with picryl chloride are challenged on each ear lobe on day 13. To gain important insights into the cellular mechanisms involved in CY-induced eosinophilia in the contact-sensitivity reaction, we examined the cytokine profile expressed in regional lymph node cells and spleen cells. CY pretreatment 2 days before sensitization enhanced expression of IL-4 mRNA in the regional lymph node cells more strongly than expressions of both IL-2 and IFN-gamma mRNA on day 13. Five days after sensitization, spleen cells expressed IL-5 mRNA and produced IL-5 in vitro. Depletion of CD4(+) cells from spleen cells completely abrogated the secretory capacity of IL-5. In vivo blocking of IL-5 on day 3 entirely inhibited spleen, bone marrow, and subsequent blood eosinophilia. When immune lymph node cells prepared on day 13 were stimulated with hapten-modified cells in vitro, the level of IL-4 secreted in the culture supernatant was enhanced by CY pretreatment, but that of IL-2 was not. One important result was that IL-5 was not produced in response to in vitro stimulation, despite the fact that marked eosinophil infiltration in the dermis was observed in vivo. Thus, eosinophilopoiesis was stimulated by IL-5-producing CD4(+) immune T cells that were present in the eosinophil production site, particularly in the spleen before elicitation. In contrast, eosinophil recruitment into the dermis in the efferent phase can be induced without production of IL-5 from lymphocytes.
Animals↗
Discovery of novel and potent retinoic acid receptor alpha agonists: syntheses and evaluation of benzofuranyl-pyrrole and benzothiophenyl-pyrrole derivatives.
In the course of our studies on retinoic acid receptor (RAR) agonists, we have designed and synthesized a series of benzofuran and benzothiophene derivatives. Some of these compounds (1a,b,e,f,j) markedly inhibited LPS-induced B-lymphocyte proliferation and exerted RARalpha selectivity. One of them, 4-[5-(4,7-dimethylbenzofuran-2-yl)pyrrol-2-yl]benzoic acid (1b), when orally administered significantly inhibited mouse antibody production and delayed type hypersensitivity (DTH) responses from a dose of 0.1 mg/kg.
Administration, Oral↗
Neuroprotection by MAPK/ERK kinase inhibition with U0126 against oxidative stress in a mouse neuronal cell line and rat primary cultured cortical neurons.
Oxidative stress is implicated in the pathogenesis of neuronal degenerative diseases. Oxidative stress has been shown to activate extracellular signal-regulated kinases (ERK)1/2. We investigated the role of these mitogen-activated protein kinases (MAPKs) in oxidative neuronal injury by using a mouse hippocampal cell line (HT22) and rat primary cortical cultures. Here, we show that a novel MAPK/ERK kinase (MEK) specific inhibitor U0126 profoundly protected HT22 cells against oxidative stress induced by glutamate, which was accompanied by an inhibition of phosphorylation of ERK1/2. U0126 also protected rat primary cultured cortical neurons against glutamate or hypoxia. However, U0126 was not protective against death caused by tumor necrosis factor alpha (TNFalpha), A23187, or staurosporine. These results indicate that MEK plays a central role in the neuronal death caused by oxidative stress.
Animals↗
Species difference in enantioselectivity for the oxidation of propranolol by cytochrome P450 2D enzymes.
We examined and compared enantioselectivity in the oxidation of propranolol (PL) by liver microsomes from humans and Japanese monkeys (Macaca fuscata). PL was oxidized at the naphthalene ring to 4-hydroxypropranolol, 5-hydroxypropranolol and side chain N-desisopropylpropranolol by human liver microsomes with enantioselectivity of [R(+)>S(-)] in PL oxidation rates at substrate concentrations of 10 microM and 1 mM. In contrast, reversed enantioselectivity [R(+)<S(-)] in PL 5-hydroxylation and N-desalkylation rates at the same substrate concentrations was observed in monkey liver microsomes, although the selectivity was the same for PL 4-hydroxylation between the two species. All oxidation reactions of the PL enantiomers in human liver microsomes showed biphasic kinetics, i.e. the reactions could be expressed as the summation of a low-K(m) phase and a high-K(m) phase. Inhibition studies using antibodies and characterization of CYP2D6 enzymes expressed in insect cells or human lymphoblastoid cells indicated that the enantioselectivity of PL oxidation, especially the ring 4- and 5-hydroxylations reflected the properties of CYP2D6 in human liver microsomes. In monkey liver microsomes, all of the oxidation reactions of S(-)-PL showed biphasic kinetics, whereas ring 4- and 5-hydroxylations were monophasic and side chain N-desisopropylation was biphasic for R(+)-PL. Similarly, from the results of inhibition studies using antibodies and inhibitors of cytochrome P450 (P450), it appears that the reversed selectivity [R(+)<S(-)] of PL oxidation rates is catalyzed by CYP2D enzyme(s) in monkey liver at low substrate concentrations. These results indicate that different properties of P450s belonging to the 2D subfamily cause the reversed enantioselectivity between human and monkey liver microsomes.
Animals↗
Improvement in exercise capacity with nitric oxide inhalation in patients with precapillary pulmonary hypertension.
BACKGROUND: Patients with precapillary pulmonary hypertension (PH) exhibit a poor exercise capacity due to an impaired vasodilatory response of their pulmonary arteries. By causing the pulmonary artery to dilate, inhaled nitric oxide (NO) may allow an increase in exercise capacity in patients with PH. METHODS AND RESULTS: On 2 separate days, 3 days apart, 14 patients with precapillary PH (10 primary PH, 4 residual PH after correction of an intracardiac shunt; age, 40+/-12 years; mean pulmonary artery pressure, 60+/-23 mm Hg) performed exercise, with and without inhalation of 20 ppm NO, on a cycle ergometer. The work rate was increased 15 W/min until their symptom-limited maximum, with breath-by-breath gas analysis. Patients were randomly and blindly selected to inhale NO on either their first or second test. Peak exercise load and anaerobic threshold tended to increase, but not significantly. Peak oxygen consumption (f1.gif" BORDER="0">O(2)) and Deltaf1.gif" BORDER="0">O(2)/DeltaW ratio increased significantly, by 18% and 22%, respectively (peak f1.gif" BORDER="0">O(2), 13.6+/-3.6 to 16.0+/-4. 1 mL. kg(-1). min(-1); Deltaf1.gif" BORDER="0">O(2)/DeltaW ratio, 5. 8+/-2.4 to 7.1+/-2.3 mL. kg(-1). min(-1). W(-1); both P<0.01). Peak f1.gif" BORDER="0">O(2) increased >10% in 12 of the 14 patients. However, respiratory quotient at peak exercise decreased from 1. 22+/-0.15 to 1.09+/-0.15 (P<0.01). CONCLUSIONS: Inhaled NO substantially increases oxygen consumption at the same workload during exercise. This finding supports the possibility of ambulatory NO inhalation therapy in patients with precapillary PH.
Administration, Inhalation↗
Halothane anesthesia decreases the extracellular level of dopamine in rat striatum: a microdialysis study in vivo.
PURPOSE: In our previous microdialysis study, sevoflurane or isoflurane anesthesia significantly decreased the extracellular level of dopamine in rat striatum in vivo. On the other hand, other investigators demonstrated that halothane anesthesia either increased or did not affect the extracellular dopamine level. To explore the differences among these volatile anesthetics, the effects of halothane and nitrous oxide on the striatal dopamine level were reinvestigated. METHODS: Halothane alone, nitrous oxide with or without halothane, or drugs known to affect the dopaminergic pathway were administered to rats. Microdialysates were collected every 20 min and directly applied to an on-line high-performance liquid chromatograph without any pretreatment. The effects of halothane on respiratory and cardiovascular variables were monitored. RESULTS: General anesthesia with halothane alone decreased the dialysate (extracellular) concentration of dopamine but increased that of dopamine metabolites. Nitrous oxide alone slightly increased dopamine metabolites in dialysates but did not affect the halothane-induced decrease in extracellular dopamine. Apomorphine and haloperidol reproduced reported results, confirming the adequacy of our methodology. Nomifensine- or methamphetamine-induced increase in extracellular dopamine was augmented by halothane. CONCLUSION: These results suggest that halothane potently enhances striatal dopamine release and activates the reuptake or metabolic process, which is consistent with our previous results for sevoflurane or isoflurane. Volatile anesthetics interfere with dopamine regulation, at least in the rat striatum.
Journal Article↗
Signal transducer and activator of transcription 6 is essential in the induction of contact hypersensitivity.
Contact hypersensitivity (CHS) is thought to be mainly associated with the activation of T helper type 1 (Th1) cells. However, there is also evidence that Th2 cells or Th2 cytokines play a role in the development of CHS. To analyze the functional contribution of Th2 cytokines interleukin (IL)-4 and IL-13, signal transducer and activator of transcription 6 (STAT6)-deficient (STAT6(-/)-) and wild-type (wt) control C57BL/6 mice were contact sensitized with 5% 2,4,6-trinitrochlorobenzene (TNCB), 0.5% 2,4-dinitrofluorobenzene, or 5% 4-ethoxyl methylene-2-phenyl-2-oxazolin-5-one, and any skin reactions were examined. Ear swelling was significantly reduced with a delayed peak response in STAT6(-/)- mice compared with wt mice.A histological analysis revealed that the infiltration of both eosinophils and neutrophils in the skin challenged after 24 h in STAT6(-/)- mice decreased substantially compared with that in wt mice. The expression of Th2 cytokines (IL-4, IL-5) in TNCB-challenged skin tissues and the supernatants from T cells stimulated by 2,4,6-trinitrobenzene sulfonate-modified spleen cells, as well as the immunoglobulin (Ig)E and IgG1 response after challenge, were also profoundly reduced in STAT6(-/)- mice, whereas the expression of interferon gamma was the same in STAT6(-/)- and wt mice after challenge. Furthermore, adoptive transfer experiments revealed that STAT6(-/)- mice induced CHS after injection of lymph node cells obtained from sensitized wt mice. Our data suggest that the STAT6 signal plays a critical role in the induction phase of CHS.
Animals↗