RESUMEN
Pod setting rate in soybean is an important trait that determines pod number, which is highly correlated with seed yield. Using two soybean cultivars with different pod setting rates, we examined the relationship between plant growth regulation by gibberellin (GA) and pod setting rate. Plant growth rate (PGR) after flowering was significantly higher in 'Fukuyutaka' (low pod setting rate) than in 'Kariyutaka' (high pod setting rate); this difference was caused by increasing of GA biosynthesis-related genes expression. Additionally, pod setting rate in 'Fukuyutaka' was lower than that in 'Kariyutaka'. Furthermore, when 'Kariyutaka' was treated with GA after flowering, the PGR increased and pod setting rate decreased. These results suggest that pod setting rate in soybean is regulated by vegetative growth after flowering through GA biosynthesis.
Asunto(s)
Giberelinas/biosíntesis , Giberelinas/metabolismo , Glycine max/metabolismo , Reguladores del Crecimiento de las Plantas/biosíntesis , Reguladores del Crecimiento de las Plantas/metabolismoRESUMEN
We determined the contribution of cytochrome P450 (CYP) isoforms to the metabolism of midazolam by kinetic analysis of human liver microsomes and CYP isoforms and by examining the effect of chemical inhibitors and monoclonal antibodies against CYP isoforms in vitro. Midazolam was metabolized to 1'-hydroxymidazolam (1'-OH MDZ) by human liver microsomes with a Michaelis-Menten constant (Km) of 4.1 (1.0) (mean (SD)) micromol litre(-1) and a maximum rate of metabolism (Vmax) of 5.5 (1.1) nmol min(-1) mg protein(-1) (n = 6). Of the nine representative human liver CYP isoforms, CYP1A2, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1, 3A4 and 3A5, three (CYP2B6, 3A4 and 3A5) showed midazolam 1'-hydroxylation activity, with Kms of 40.7, 1.7 and 3.0 micromol litre(-1), respectively, and Vmax values of 12.0, 3.3 and 13.2 nmol min(-1) nmol P450(-1), respectively (n = 4). Midazolam 1'-hydroxylation activity of human liver microsomes correlated significantly with testosterone 6beta-hydroxylation activity, a marker of CYP3A activity (r2 = 0.77, P = 0.0001), but not with S-mephenytoin N-demethylation activity, a marker of CYP2B6 activity (r2 < 0.01, P = 0.84) (n = 11). Troleandomycin and orphenadrine, chemical inhibitors of CYP isoforms, inhibited the formation of 1'-OH MDZ by human liver microsomes. Monoclonal antibody against CYP3A4 inhibited the formation of 1'-OH MDZ by 79%, whereas monoclonal antibody against CYP2B6 had no effect on midazolam 1'-hydroxylation by human liver microsomes (n = 5). These results indicate that only CYP3A4, but not CYP2B6 or CYP2C, is involved in the metabolism of midazolam in vitro.
Asunto(s)
Anestésicos Intravenosos/metabolismo , Ansiolíticos/metabolismo , Hidrocarburo de Aril Hidroxilasas , Sistema Enzimático del Citocromo P-450/fisiología , Midazolam/metabolismo , Oxidorreductasas N-Desmetilantes/fisiología , Esteroide 16-alfa-Hidroxilasa , Esteroide Hidroxilasas/fisiología , Anticuerpos Monoclonales/inmunología , Citocromo P-450 CYP1A2 , Citocromo P-450 CYP2B6 , Inhibidores Enzimáticos del Citocromo P-450 , Sistema Enzimático del Citocromo P-450/inmunología , Sistema Enzimático del Citocromo P-450/metabolismo , Inhibidores Enzimáticos/farmacología , Humanos , Hidroxilación , Microsomas Hepáticos/metabolismo , Orfenadrina/farmacología , Oxidorreductasas N-Desmetilantes/antagonistas & inhibidores , Oxidorreductasas N-Desmetilantes/inmunología , Oxidorreductasas N-Desmetilantes/metabolismo , Esteroide Hidroxilasas/antagonistas & inhibidores , Esteroide Hidroxilasas/inmunología , Esteroide Hidroxilasas/metabolismo , Troleandomicina/farmacologíaRESUMEN
AIMS: To determine the cytochrome P450 (CYP) isoforms involved in the oxidation of propofol by human liver microsomes. METHODS: The rate constant calculated from the disappearance of propofol in an incubation mixture with human liver microsomes and recombinant human CYP isoforms was used as a measure of the rate of metabolism of propofol. The correlation of these rate constants with rates of metabolism of CYP isoform-selective substrates by liver microsomes, the effect of CYP isoform-selective chemical inhibitors and monoclonal antibodies on propofol metabolism by liver microsomes, and its metabolism by recombinant human CYP isoforms were examined. RESULTS: The mean rate constant of propofol metabolism by liver microsomes obtained from six individuals was 4.2 (95% confidence intervals 2.7, 5.7) nmol min(-1) mg(-1) protein. The rate constants of propofol by microsomes were significantly correlated with S-mephenytoin N-demethylation, a marker of CYP2B6 (r = 0.93, P < 0.0001), but not with the metabolic activities of other CYP isoform-selective substrates. Of the chemical inhibitors of CYP isoforms tested, orphenadrine, a CYP2B6 inhibitor, reduced the rate constant of propofol by liver microsomes by 38% (P < 0.05), while other CYP isoform-selective inhibitors had no effects. Of the recombinant CYP isoforms screened, CYP2B6 produced the highest rate constant for propofol metabolism (197 nmol min-1 nmol P450-1). An antibody against CYP2B6 inhibited the disappearance of propofol in liver microsomes by 74%. Antibodies raised against other CYP isoforms had no effect on the metabolism of propofol. CONCLUSIONS: CYP2B6 is predominantly involved in the oxidation of propofol by human liver microsomes.
Asunto(s)
Hidrocarburo de Aril Hidroxilasas , Sistema Enzimático del Citocromo P-450/metabolismo , Microsomas Hepáticos/enzimología , Oxidorreductasas N-Desmetilantes/metabolismo , Propofol/metabolismo , Analgésicos Opioides/metabolismo , Citocromo P-450 CYP2B6 , Sistema Enzimático del Citocromo P-450/aislamiento & purificación , Humanos , Técnicas In Vitro , Microsomas Hepáticos/metabolismo , Oxidación-Reducción , Oxidorreductasas N-Desmetilantes/aislamiento & purificaciónRESUMEN
Fentanyl decreases clearance of midazolam administered i.v., but the mechanism remains unclear. To elucidate this mechanism, we have investigated the effect of fentanyl on metabolism of midazolam using human hepatic microsomes and recombinant cytochrome P450 isoforms (n = 6). Midazolam was metabolized to l'-hydroxymidazolam (l'-OH MDZ) by human hepatic microsomes, with a Michaelis-Menten constant (K(m)) of 5.0 (SD 2.7) mumol litre-1. Fentanyl competitively inhibited metabolism of midazolam in human hepatic microsomes, with an inhibition constant (Ki) of 26.8 (12.4) mumol litre-1. Of the seven representative human hepatic P450 isoforms, CYP1A2, 2A6, 2C9, 2C19, 2D6, 2E1 and 3A4, only CYP3A4 catalysed hydroxylation of midazolam, with a K(m) of 3.6 (0.8) mumol liter-1. Fentanyl competitively inhibited metabolism of midazolam to l'-OH MDZ by CYP3A4, with a Ki of 24.2 (6.8) mumol litre-1, comparable with the Ki obtained in human hepatic microsomes. These findings indicate that fentanyl competitively inhibits metabolism of midazolam by CYP3A4.
Asunto(s)
Anestésicos Intravenosos/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Fentanilo/metabolismo , Isoenzimas/metabolismo , Microsomas Hepáticos/metabolismo , Midazolam/metabolismo , Oxigenasas de Función Mixta/metabolismo , Unión Competitiva , Técnicas de Cultivo , Citocromo P-450 CYP3A , Humanos , Proteínas Recombinantes/metabolismoRESUMEN
Samples of feces and vomit collected from patients during 13 non-bacterial gastroenteritis outbreaks which occurred in Hokkaido between 1995 and 1998 were examined by electron microscopy (EM) and reverse-transcription polymerase chain reaction (RT-PCR) for evidence of infection with human caliciviruses (HuCVs). In 6 food-borne outbreaks, oysters were the probable source of infection, while the origin of HuCVs was not found out for the other 7 outbreaks. One-hundred-eleven of 214 stool, vomit and oyster specimens examined gave positive results by RT-PCR, while HuCVs were detected by EM in 36 of 121 stool specimens examined. We determined the nucleotide sequences of 470-bp or 373-bp PCR products amplified from the RNA polymerase region of the HuCV genomes with primer sets MR3/4 and Yuri22F/R, respectively. The sequences of different strains revealed great heterogenicity, with a range of 60 to 100% homology among strains. In a few cases, a mixed genotype was found in the same patient or same outbreak by means of nested PCR and cloning of PCR products into an appropriate vector. Of the 19 different strains found, 4 strains could be classified as Norwalk virus (genogroup 1) and the other 15 strains as Snow Mountain agent (genogroup 2) based on genotyping with homology analysis. Furthermore, the strains belonging to genogroup 2 could be classified into 4 subgroups with more than 93% homology in amino acids among strains in the subgroup.
Asunto(s)
Infecciones por Caliciviridae/virología , Caliciviridae/genética , Caliciviridae/aislamiento & purificación , Gastroenteritis/virología , Animales , Infecciones por Caliciviridae/epidemiología , Cartilla de ADN , ADN Viral/análisis , ADN Viral/genética , Brotes de Enfermedades , Heces/virología , Gastroenteritis/epidemiología , Humanos , Japón/epidemiología , Microscopía Electrónica , Datos de Secuencia Molecular , Ostreidae/virología , Filogenia , Reacción en Cadena de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ADNRESUMEN
OBJECTIVE: To examine the effect of propofol on the pharmacokinetics of midazolam in vivo and to elucidate the mechanism of the pharmacokinetic changes of midazolam by propofol with the use of human liver microsomes and recombinant CYP3A4. METHODS: In an in vivo, double-blind randomized study, 24 patients received 0.2 mg/kg midazolam and either 2 mg/kg propofol (propofol group) or placebo (placebo group) for induction of anesthesia. In the propofol group, continuous infusion of propofol at 9 mg/kg/h was started immediately after the bolus infusion of propofol and was maintained for an hour. In the placebo group the same dose of soybean emulsion as a placebo was given and infused intravenously for an hour instead of propofol. In an in vitro study the effect of propofol on the metabolism of midazolam was studied with human liver microsomes and recombinant CYP3A4. RESULTS: In the propofol group the mean clearance of midazolam was decreased by 37% (P = .005) and the mean elimination half-life was prolonged by 61% (P = .04) compared with the placebo group. The mean plasma concentrations of 1'-hydroxymidazolam were lower in the propofol group than in the placebo group at 5, 10, 15, 20, and 30 minutes after midazolam was administered (P < .05). The mean (+/-SD) Michaelis-Menten constant for midazolam 1'-hydroxylation by human liver microsomes was 5.6 +/- 3.3 micromol/L. The formation of 1'-hydroxymidazolam was competitively inhibited by propofol, and the mean inhibition constant was 56.7 +/- 16.6 micromol/L. The mean Michaelis-Menten constant and mean inhibition constant values for midazolam 1'-hydroxylation by recombinant CYP3A4 were 4.0 micromol/L and 61.0 micromol/L, respectively, consistent with the mean values obtained from human liver microsomes. CONCLUSION: Propofol decreases the clearance of midazolam, and the possible mechanism is the competitive inhibition of hepatic CYP3A4.