Effect of long-term multiple nifedipine administration on the antinociceptive activity of acetaminophen

The influence of long-term nifedipine administration on the antinociceptive activity of acetaminophen on hexobarbital sleeping time and liver monooxygenase and synthetase activities was studied in male albino mice. Nifedipine was administered orally at a dose of 25 mg/kg daily for 14 and 21 days. The nociceptive response was determined by the acetic acid writhing test. There was no significant difference in the antinociceptive effect of acetaminophen after treatment with nifedipine for 14 days. Nifedipine caused enzyme induction, which was demonstrated by shortened hexobarbital sleeping time, enhanced ethylmorphine-N-demethylase (EMND), aniline-4-hydroxylase (AH), ethoxycoumarine-O-deethylase (ECOD), UDP-glucuronyl transferase (UDPGT), glutathione-S-transferase (GST) and NADPH-cytochrome c reductase activity and increased content of cytochrome P450 and cytochrome b5. It is assumed that this effect of nifedipine on acetaminophen analgesia is associated with the changes (acceleration) in acetaminophen metabolism in the liver after repeated administration of the drug.     

Changes in rat liver monooxygenase activities after administration of atenolol, nifedipine and diltiazem alone and in combination

The effects of the Ca2+ antagonists nifedipine (NF) and diltiazem (DL) and of the cardioselective beta 1-adrenergic blocking agent atenolol (AT) on the hexobarbital (HB) sleeping time and on the activity of some liver drug-metabolizing enzyme systems in male Wistar rats were studied. Two hours after single oral administration, atenolol (150 mg/kg) did not change hexobarbital sleeping time, while nifedipine (50 mg/kg) and diltiazem (30 mg/kg) prolonged it by 171.2 and 99.6%, respectively. Coadministration of atenolol with diltiazem or with nifedipine significantly prolonged hexobarbital sleep by 205 and 283%, respectively. Administered alone, atenolol decreased the ethylmorphine-N-demethylase (EMND) activity, but the amidopyrine-N-demethylase (APND) activity was not changed in any of the treated groups. Atenolol and nifedipine significantly increased aniline-4-hydroxylase (AH) activity and this effect was also observed with the combinations AT + NF and AT + DL. The NADPH cytochrome P-450 reductase activity was significantly decreased by nifedipine and diltiazem. Only nifedipine increased the total content of cytochrome P-450 (by 23.8%). Atenolol and diltiazem tended to increase the content of cytochrome b5 which was increased by nifedipine by 97.6%. The same effect was observed with the combinations AT + NF and AT + DL. The results suggest that NF, AT + NF and AT + DL produced the manifested changes in hepatic oxidative metabolism. The decreased EMND activity by atenolol, however, and the prolongation of hexobarbital sleeping time by nifedipine, diltiazem and their coadministration with atenolol did not correlate with enhanced microsomal P-450 and b5 content.     

Interactions of melatonin with the liver microsomal cytochrome P450 system of rats and humans in vitro and effects on the P450 system and the antioxidative status in rat liver after acute treatment.

In vitro melatonin binds to human and rat liver microsomal cytochrome P-450 (P450) according to a type II substrate. The affinity is similar to that of aniline with a general left-shift. Melatonin interferes with model monooxygenase reactions indicative of different P450 forms in humans and rats (in humans according to the lower specific P450 content less pronounced): the strongest inhibition was found for ethoxyresorufin O-deethylation, indicating the binding to P450 1A, the binding to P450 2B (ethoxycoumarin O-deethylation) was less pronounced, the least inhibition was found for P450 3A (ethylmorphine N-demethylation) reaction. The oxidase function was also inhibited: luminol amplified chemiluminescence was more inhibited than the lucigenin amplified one, hydrogen peroxide formation was inhibited at concentrations higher than 10(-4) M, microsomal NADPH/Fe stimulated lipid peroxidation was inhibited at concentrations higher than 10(5) M. In vivo melatonin prolonged hexobarbital sleeping time in rats in a dose dependent manner (ip. co-administration of 1, 5 and 20 mg/kg b.w. melatonin with 100 mg/kg hexobarbital). Immediately after awakening the animals were sacrificed: a small increase in P450 concentrations cannot be explained, no changes in P450 monooxygenase or oxidase activities nor in microsomal lipid peroxidation or GSH status could be observed.     

Protective effect of diethyldithiocarbamate and carbon disulfide against liver injury induced by various hepatotoxic agents

Diethyldithiocarbamate (DTC) and carbon disulfide (CS2), at nearly equimolar oral dose levels, protected mice against liver damage induced by carbon tetrachloride, chloroform, bromotrichloromethane, thioacetamide, bromobenzene, furosemide, acetaminophen, dimethylnitrosamine and trichloroethylene, as evidenced by the suppression of elevations in plasma GPT activity and liver calcium content, and of histopathological alterations. Both agents also prolonged hexobarbital sleeping time and zoxazolamine paralysis time in mice. DTC and SC, alone, given orally, decreased microsomal metabolism of several substrates (aniline, p-nitroanisole, hexobarbital, zoxazolamine, aminopyrine and 3,4-benzopyrene), CC14-induced lipid peroxidation, and cytochrome P-450 content. The loss of microsomal drug-metabolizing enzyme activity was also observed in the experiments in vitro using liver slices and isolated microsomes. Since a characteristic common to such diverse hepatotoxins is that they require metabolic activation before exhibiting hepatotoxicity, the protective mechanisms of DTC and CS2 may involve their interference with the process of metabolic activation of these hepatotoxins. The protective action of DTC may be mediated almost entirely through CS2 when administered orally and at least partly with parenteral administration, since, in CCl4-induced liver injury, DTC was most effective when given orally, while the action of CS2 was less dependent on the route of administration. Thus CS2 and CS2-producing agents in vivo such as dithiocarbamate derivatives and disulfiram may modify toxicological and pharmacological effects of foreign compounds by inhibiting microsomal drug-metabolizing enzyme activity in the liver.     

年龄对大鼠肝脏生物转化功能及膜流动性的影响

通过与青年(3～4月)及中年(14月)组比较,研究了老年(24月)大鼠肝脏生物转化酶活性改变及膜流动性变化。结果表明,老年大鼠肝微粒体P-450含量、NADPH-细胞色素C还原酶活性无明显改变,但氨基比林N-脱甲基酶、苯胺羟化酶活性明显降低,且微粒体及胞浆GST、胞浆GSH-Px活性也明显下降;同时肝微粒体膜脂区流动性明显降低,膜Ch/PL值显著增大。研究提示,微粒体膜脂质环境及流动性变化与上述生物转化功能改变可能有一定的联系。     

Effect of glucose administration on various rat hepatic constituents and on the spectral binding of hexobarbital and methadone to rat hepatic cytochrome P450

The administration of glucose to rats for 48 h resulted in an increase in the duration of anesthesia produced by intraperitoneal injection of pentobarbital. The effect of this glucose treatment on hepatic glycogen and on microsomal protein, cytochrome P450 (P450), cholesterol, lipid and phospholipid content, as well as on the spectral binding of hexobarbital and methadone to microsomal P450, was examined. The glucose treatment appeared to have no effect on microsomal protein or P450 content. The binding of hexobarbital and methadone by P450 produced a type 1 spectrum in both control and glucose-treated animals. The glucose treatment resulted in a decrease in the spectral dissociation constant (Ks) and in the maximal spectral binding (delta Amax) of hexobarbital to P450 while with methadone there was an increase in Ks and a decrease in delta Amax. Total lipid, phosphatidylcholine and phosphatidylethanolamine were increased in the microsomal fractions from glucose-treated animals. The changes in the parameters for drug binding to microsomal P450 probably relate to the increased lipid content of the microsomes although changes in the proportion of P450 isoenzymes could be involved. The previously observed decrease in the microsomal metabolism of hexobarbital and methadone following glucose treatment may be due to decreased binding of these compounds to microsomal P450.     

硒多糖、亚砷酸钠对大鼠肝微粒体酶和GSH-Px等的影响

研究了硒多糖、亚砷酸钠在体内、外对大鼠肝微粒体酶细胞色素Ｐ－４５０、ｂ５、ＮＡＤ（Ｐ）Ｈ－细胞色素Ｃ还原酶、谷胱甘肽硫转移酶（ＧＳＴ）的影响;并通过测定硒多糖、亚砷酸钠对肝谷胱甘肽过氧化物酶（ＧＳＨ－Ｐｘ）和脂质过氧化（ＬＰＯ）的影响,探讨了硒、砷相互作用的机理。结果表明：连续７天腹腔注射０．２ｍｇ／ｋｇ硒多糖,细胞色素Ｐ－４５０、ｂ５的含量、ＧＳＴ的活性降低（Ｐ＜０．０５）;硒多糖明显诱导ＧＳＨ－Ｐｘ的活性,降低脂质过氧化,拮抗亚砷酸钠对ＬＰＯ的作用。亚砷酸钠显著增强肝细胞脂质过氧化（Ｐ＜０．０５）,对ＧＳＨ－Ｐｘ和肝微粒体酶无明显影响     

苯巴比妥诱导下大鼠肝微粒体药酶活性与膜流动性变化的相关性

本文用ANS和DPH为荧光探剂,研究苯巴比妥(PB)诱导下大鼠肝微粒体膜脂区流动性与膜药酶活性变化的相关性。结果表明,经PB诱导后在增加肝微粒体蛋白质含量,P-450含量及NADPH-细胞色素C还原酶等酶活性的同时,肝微粒体膜流动性明显增大,且膜深层流动性的增大与膜氨基比林N-脱甲基酶、细胞色素C还原酶活性增加有明显的直线正相关。膜胆固醇/碑脂比值明显降低。此结果提示,肝微粒体膜流动性的适当增大与PB增加单胺氧化酶系统活性之间可能存在着某种联系。     

Effect of excess and deficient copper intake on rat liver microsomal enzyme activity

The effect of copper loading and copper deficiency on rat liver microsomal enzyme activity was studied. A significant reduction of liver aniline hydroxylase activity was produced by the administration of 450 ppm copper in the drinking water to rats for 15 and 30 days. Lower levels of copper (50 and 150 ppm) administered to rats for the same time had no significant effect on enzyme activity. Copper loading did not alter the activities of liver glucose 6-phosphatase or benzpyrene hydroxylase. Aniline hydroxylase and hexobarbital oxidase activities were significantly reduced, and hexobarbital sleeping time was prolonged in copper-deficient rats. Liver enzyme activities and hexobarbital sleeping times were restored to control levels by feeding copper- deficient rats a copper-containing diet for 14 days. The addition of divalent copper in vitro to 10,000 g supernatants from liver homogenates from copper-deficient rats did not restore enzyme activity to control levels. Instead, high levels of added copper produced a decreased rate of aniline hydroxylation in vitro. Copper deficiency did not prevent the induction of liver microsomal enzyme activity by phenobarbital. Phenobarbital administration significantly increased microsomal copper in rats receiving a normal copper intake and increased both whole liver and microsomal copper concentration in copper-deficient rats. The normal developmental increase in liver aniline hydroxylase and hexobarbital oxidase activities with age was delayed in the offspring of female rats maintained on a copper-deficient diet.     

The effects of adriamycin in vitro and in vivo on hepatic microsomal drug-metabolizing enzymes: role of microsomal lipid peroxidation

The quinone-containing anticancer drug adriamycin augments the reduction of dioxygen to reactive oxygen species and thereby stimulates (sixfold) NADPH-dependent microsomal lipid peroxidation. In vitro the extensive adriamycin-promoted peroxidation depleted (85%) rat liver microsomal cytochrome P-450, severely inhibited cytochrome P-450-dependent monooxygenation (70%), and glucose-6-phosphatase activity (80%), and activated (450%) UDP-glucuronyltransferase activity. When lipid peroxidation was blocked by EDTA, adriamycin selectively decreased cytochrome P-450 and aminopyrine N-demethylase activity; NADPH-cytochrome c reductase, UDP-glucuronyltransferase, and glucose-6-phosphatase activities were unchanged. Washing and resedimenting peroxidized microsomes to remove adriamycin and soluble lipid peroxidation products failed to restore enzyme activities to control values. Adriamycin administered subacutely (5 mg/kg × three doses) to rats significantly descreased hepatic microsomal cytochrome P-450 content and reduced aminopyrine N-demethylase and NADPH-cytochrome c reductase activities compared to saline-treated controls. Microsomal lipid peroxidation was increased following the above adriamycin treatment. Thus, these data suggested that adriamycin was capable of impairing hepatic drug metabolism in vitro by stimulating membrane lipid peroxidation in a manner similar to carbon tetrachloride; the mechanism by which adriamycin treatment in vivo decreased the activity of the drug monooxygenase system remains unclear.     

苯巴比妥诱导下大鼠肝微粒体药酶活性与膜流动性变化的相关性

本文用ANS和DPH为荧光探剂,研究苯巴比妥(PB)诱导下大鼠肝微粒体膜脂区流动性与膜药酶活性变化的相关性。结果表明,经PB诱导后在增加肝微粒体蛋白质含量,P-450含量及NADPH-细胞色素C还原酶等酶活性的同时,肝微粒体膜流动性明显增大,且膜深层流动性的增大与膜氨基比林N-脱甲基酶、细胞色素C还原酶活性增加有明显的直线正相关。膜胆固醇/碑脂比值明显降低。此结果提示,肝微粒体膜流动性的适当增大与PB增加单胺氧化酶系统活性之间可能存在着某种联系。     

Metabolic fate of the new anti-ulcer drug enprostil in animals. 4th communication: effect on hepatic drug metabolizing enzyme system in rats

Effects of multiple oral administration of enprostil ((+/-)-11 alpha,15 alpha-dihydroxy-9-oxo-16-phenoxy-17,18,19,20-tetranorprosta- 4,5,13(t)-trienoic acid methyl ester, TA 84135) (20 micrograms/kg/d) and its solvent propylene carbonate (PC, 100 microliters/kg/d) on body weight gain, liver weight, hepatic drug metabolizing enzyme system and hexobarbital sleeping time were investigated in male rats during a 14-day period. Cytochrome P-450 content (as compared to the untreated control) and cytochrome b5 content (as compared to PC treated group) were slightly, but significantly, reduced in the group given a single oral dose of enprostil. However, these slight reductions were not augmented significantly by repeated administrations of enprostil. Slight but significant increase in microsomal protein content was observed in the group given 14 oral doses of enprostil and PC. Enprostil did not affect the other indicators used to evaluate the status of the hepatic drug metabolizing enzyme system. Additionally, single or multiple oral doses of enprostil or PC showed no effect on the hexobarbital-induced sleeping time. It therefore may be safely concluded that multiple oral administration, both of enprostil and of PC, has very little effect on drug metabolizing enzyme inducing or inhibiting activity in rats.     

Species differences of mixed-function oxidase induction between rabbits and rats after pretreatment with polychlorinated biphenyls (PCBs).

The inducing effect of PCB on mixed-function oxidase was investigated in rabbits. No induction of microsomal aminopyrine- and p-nitroanisole-demethylation as well as of aniline- and 4-chlorobiphenyl-p-hydroxylation was found, whereas the cytochrome P-450 level was increased by 150–300 percent normal after Clophen A 50 administration. In rat liver microsomes corresponding enzyme activities and P-450 levels were increased 2-5-fold after identical pretreatment. Similarly, hexobarbital half life remained unchanged in rabbits when hexobarbital sleeping time was reduced by more than 50 percent after the same dose of Clophen A 50 in the rat. Pretreatment of rabbits with different acute doses or with low chronic doses of Clophen A 50 increased P-450-content but confirmed the noninducibility of the enzyme activities measured. Evidence is presented that this phenomenon is not caused by inhibitory action of residual PCBs on mixed-function oxidase. The results are discussed with regard to the well known response of mixed-function oxidase in rabbit liver to 3-methylcholanthrene (3-MC)-treatment.     

Comparison of effects of acetaminophen on liver microsomal drug metabolism and lipid peroxidation in rats and mice.

Studies were conducted to determine the in vivo effect of acetaminophen (AAP) on the lipid peroxidation, drug metabolizing enzyme activity and microsomal electron transfer system of rat and mouse liver. AAP was found to inhibit ethylmorphine N-demethylase activity in the presence of NADPH and this inhibition of the enzyme was due to decrease in cytochrome P-450 content, but not due to change in lipid peroxidation in liver microsomes. Kinetical data showed that AAP administration had no effect on Km values of ethylmorphine N-demethylase, however, a decrease in the Vmax values was seen in rats and mice. There was no significant effect of AAP on both NADPH-cytochrome c reductase and the content of cytochrome b5 3 hours after this administration to rats and mice. On the other hand, AAP induced a significant decrease in NADH-ferricyanide reductase in mice, but not in rats. The greatest decrease in cytochrome P-450 observed among the components of the liver microsomal electron transfer system of rats and mice.     

Stimulation of lipid peroxidation and heme oxygenase activity with inhibition of cytochrome P-450 monooxygenase in the liver of rats repeatedly exposed to cadmium

Exposure of rats to 0.1 and 0.5 mg Cd/kg subcutaneously (s.c.) thrice weekly for 5 weeks resulted in an accumulation of cadmium in the liver in concentrations of 40 and 95 micrograms/g tissue, respectively, and a microsomal burden of Cd amounting to approx. 2-3% of the retained cadmium. The cytoplasm contained about 80% of the cadmium. At an exposure dose of 0.1 mg Cd/kg, stimulation of lipid peroxidation by 22% and inhibition of ALA synthetase by 16% in the liver were observed. The higher exposure of 0.5 mg Cd/kg caused an inhibition of microsomal monooxygenase with depression of cytochrome P-450 and cytochrome b5 by 20% (over 2-fold prolongation of hexobarbital sleeping time and statistically significant decrease of activity of aniline p-hydroxylase). The loss of cytochrome P-450 probably was due to an intensified lipid peroxidation and induction of heme oxygenase (30% and 60% over control, respectively). Sequestration of cadmium by cytoplasm (metallothionein) does not protect microsomes against cadmium accumulation and specific biochemical disturbances.     

Further studies on the in vivo effect of diisopropyl 1,3-dithiol-2-ylidenemalonate (NKK-105) on the liver microsomal drug oxidation system in rats

The effect of diisopropyl 1,3-dithiol-2-ylidenemalonate (NKK-105) on microsomal electron transport systems in relation to drug oxidation was studied in rat liver. A single oral dose (250 mg/kg) of NKK-105 increased the ratio of liver to body weight, the microsomal protein content, the cytochrome b₅ content, and the NADPH cytochrome c reductase activity at 24–48 hr after drug administration. The cytochrome P-450 content was decreased at 2–6 hr and slightly increased at 24–48 hr after drug administration. Upon daily administration of NKK-105 at a dose of 250 mg · kg^?1 · day^?1 for 21 days, cytochrome b₅ content and NADPH cytochrome c reductase activity were increased, but cytochrome P-450 content and NADH cytochrome b₅ reductase activity remained unchanged. Despite the increase of NADPH cytochrome c reductase activity, NADPH-dependent lipid peroxidation tended to decrease rather than increase. NADPH stearoyl-CoA desaturase activity increased prior to the increase of cytochrome b₅. Benzphetamine N-demethylase and p-nitroanisole O-demethylase activities were enhanced, accompanied by an increase of cytochrome b₅. Aniline hydroxylase activity was decreased by NKK-105 administration. These results indicate that the induction pattern of liver microsomal electron transport systems by NKK-105 is characteristic.     

Modulatory influence of noscapine on the ethanol-altered hepatic biotransformation system enzymes, glutathione content and lipid peroxidation in vivo in rats.

The modulatory potential of noscapine, an opium alkaloid was assessed on the ethanol-induced changes in hepatic drug metabolizing enzyme systems, glutathione content and microsomal lipid peroxidation. Noscapine was administered orally to male Wistar rats at a dose level of 200 mg/kg bw alone as well as in combination with 50% ethanol (v/v) for 5 days. Noscapine administration was associated with a approximately 91% decrease in hepatic microsomal cytochrome P-450 content. A decline of approximately 36% was observed in the NADPH-cytochrome c reductase activity on noscapine administration. The lowering of cytochrome P-450 levels on noscapine administration was accompanied by a concomitant increase in heme oxygenase activity as well as serum bilirubin levels. Our results indicate that the combination dosage of noscapine and ethanol antagonised the ethanol-induced elevation of cytochrome P-450 levels. Noscapine fed rats had decreased glutathione (GSH) content and enhanced lipid peroxidation compared to control rats as indexed by MDA method. Further, noscapine and ethanol coexposure produced a more pronounced elevation in lipid peroxidation and the glutathione levels also decreased significantly. We speculate on the basis of our results that the significant enhancement of lipid peroxidation on combination dosage of noscapine and ethanol is a consequence of depletion of glutathione to certain critical levels. The inhibition of glutathione-S-transferase (GST) as well as lowering of cytochrome P-450 suggests that the biotransformation of noscapine and ethanol is significantly altered following acute coexposures.     

Effects of motorcycle exhaust inhalation exposure on cytochrome P-450 2B1, antioxidant enzymes, and lipid peroxidation in rat liver and lung

The effects of motorcycle exhaust (ME) on metabolic and antioxidant enzymes and lipid peroxidation were determined using male rats exposed to 1:10 diluted ME by inhalation 2 h daily for 4 wk. For microsomal cytochrome P-450 enzymes, ME resulted in threefold increases of 7-ethoxyresorufin and pentoxyresorufin O-deethylase activities in liver and a sixfold increase of 7-ethoxyresorufin O-deethylase activity and an 80% decrease of pentoxyresorufin O-dealkylase activity in lung. The results of immunoblot analysis of microsomal proteins revealed that ME increased liver and lung cytochrome P-450 1A1 with minimal effects on cytochrome P-450 2E1. ME increased cytochrome P-450 2B1/2 proteins in liver but decreased cytochrome P-450 2B1 in lung. ME did not change microsomal cytochrome P-450 enzyme activity or protein level in kidney. For phase II enzymes, ME resulted in 53% and twofold increases of cytosolic NAD(P)H:quinone oxidoreductase activities in liver and lung, respectively, and no effect on microsomal UDP-glucuronosyltransferase activities. For antioxidant enzymes, ME produced 23% and 35% decreases of superoxide dismutase, 9% and 27% decreases of catalase, and no changes of glutathione peroxidase activities in liver and lung cytosols, respectively. For lipid peroxidation, the results of thiobarbituric acid assay showed that ME resulted in a twofold increase of formation of malondialdehyde by liver microsomes incubated with FeCl(3) -ADP. ME produced a threefold increase of malondialdehyde formation by lung microsomes. The present study demonstrates that ME inhalation exposure differentially modulates cytochrome P-450 2B1 and antioxidant enzymes and increases susceptibility to lipid peroxidation in rat liver and lung.     

Depression of cytochrome P-450-dependent drug biotransformation by adriamycin

The cytochrome P-450-dependent mixed function oxidase system was depressed following the administration of adriamycin. Cytochrome P-450, aminopyrine N-demethylase, and benzo(a)pyrene hydroxylase were significantly decreased in hepatic microsomes prepared from rats treated with a single dose of adriamycin (10 mg/kg subcutaneously) 4 days previously. Total microsomal protein levels and cytochrome b₅ levels remained unchanged. The administration of cysteamine 1 hr prior to adriamycin prevented the loss of the cytochrome P-450 and the decrease in drug biotransformation. The administration of diethylmaleate had no effect on the ability of adriamycin to decrease this enzyme system. The loss of drug biotransformation and the protection offered by cysteamine is correlated with the lipid peroxidation activity in hepatic microsomes. This suggests that the loss of cytochrome P-450 and related drug biotransformation is related to the generation of free radicals and subsequent lipid peroxidation in the liver.     

Hexobarbital sleeping time and drug metabolism in rats with ligated bile ducts—A lack of correlation

Hexobarbital sleeping time is commonly used as an index of the activity of hepatic microsomal drug-metabolizing enzymes in animals. This report describes anomalies between hexobarbital sleeping time and the rate of metabolism in vitro by microsomal enzymes in rats after bile duct ligation (BDL). The duration of hexobarbital sleeping time, 2–24 hr after BDL, was approximately twice that of sham-operated controls. No significant decrease in the activity of microsomal aminopyrine demethylase, aniline hydroxylase, hexobarbital oxidase or the amount of cytochrome P-450 was detected during this period. A further prolongation of hexobarbital sleeping time was observed 48–72 hr after BDL, and this was accompanied by a significant impairment of drug metabolism in vitro. The effect of BDL on hexobarbital sleeping time was independent of the route of administration. Thiopental sleeping time was prolonged at 12 and 72 hr after BDL. Zoxazolamine paralysis time was prolonged at 72 hr after BDL, but not at 12 hr. Plasma protein binding of hexobarbital and thiopental was not altered by hyperbilirubinemia. These data suggest that changes in drug metabolism are not responsible for the prolongation of hexobarbital sleeping time during the early phase of cholestasis caused by BDL.