Hepatic Cytochrome P-450 Content Research Articles

Mode of action analysis for fluxapyroxad-induced rat liver tumour formation: evidence for activation of the constitutive androstane receptor and assessment of human relevance

The fungicide fluxapyroxad (BAS 700F) has been shown to significantly increase the incidence of liver tumours in male Wistar rats at dietary levels of 1500 and 3000 ppm and in female rats at a dietary level of 3000 ppm via a non-genotoxic mechanism. In order to elucidate the mode of action (MOA) for fluxapyroxad-induced rat liver tumour formation a series of in vivo and in vitro investigative studies were undertaken. The treatment of male and female Wistar rats with diets containing 0 (control), 50, 250, 1500 and 3000 ppm fluxapyroxad for 1, 3, 7 and 14 days resulted in a dose-dependent increases in relative weight at 1500 and 3000 ppm from day 3 onwards in both sexes, with an increase in relative liver weight being also observed in male rats given 250 ppm fluxapyroxad for 14 days. Examination of liver sections revealed a centrilobular hepatocyte hypertrophy in some fluxapyroxad treated male and female rats. Hepatocyte replicative DNA synthesis (RDS) was significantly increased in male rats given 1500 and 3000 ppm fluxapyroxad for 3 and 7 days and in female rats given 50-3000 ppm fluxapyroxad for 7 days and 250-3000 ppm fluxapyroxad for 3 and 14 days; the maximal increases in RDS in both sexes being observed after 7 days treatment. The treatment of male and female Wistar rats with 250-3000 ppm fluxapyroxad for 14 days resulted in significant increases in hepatic microsomal total cytochrome P450 (CYP) content and CYP2B subfamily-dependent enzyme activities. Male Wistar rat hepatocytes were treated with control medium and medium containing 1-100μM fluxapyroxad or 500μM sodium phenobarbital (NaPB) for 4 days. Treatment with fluxapyroxad and NaPB increased CYP2B and CYP3A enzyme activities and mRNA levels but had little effect on markers of CYP1A and CYP4A subfamily enzymes and of the peroxisomal fatty acid β-oxidation cycle. Hepatocyte RDS was significantly increased by treatment with fluxapyroxad, NaPB and 25ng/ml epidermal growth factor (EGF). The treatment of hepatocytes from two male human donors with 1-100μM fluxapyroxad or 500μM NaPB for 4 days resulted in some increases in CYP2B and CYP3A enzyme activities and CYP mRNA levels but had no effect on hepatocyte RDS, whereas treatment with EGF resulted in significant increase in RDS in both human hepatocyte preparations. Hepatocytes from male Sprague-Dawley wild type (WT) and constitutive androstane receptor (CAR) knockout (CAR KO) rats were treated with control medium and medium containing 1-16μM fluxapyroxad or 500μM NaPB for 4 days. While both fluxapyroxad and NaPB increased CYP2B enzyme activities and mRNA levels in WT hepatocytes, only minor effects were observed in CAR KO rat hepatocytes. Treatment with both fluxapyroxad and NaPB only increased RDS in WT and not in CAR KO rat hepatocytes, whereas treatment with EGF increased RDS in both WT and CAR KO rat hepatocytes. In conclusion, a series of in vivo and in vitro investigative studies have demonstrated that fluxapyroxad is a CAR activator in rat liver, with similar properties to the prototypical CAR activator phenobarbital. A robust MOA for fluxapyroxad-induced rat liver tumour formation has been established. Based on the lack of effect of fluxapyroxad on RDS in human hepatocytes, it is considered that the MOA for fluxapyroxad-induced liver tumour formation is qualitatively not plausible for humans.

CYP2B6 allelic variants and non-genetic factors influence CYP2B6 enzyme function

Human CYP2B6 enzyme although constitutes relatively low proportion (1–4%) of hepatic cytochrome P450 content, it is the major catalyst of metabolism of several clinically important drugs (efavirenz, cyclophosphamide, bupropion, methadone). High interindividual variability in CYP2B6 function, contributing to impaired drug-response and/or adverse reactions, is partly elucidated by genetic polymorphisms, whereas non-genetic factors can significantly modify the CYP2B6 phenotype. The influence of genetic and phenoconverting non-genetic factors on CYP2B6-selective activity and CYP2B6 expression was investigated in liver tissues from Caucasian subjects (N = 119). Strong association was observed between hepatic S-mephenytoin N-demethylase activity and CYP2B6 mRNA expression (P < 0.0001). In less than one third of the tissue donors, the CYP2B6 phenotype characterized by S-mephenytoin N-demethylase activity and/or CYP2B6 expression was concordant with CYP2B6 genotype, whereas in more than 35% of the subjects, an altered CYP2B6 phenotype was attributed to phenoconverting non-genetic factors (to CYP2B6-specific inhibitors and inducers, non-specific amoxicillin + clavulanic acid treatment and chronic alcohol consumption, but not to the gender). Furthermore, CYP2B6 genotype–phenotype mismatch still existed in one third of tissue donors. In conclusion, identifying potential sources of CYP2B6 variability and considering both genetic variations and non-genetic factors is a pressing requirement for appropriate elucidation of CYP2B6 genotype–phenotype mismatch.

Piperonyl butoxide: Mode of action analysis for mouse liver tumour formation and human relevance

In a 79 week bioassay the pesticide synergist piperonyl butoxide (PBO) was shown to significantly increase the incidence of hepatocellular adenoma (but not hepatocellular carcinoma) in male CD-1 mice at dietary levels of 100 and 300 mg/kg/day PBO and in female mice at a dietary level of 300 mg/kg/day. As PBO is not a genotoxic agent, a series of investigative studies were undertaken to elucidate the mode of action (MOA) for PBO-induced mouse liver tumour formation. Male CD-1 mice were fed diets to provide intakes of 0 (control), 30, 100 and 300 mg/kg/day PBO and for purposes of comparison 500 ppm sodium phenobarbital (NaPB), a known constitutive androstane receptor (CAR) activator, for 7 and 14 days. Treatment with 100 and 300 mg/kg/day PBO and 500 ppm NaPB increased relative liver weight which was associated with hepatocyte hypertrophy, with hepatocyte replicative DNA synthesis (RDS) being increased after 7 days treatment. The treatment of CD-1 mice with 30−300 mg/kg/day PBO for 14 days resulted in significant dose-dependent increases in hepatic microsomal cytochrome P450 (CYP) content and 7-pentoxyresorufin O-depentylase (PROD) activity and in hepatic Cyp2b10 mRNA levels. In contrast, PBO produced a biphasic effect on markers of activation of the peroxisome proliferator-activated receptor alpha (PPARα), with small increases in microsomal lauric acid 12-hydroxylase activity and hepatic Cyp4a10 mRNA levels being observed in mice given 100 mg/kg/day with PBO, with either no increase or a significant inhibition being observed in mice given 300 mg/kg/day PBO. The hepatic effects of PBO in male CD-1 mice were generally similar to those produced by NaPB and were reversible after the cessation of treatment for 28 days. Studies were also performed in male C57BL/6J (wild type) mice and in hepatic CAR and pregnane X receptor (PXR) knockout mice (CAR KO/PXR KO mice), where in the CAR KO/PXR KO mice PBO had little effect on markers of CAR activation, but produced some increases in markers of PPARα activation. The treatment of male CD-1 mouse hepatocytes for 4 days with 5−50 μM PBO, 10−1000 μM NaPB and 25 ng/mL epidermal growth factor (EGF) resulted in significant increases in hepatocyte RDS. While treatment of hepatocytes from one male and one female human donor with 5−500 μM PBO and 10−1000 μM NaPB for 4 days had no effect on hepatocyte RDS, treatment with EGF resulted in significant increases in RDS in both human hepatocyte preparations. In summary, PBO is predominantly a hepatic CAR activator at carcinogenic dose levels in CD-1 mice, with activation of hepatic CAR resulting in a suppression of the effect of PBO on hepatic PPARα. A robust MOA for PBO-induced mouse liver tumour formation has been established, this MOA being similar to that previously identified for NaPB and some other rodent liver CAR activators. Based on the lack of effect of PBO on RDS in human hepatocytes, it is considered that the MOA for PBO-induced mouse liver tumour formation is qualitatively not plausible for humans.

Trigonella stellata reduced the deleterious effects of diabetes mellitus through alleviation of oxidative stress, antioxidant- and drug-metabolizing enzymes activities

Ethnopharmacological relevanceGenus Trigonella has a history of folkloric medicinal uses in China, Japan, Egypt and India. There are a variety of therapeutic actions of Trigonella including hypocholesterolemia, hypoglycemia, antibacterial, antiviral, anti-inflammatory activities, antioxidants and appetite stimulant. Aim of the studyThe prevalence of diabetes mellitus is increasing annually. The present study aims at investigating the protective effects of Trigonella stellata against the adverse effects of diabetes mellitus through investigation of the changes in phase I & II drug-metabolizing enzyme activities, protein expression of cytochrome P450 isoenzymes [CYP2E1 & 3A4], oxidative stress, antioxidant enzymes as well as histopathology of both liver and kidney tissues. MethodsGC-MS and MALDI-TOF were used to analyze the main constituents of the aqueous and the ethanolic extract of Trigonella stellata. Western immunoblotting technique used to show the protein expression of CYP450 isozymes in different groups. Spectrophotometric- and fluorophotometric techniques were also used for assessment of different hepatic integrity enzymes. Histopathological techniques used to illustrate the changes in the tissues of both livers and kidneys after different treatments. ResultsTrigonelline was found to be the main constituent of both aqueous and ethanolic extract of Trigonella stellata. Administration of the aqueous and/or the ethanolic extracts of Trigonella stellata to the diabetic rats was found to decrease the blood glucose level, the biochemical markers of both liver (transaminases activities, Lactate dehydrogenase, gamma-glutamyl transferase) and the renal functions (urea, creatinine and bilirubin) which were increased in diabetic-treated rats relative to their normal levels. Diabetes mellitus potentially induced the oxidative stress, and also activities of dimethylnitrosamine N-demethylase I, cytochrome c-reductase, ethoxyresourfin O-deethylase, and the total hepatic content of cytochrome P450. On the other hand, the activity of catalase [CAT], superoxide dismutase [SOD], glutathione S-transferase [GST], glutathione reductase [GR], glutathione peroxidase [GPx] and levels of reduced glutathione [GSH] were potentially inhibited in diabetic rats compared to the control rats. However, treatments of diabetic rats with either aqueous and/or ethanolic extracts of Trigonella stellata restored such changes caused by diabetes almost nearly to their normal levels compared to the control group. Supporting the activity of dimethyl nitrosamine N-demethylase I activity, the protein expression of CYP2E1 was also induced in diabetic rats. However, the aqueous extract of Trigonella stellata was more effective than ethanolic extract in restoring the changes in the protein expression of CYP2E. On the other hand, the protein expression of CYP3A4 was markedly decreased in diabetic rats, and this decrease was partially restored to its normal level after treatment of diabetic rats with aqueous and/or ethanolic extracts. In addition, Trigonella stellata extracts alleviated the histopathological changes in livers and kidneys caused by diabetes mellitus. ConclusionIt is concluded that diabetes mellitus induced changes in oxidative stress, phase I & II drug-metabolizing enzymes, and antioxidant enzymes activities, whereas both extracts of Trigonella stellata alleviated such changes. Alterations in cytochrome P450 system should be considered when therapeutic drugs are administered to diabetic patients since most of xenobiotic are mainly metabolized by this system.

Metazachlor: Mode of action analysis for rat liver tumour formation and human relevance

In a 2-year study the herbicide metazachlor (BAS 479H) was shown to significantly increase the incidence of liver tumours in female Wistar rats at a dietary level of 8000 ppm. As metazachlor is not a genotoxic agent, a series of in vivo and in vitro investigative studies were undertaken to elucidate the mode of action (MOA) for metazachlor-induced female rat liver tumour formation. Male and female Wistar rats were given diets containing 0 (control), 200 and 8000 ppm metazachlor for 3, 7, 14 and 28 days. The treatment of male rats with 200 and 8000 ppm metazachlor and female rats with 8000 ppm metazachlor resulted in significant increases in relative liver weight, which was associated with a centrilobular hepatocyte hypertrophy. Hepatocyte replicative DNA synthesis (RDS) was significantly increased in male rats given 8000 ppm metazachlor for 3 and 7 days and in female rats given 200 ppm metazachlor for 7–28 days and 8000 ppm metazachlor for 3–28 days. Significant increases in relative liver weight, centrilobular hepatocyte hypertrophy and hepatocyte RDS were also observed in male and female Wistar rats given and 500 ppm sodium phenobarbital (NaPB) for 3–28 days. The treatment of female Wistar rats with either 8000 ppm metazachlor for 7 days or with 500 ppm NaPB for 3 and 7 days resulted in the nuclear translocation of the hepatic constitutive androstane receptor (CAR). Treatment of male and female Wistar rats with 8000 ppm metazachlor for 14 days resulted in significant increases in hepatic microsomal total cytochrome P450 (CYP) content, CYP2B subfamily-dependent enzyme activities and mRNA levels, together with some increases in CYP3A enzyme activity and mRNA levels. The treatment of male Wistar rat hepatocytes with metazachlor (concentration range 0.5–50 μM) and NaPB (500 μM) for 4 days resulted in increased CYP2B enzyme activities and mRNA levels; with metazachlor and NaPB also producing significant increases in hepatocyte RDS levels. Studies were also performed with hepatocytes from male Sprague-Dawley wild type (WT) rats and CAR knockout (CAR KO) rats. While both treatment with metazachlor and NaPB for 4 days increased CYP2B enzyme activities and mRNA levels in WT rat hepatocytes, only minor effects were observed in CAR KO rat hepatocytes. Treatment with both metazachlor and NaPB only increased RDS in WT but not in CAR KO rat hepatocytes. The treatment of hepatocytes from two male human donors with 0.5–25 μM metazachlor or 500 μM NaPB for 4 days resulted in increases in CYP2B6 and CYP3A4 mRNA levels but had no effect on hepatocyte RDS. EGF as concurrently used positive control demonstrated the expected RDS response in all rat and human hepatocyte cultures. In conclusion, a series of in vivo and in vitro investigative studies have demonstrated that metazachlor is a CAR activator in rat liver, with similar properties to the prototypical CAR activator phenobarbital. A robust MOA for metazachlor-induced female rat liver tumour formation has been established. Based on the lack of effect of metazachlor on RDS in human hepatocytes, it is considered that the MOA for metazachlor-induced rat liver tumour formation is qualitatively not plausible for humans.

The role of microsomal oxidation in the regulation of monoamine oxidase activity in the brain and liver of rats

It has been shown in our previous study that monoamine oxidase (MAO) activity in different brain regions are correlated with a microsomal oxidation phenotype. The data obtained in this study, using the microsomal oxidation inhibitor SKF525, and using animals with different duration of hexobarbital sleep, has shown that increased intensity of microsomal oxidation might be associated with increased MAO activity. Since the rats with short hexobarbital sleep time had higher content of hepatic microsomal cytochrome P450 than did rats with long hexobarbital sleep time. In addition, the rats with higher hepatic content of CYP450 had higher activities of MAO-A and MAO-B. Moreover, the microsomal oxidation inhibitor SKF-525 reduced brain and liver activities of MAOA and MAO-B. Consequently, MAO activities in a brain and a liver depend on the microsomal oxidation process.

The effects of P-gp and CYP450 modulated by rifampicin on the steroid-induced osteonecrosis of the femoral head.

This study investigated the effects of rifampicin-modulated P-glycoprotein (P-gp) and cytochrome P450 (CYP450) activity on the development of steroid-induced osteonecrosis of the femoral head. Thirty-two rabbits were equally divided into four groups: control group, oral administration group, intramuscular injection group, and local release group, in which rifampicin-loaded artificial bone graft was implanted in the left femur cavity and blank bone graft was implanted in the right femur cavity. Dexamethasone was given 1week after rifampicin administration. Peripheral P-gp activity and hepatic CYP450 content were investigated 4weeks later. Hematoxylin and eosin, Massson, and tetracycline-fluorescence staining of the femoral head were compared. In vitro, the effects of intracellular dexamethasone concentration modulated by P-gp on osteoprotegerin (OPG)/receptor activator of nuclear factor κB ligand (RANKL) expression and differentiation of mesenchymal stem cells were further investigated. Peripheral P-gp activity and hepatic CYP450 content in the oral administration group and the intramuscular injection group were significantly higher than those in the local release group. P-gp activity of mesenchymal stem cells in rifampicin-implanted femoral head was significantly higher than that in the blank control. Histological study showed that rifampicin could prevent steroid-induced bone loss and lipid formation, and promote new bone formation and maturation. In vitro study confirmed that intracellular dexamethasone concentration modulated by P-gp could influence the OPG/RANKL ratio and the differentiation of mesenchymal stem cells. Enhanced levels of peripheral P-gp and hepatic CYP450 can reduce the incidence of steroid-induced osteonecrosis of the femoral head. P-gp activity locally enhanced by rifampicin decreases the intracellular steroid concentration, but rifampicin does not have significant effects on peripheral P-gp and hepatic CYP450.

Duration of hexobarbital-induced sleep and monoamine oxidase activities in rat brain: Focus on the behavioral activity and on the free-radical oxidation.

The present study is focused on the relationship between monoamine oxidase (MAO) activity and hepatic content of cytochrome P450 (CYP), which reflects the status of microsomal oxidation. For vital integrative evaluation of hepatic microsomal oxidation in rats, the hexobarbital sleep test was used, and content of CYP was measured in hepatic microsomes. Rats with short hexobarbital sleep time (SHST) had higher content of microsomal CYP than rats with long hexobarbital sleep time (LHST). Whole brain MAO-A and MAO-B activities, serotonin and carbonylated protein levels were higher in SHST than in LHST rats. MAO-A and MAO-B activities were higher in brain cortex of SHST rats; MAO-A activity was higher only in hypothalamus and medulla of LHST. The same brain regions of LHST rats had higher concentrations of carbonylated proteins and lipid peroxidation products than in SHST rats. MAO activity was correlated with microsomal oxidation phenotype. Rats with higher hepatic content of CYP had higher activities of MAO-A and MAO-B in the brain and higher plasma serotonin levels than rats with lower microsomal oxidation. In conclusion, data obtained in this study showed a correlation between MAO activity and microsomal oxidation phenotype.

Role of Kupffer cells in ischemic injury in alcoholic fatty liver

BackgroundThis study was designed to evaluate the role of Kupffer cells (KCs) in hepatic drug metabolizing dysfunction after hepatic ischemia–reperfusion (IR) in alcoholic fatty liver. Materials and methodsRats were fed the Lieber-DeCarli diet for 5 wk to develop alcoholic fatty liver, then were subjected to 90 min of hepatic ischemia and 5 h of reperfusion. For ablation of KCs, rats were pretreated with gadolinium chloride (GdCl3) 48 and 24 h before the IR procedure. ResultsAfter the IR procedure, ethanol diet (ED)-fed rats had higher serum aminotransferase activity compared with the control diet-fed rats. These changes were attenuated by GdCl3. The ED-fed rats exhibited increased hepatic microsomal total cytochrome P450 (CYP) content and nicotinamide adenine dinucleotide phosphate-CYP reductase and CYP1A1, 1A2, 2B1, and 2E1 isozyme activity. After hepatic IR, these increases were reduced to lower levels than observed in the sham group, except CYP2E1 activity. Increases in CYP2E1 activity and its expression were augmented after hepatic IR in ED-fed animals, but were attenuated by GdCl3. Finally, toll-like receptor 4 and myeloid differentiation primary response gene 88 protein expression, nuclear translocation of nuclear factor-κB and activator protein 1, and levels of proinflammatory mediators were further increased in ED-fed animals compared with control diet-fed animals after IR. These increases were attenuated by GdCl3. ConclusionsWe suggest that KCs contribute to hepatic drug metabolizing dysfunction during hepatic IR in alcoholic fatty liver via the toll-like receptors 4-mediated inflammatory response.

Impact of age on hepatic cytochrome P450 of domestic male Camborough-29 pigs.

Swine is not only an important species in veterinary medicine research but also a popular animal model for human drug discovery. It is valuable to understand the impact of pig age on abundance and activity of porcine hepatic cytochrome P450 (CYP450). Liver microsomes were prepared from Camborough-29 intact male pigs at the age of 1day and 2weeks and the castrated male pigs at the age of 5, 10, and 20weeks. Hepatic CYP450 content in the liver microsomes was measured using a UV/visible spectroscopic method. The activities of CYP450s were evaluated by metabolism of phenacetin, coumarin, tolbutamide, bufuralol, chlorzoxazone, and midazolam. The porcine hepatic CYP450 content increased with age with a plateau between age 2 and 5weeks. Activities of all CYP450 enzymes increased with age of pigs too. The bufuralol 1'-hydroxylase showed the highest hepatic activities compared with other CYP enzymes at all ages of pigs. The average activities at the age of 20weeks were about five times higher than those at the age of 5weeks for most of the CYP enzymes. With compensation of the ratio of liver to body weights, the overall CYP450 metabolism capability of the pigs may be peaked around ages of 10 to 20weeks. Those findings suggest that metabolism can be significantly different in growing phase of pigs and that the age may be an important factor in porcine medicine evaluation and pig model development.

Ascorbic acid deficiency decreases hepatic cytochrome P-450, especially CYP2B1/2B2, and simultaneously induces heme oxygenase-1 gene expression in scurvy-prone ODS rats.

The mechanisms underlying the decrease in hepatic cytochrome P-450 (CYP) content in ascorbic acid deficiency was investigated in scurvy-prone ODS rats. First, male ODS rats were fed a diet containing sufficient ascorbic acid (control) or a diet without ascorbic acid (deficient) for 18 days, with or without the intraperitoneal injection of phenobarbital. Ascorbic acid deficiency decreased hepatic microsomal total CYP content, CYP2B1/2B2 protein, and mitochondrial cytochrome oxidase (COX) complex IV subunit I protein, and simultaneously increased heme oxygenase-1 protein in microsomes and mitochondria. Next, heme oxygenase-1 inducers, that is lipopolysaccharide and hemin, were administered to phenobaribital-treated ODS rats fed sufficient ascorbic acid. The administration of these inducers decreased hepatic microsomal total CYP content, CYP2B1/2B2 protein, and mitochondrial COX complex IV subunit I protein. These results suggested that the stimulation of hepatic heme oxygenase-1 expression by ascorbic acid deficiency caused the decrease in CYP content in liver.

Role of hepatic microsomal oxidation in regulation of monoamine oxidase mediated processes in rat brain (727.1)

Monoamine oxidase (MAO) plays a key role in metabolism of biogenic amines and serves as a marker of alcoholism and depressive disorders. Various isoforms of cytochrome This study was focused on the relationship between MAO activity and hepatic content of cytochrome P450 which reflects the state of microsomal oxidation. Methods. For vital integrative evaluation of hepatic microsomal oxidation in rats, the hexobarbital sleep test was used. Content of cytochrome P450 was measured in hepatic microsomes. Activities of MAO‐A and MAO‐B were measured in the whole brain and brain structures. Content of serotonin, a MAO substrate was measured in the brain and plasma. Intensity of oxidative stress was evaluated by measuring lipid peroxidation products and carbonylated proteins in brain. Results. Rats with short hexibarbital sleep time (SHST) had higher content of microsomal cytochrome P450 than rats with long hexobarbital sleep time (LHST). Whole brain MAO‐A and MAO‐B activities, serotonin and carbonylated protein levels were higher in SHST than LHST rats. MAO‐A and MAO‐B activities were higher in brain cortex of SHST rats; only MAO‐A activity was higher in hypothalamus and medulla of LHST. The same brain regions of LHST rats had higher concentrations of carbonylated proteins and lipid peroxidation products. Conclusion. Data obtained in this study showed a correlation between MAO activity and microsomal oxidation phenotype. Rats with higher hepatic content of cytochrome P450 had higher activities of MAO‐A and MAO‐B in the brain and higher plasma serotonin levels than rats with lower microsomal oxidation. These correlations may reflect contribution of biogenic amines to regulation of cytochrome P450‐dependent monooxygenases and a role of cytochrome P450 in metabolism of biogenic amines and hormones regulating MAO activity including glucocorticoidsGrant Funding Source: Russian Foundation for Basic Research grant 11‐04‐01378‐а&amp;14‐04‐01381‐a

Growth hormone-independent suppression of growth hormone-dependent female isoforms of cytochrome P450 by the somatostatin analog octreotide

Octreotide is a potent somatostatin analog therapeutically used to treat several conditions including hyper growth hormone secretion in patients with acromegaly. We infused octreotide into female Sprague Dawley rats every 12h for 6 days at levels considerably greater than typical human therapeutic doses. Resulting circulating growth hormone profiles were characterized by ∼25% reduction in plasma levels, including both pulse and interpulse components, but still contained in an otherwise female-like “continuous” secretory profile. The normally elevated feminine expression levels (protein and/or mRNA) of CYP2C12, CYP2A1, CYP2C7 and insulin-like growth factor-1 (IGF-1), all dependent on the continuous feminine growth hormone profile, were dramatically down-regulated. Octreotide suppression of the female-dependent levels of CYPs (cytochromes P450) and IGF-1 could not be explained by the apparently inconsequential alterations in the feminine circulating growth hormone profile. In this regard, somatostatin and its analogs are known to have a myriad of extra-pituitary actions effecting nearly all tissues in the body. Focusing our attention on CYP2C12, accounting for >40% of the total hepatic cytochrome P450 content in the female rat liver, we found a ∼4-fold increase in hepatic ubiquitin-CYP2C12 levels in octreotide treated rats suggesting a possible contributing factor for the >60% suppression of CYP2C12 protein concentrations.

Effect of age on hepatic cytochrome P450 of Ross 708 broiler chickens

Age has significant impact on hepatic cytochrome P450 (CYP450) systems in animals. Ross 708 broiler chicken is a breed of chicken with fast growth characteristics. Cytochrome P450 in the livers of Ross 708 broiler chicken of different ages has been investigated. The birds were raised under standard husbandry conditions. A certain number of chickens was randomly sampled weekly for liver collection from d 1 to 56 posthatch. The chicken body and liver weights were recorded. The chicken livers were processed for liver microsomes though a multiple-step procedure at low temperature. Total CYP450 content in chicken liver homogenates and liver microsomes was measured using a UV/visible spectroscopic method. The enzymatic activities of CYP450 in the chicken liver microsomes were determined through incubation of CYP450 isoform substrates followed by measurement of formation of their metabolites. The chicken showed an opposite age pattern in hepatic CYP450 content and activities compared with most mammals. The hepatic CYP450 content and activities of chicken at d 1 posthatch were higher than at other ages. The total hepatic CYP450 content in chickens at d 1 posthatch was more than twice the average hepatic value of the chickens at d 7 to 28. This high CYP450 fell quickly in the first week posthatch and slightly rose from d 28 to 56. Hepatic CYP450 activities of CYP1A, 3A, 2C, 2D, and 2H were much higher in the chicken at d 1 posthatch. The differences of these enzymatic activities between d 1 and other ages of chicken were CYP450 isoform dependent. This result suggests that embryonic development of chicken livers has a significant impact on the age profile of hepatic CYP450 content and activities of posthatch chickens.

Cytochrome P450 2B6: function, genetics, and clinical relevance

Cytochrome P450 (CYP) 2B6 belongs to the set of important hepatic drug-metabolizing CYPs. It makes up roughly 3%-6% of total hepatic CYP content and metabolizes several pharmaceuticals including bupropion, efavirenz, cyclophosphamide, pethidine, ketamine and propofol. The enzyme is susceptible to drug-drug interactions by enzyme induction and inhibition. In addition to drugs, CYP2B6 is able to both detoxify and bioactivate a number of procarcinogens and environmental agents including pesticides and herbicides. There is an extensive interindividual variability in the expression of CYP2B6, which is in part explained by extensive genetic polymorphism. CYP2B6 is one of the most polymorphic CYP genes in humans with over 100 described SNPs, numerous complex haplotypes and distinct ethnic and racial frequencies. This review summarizes the basic properties of CYP2B6 and the main characteristics of clinical relevance.

Noncanonical suppression of GH-dependent isoforms of cytochrome P450 by the somatostatin analog octreotide

Octreotide is a potent somatostatin analog therapeutically used to treat several conditions including hyper GH secretion in patients with acromegaly. We infused, over 30 s, octreotide into male rats every 12 h for 6 days at levels considerably greater than typical human therapeutic doses. Unexpectedly, resulting circulating GH profile was characterized by pulses of higher amplitudes, longer durations, and greater total content than normal, but still contained an otherwise male-like episodic secretory profiles. In apparent disaccord, the normally elevated masculine expression levels (protein and/or mRNA) of CYP2C11 (accounting for >50% of the total hepatic cytochrome P450 content), CYP3A2, CYP2C7, and IGF1, dependent on the episodic GH profile, were considerably downregulated. We explain this contradiction by proposing that the requisite minimal GH-devoid interpulse durations in the masculine profile that solely regulate expression of at least CYP2C11 and IGF1 may be sufficiently reduced to suppress transcription of the hepatic genes. Alternatively, we observed that octreotide infusion may have acted directly on the hepatocytes to induce expression of immune response factors postulated to suppress CYP transcription and/or upregulate expression of several negative regulators (e.g. phosphatases and SOCS proteins) of the JAK2/STAT5B signaling pathway that normally mediates the upregulation of CYP2C11 and IGF1 by the masculine episodic GH profile.

Effect of Schistosoma mansoni infection and treatment on drug metabolizing enzymes.

The effect of Schistosoma mansoni infection on drug-metabolizing enzymes was investigated before and after treatment of S. mansoni-infected male albino mice with the antibilharzial drug praziquantel (CAS 55268-74-1). The drug was given in a total dose of 1000 mg/kg, and was administered at 500 mg/kg of body weight for two consecutive days each of 500 mg/kg of body weight. The drug was given 49 days after infection with 80 Egyptian strains of S. mansoni cercariae/mouse. The hepatic content of cytochrome P-450, cytochrome b-5 and NADPH cytochrome P-450 reductase were determined at 4, 8, 24, 72 h, one and two weeks after the second dose of treatment. The enzymes were determined in the microsomal fraction after homogenization and ultracentrifugation at 105,000 x g. The results indicate a marked decrease of most enzymes in the infected groups compared to normal controls. Two weeks after treatment there was an improvement of the level of most enzymes towards the normal values. The levels of liver function tests were also improved. Concerning the oogram pattern, there was a rapid change even after 8 h after the second dose of treatment. It was concluded that the antibilharzial drug could be considered as a safe drug with a rapid onset of action.

Coleus forskohlii extract induces hepatic cytochrome P450 enzymes in mice

Coleus forskohlii root extract (CFE) is popular for use as a weight loss dietary supplement. In this study, the influence of standardized CFE containing 10% active component forskolin on the hepatic drug metabolizing system was investigated to evaluate the safety through its drug interaction potential. Male ICR mice were fed AIN93G-based diets containing 0–5% CFE or 0.05% pure forskolin for 2–3weeks. Intake of two different sources of 0.5% CFE significantly increased the relative liver weight, total content of hepatic cytochrome P450 (CYP) and induced CYPs (especially 2B, 2C, 3A types) and glutathione S-transferase (GST) activities. CFE significantly increased mRNA expression of CYPs and GST with dose related responses. However, unlike the CFE, intake of 0.05% pure forskolin was found to be associated with only weak induction in CYP3A and GST activities with no significant increases in relative liver weight, total hepatic content or other CYPs activities. The inductions of CYPs and GST by CFE were observed at 1week of feeding and rapidly recovered by discontinuation of CFE. These results indicated the induction potential of CFE on CYPs, and that this effect was predominantly due to other, as yet unidentified constituents, and not forskolin contained in CFE.

Intrasplenic Transplantation of Isolated Adult Rat Hepatocytes

Adult male and female rat hepatocytes were individually transplanted into the spleens of adult male and female rats. The recipients were euthanized at either eight, sixteen, thirty, or forty-five weeks following transplantation, at which time hepatic and splenic levels of liver-specific rat albumin mRNA as well as sex-dependent transcript levels of CYP2C11, -2C12, -2C7, -2A1, and -3A2-which accounts for > 60% of the total concentration of hepatic constituent cytochrome P450-were determined. Whereas the pre-infused hepatocytes expressed their expected cytochrome P450 sexual dimorphisms (female-specific CYP2C12, male-specific CYP3A2, and female-predominant CYP2A1), their post-transplantational competence now reflected the sexual dimorphisms of the recipient (as observed in the host's liver), which supports the concept that the sex-dependent growth hormone circulating profiles are the determinants regulating the expression levels of hepatic cytochrome P450. Also expressed at normal concentrations in the pre-infused hepatocytes, male-specific CYP2C11 and female-predominant CYP2C7 were inexplicably undetectable in the spleens of both recipient males and females, regardless of the sex of the donor hepatocytes, almost one year after transplantation.

Repeated preexposure or coexposure to arsenic differentially alters acetaminophen‐induced oxidative stress in rat kidney

Acetaminophen (AP) is a widely used, cheap, and over-the-counter nonsteroidal anti-inflammatory drug. Its toxicity depends on the cytochrome P-450 (CYP)-mediated oxidation to the toxic metabolite N-acetyl-p-benzoquinoneimine. On the other hand, arsenic, a global groundwater and environmental contaminant of major public health concern, decreases hepatic CYP content and its dependent monoxygenase activities. We hypothesized that arsenic exposure would reduce the AP toxicity. Our aim was to evaluate the effects of repeated preexposure or coexposure to arsenic on the oxidative stress induced by a single or repeated oral administration of AP in rat kidney and its possible relationship with the effects of arsenic on certain antioxidants. Rats were exposed to arsenic through drinking water at 25 ppm for 28 days. The dosages of AP used for a single administration after arsenic preexposure for 28 days were 420 and 1000 mg kg(-1) , while for daily concurrent administration with arsenic for 28 days were 105 and 420 mg kg(-1) body weight. AP increased lipid peroxidation (LPO) in rat kidney where its acute administration caused more LPO than its subacute dosing. Repeated arsenic exposure differentially altered the AP-induced LPO. Arsenic preexposure antagonized LPO induced by the acute AP administration; in contrast, arsenic coexposure aggravated the repeated dose (AP)-mediated LPO. Arsenic-mediated alterations in renal sensitivity to LPO did not appear to be linked to the antioxidants such as reduced glutathione, superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase; nor could it be related to glutathione-S-transferase activity. The results indicated that repeated arsenic preexposure decreased susceptibility of rat kidney to acute AP-mediated oxidative stress; on the contrary, its coexposure rendered the rat kidney more vulnerable to oxidative stress induced by the repeated dosing of AP.