Total Hepatic Cytochrome P450 Content Research Articles

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.

CYP2B6: New Insights into a Historically Overlooked Cytochrome P450 Isozyme

Human CYP2B6 has been thought to account for a minor portion (<1%) of total hepatic cytochrome P450 (CYP) content and to have a minor function in human drug metabolism. Recent studies, however, indicate that the average relative contribution of CYP2B6 to total hepatic CYP content ranges from 2% to 10%. An increased interest in CYP2B6 research has been stimulated by the identification of an ever-increasing substrate list for this enzyme, polymorphic and ethnic variations in expression levels, and evidence for cross-regulation with CYP3A4, UGT1A1 and several hepatic drug transporters by the nuclear receptors pregnane X receptor and constitutive androstane receptor. Moreover, 20- to 250-fold interindividual variation in CYP2B6 expression has been demonstrated, presumably due to transcriptional regulation and polymorphisms. These individual differences may result in variable systemic exposure to drugs metabolized by CYP2B6, including the antineoplastics cyclophosphamide and ifosfamide, the antiretrovirals nevirapine and efavirenz, the anesthetics propofol and ketamine, the synthetic opioid methadone, and the anti-Parkinsonian selegiline. The potential clinical significance of CYP2B6 further enforces the need for a comprehensive review of this xenobiotic metabolizing enzyme. This communication summarizes recent advances in our understanding of this traditionally neglected enzyme and provides an overall picture of CYP2B6 with respect to expression, localization, substrate-specificity, inhibition, regulation, polymorphisms and clinical significance. Emphasis is given to nuclear receptor mediated transcriptional regulation, genetic polymorphisms, and their clinical significance.

Pomegranate Juice Effects on Cytochrome P450s Expression:In VivoStudies

Beneficial health effects have recently been claimed for pomegranate juice. In vitro and in vivo studies have demonstrated its anti-atherosclerotic capacity, chemoprevention and chemotherapy of prostate cancer, and antiproliferative, apoptotic, and antioxidant activity, among others. On the other hand, there is a complex interplay between tumor initiation, promotion, and progression and xenobiotic biotranformation. This led us to investigate the effect of pomegranate juice consumption on cytochrome P450 (CYP) activity and expression. For this purpose, male mice consumed this fruit juice for 4 weeks, and pentobarbital-induced sleeping time and total hepatic CYP content, activity, and expression were evaluated. Moreover, the activity of CYP isoform 2E1 and expression of the main CYP isoforms, namely, CYP1A1/2, CYP2E1, and CYP3A, were also assessed. It was found that pomegranate juice consumption decreased total hepatic CYP content as well as the expression of CYP1A2 and CYP3A. Prevention of procarcinogen activation through CYP activity/expression inhibition may be involved in pomegranate juice's effect on tumor initiation, promotion, and progression.

Modulation of Hepatic Cytochrome P450 during Listeria Monocytogenes Infection of the Brain

Hepatic cytochrome P450 enzymes can be modulated during systemic infections. Inflammatory responses in the brain have also been shown to cause a significant decrease in the levels and activities of important cytochrome P450 isoforms in the liver. We determined some of the effects of central nervous system (CNS) Listeria monocytogenes infection on hepatic cytochrome P450 systems in rats. Intracerebroventricular injection of L. monocytogenes resulted in a time-dependent modulation of CYP1A, CYP2B, and CYP3A activities in the liver. Total hepatic cytochrome P450 content was significantly lowered 48h after administration of the bacterium, and hepatic CYP1A and CYP2B activities were significantly altered 48 and 72h after infection, respectively, whereas CYP3A activity and protein content were depressed 72h after the insult. Bacterial load in the brain increased dramatically over a 72-h period, but the number of bacteria cultured from liver over this time period was relatively small. Therefore, an infection largely confined to the CNS in the rat results in abnormal activity levels of certain hepatic cytochrome P450 enzymes crucial in drug metabolism. If such a response also occurs in humans, this has the potential to produce serious complications with drug and endogenous substrate metabolism in patients with an infectious disease involving the CNS. © 2003 Wiley-Liss, Inc. and the American Pharmacists Association.

Effects of 5-Bromo-2'Deoxyuridine (BrdU) Implants on Hepatic Cytochrome P-450 Content and Beta-Oxidation Activity in Rats and Mice

ABSTRACTStandard regulatory toxicity tests are frequently supplemented with additional compound specific analysis. Analysis of hepatic cytochrome P-450 content, hepatic β-oxidation activity (biochemical analysis), and cell proliferation rates are examples of these analyses that are included when past experience or similarity to other compounds, suggest that a presently tested compound may have an effect. Until now, separate subsets of animals have been designated for cell proliferation analysis and biochemical analysis, because it was unknown if implantation of 5-bromo-2'deoxyuridine (BrdU) filled osmotic pumps (BrdU implants) would effect the rate of hepatic-β or hepatic cytochrome P-450 content.The purpose of the current study was to determine if BrdU implants had an effect on hepatic cytochrome P-450 content, β-oxidation activity, or the measurement of these enzymes in rats and mice. The BrdU was administered through subcutaneous osmotic pump implants.The rate of hepatic peroxisomal p-oxidation was not altered in male or female rats or mice with the BrdU implants when compared to those of the control groups. The total hepatic cytochrome P-450 content was also not altered in male or female rats or mice with the BrdU implants when compared to those of the control groups.BrdU implants do not appear to have an effect on the rate of hepatic peroxisomal β-oxidation or the total hepatic cytochrome P-450 content in male or female rats and mice. It can be concluded that in future studies, rats or mice which are designated for cell proliferation analysis using BrdU implants are also suitable for use in evaluating chemically induced effects on hepatic peroxisomal β-oxidation activity and/or total hepatic cytochrome P-450 content.

Modulation of cytochrome P450 by 5,5′-bis-trifluoromethyl-2,2′-dichlorobiphenyl, a unique environmental contaminant

5,5′-Bis-trifluoromethyl-2,2′-dichlorobiphenyl (5,5′CF 3-2,2′PCB) is representative of a unique class of trifluoromethyl polychlorinated biphenyls (CF 3-PCBs) found in sediments and fish of Lake Ontario, the Niagara river, and their tributaries. The potential hazard of 5,5′CF 3-2,2′PCB was assessed by exposing male Wistar rats to this agent in corn oil at a dose of 1 mg/kg/day or 75 mg/kg/day or corn oil alone (control) by oral intubation for 7 consecutive days. No lethality occurred during the course of exposure. A significant increase in liver weight and liver/body weight ratio and significant decrease in body weight gain were observed following exposure to the high dose of CF 3-PCB, relative to control. Exposure to the CF 3-PCB also resulted in a dose-related increase in the total hepatic cytochrome P450 content. This was associated with a dose-related increase in the O-deethylation of ethoxycoumarin, an activity which is mediated by several cytochrome P450s and thus provides a general representation of cytochrome P450 status. Specifically, a dose-dependent induction of the cytochrome P450s 1A1 and 1A2 (CYP1A1 and CYP1A2) proteins and their respective activities was observed, with significant induction occurring in both the low and high dose CF 3-PCB groups, compared to control. Additionally, CYP2B1 and CYP2B2 proteins and activities were induced following treatment with the high dose of 5,5′CF 3-2,2′PCB. Since, 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) and related compounds selectively induce CYP1A1 and CYP1A2, the results suggest that 5,5′CF 3-2,2′PCB has significant dioxin-like activity. Furthermore, 5,5′CF 3-2,2′PCB appears to be acting as a mixed-type inducer since phenobarbitallike induction of CYP2B1 and 2B2 was also associated with exposure.

Exercise training does not alter cytochrome P-450 content and microsomal metabolism

The purpose of this investigation was to determine whether increased endurance exercise capacity alters total hepatic cytochrome P-450 content and cytochrome P-450 (CYP1A and CYP2B) mediated hepatic microsomal mixed-function oxidase drug metabolism. Twenty adult male Sprague-Dawley rats were randomly assigned to either a control (C) or an endurance trained group (ET). ET rats were progressively trained 5 d.wk-1 for 11 wk. Both C and ET rats were administered in random order single posttraining doses of probe drugs theophylline (probe for CYP1A) and antipyrine (probe for CYP2B). Soleus muscle citrate synthase activity of ET rats was significantly greater (P < 0.01) than for C rats (mean +/- SD; C, 26.4 +/- 1.3 mumol.g-1.min-1; ET, 46.1 +/- 2.7). In contrast, total liver cytochrome P-450 content was not significantly different (P > 0.01) among C and ET rats (mean +/- SD; C, 0.554 +/- 0.055 nmol.mg-1 liver protein; ET, 0.604 +/- 0.080). Likewise, the posttraining C and ET single-sample plasma clearances of theophylline (mean +/- SD; C, 1.89 +/- 0.360 1.h-1.kg-1 total liver weight; ET, 2.08 +/- 0.49) and antipyrine (mean +/- SD; C, 6.44 +/- 1.56 1.h-1.kg-1 total liver weight; ET, 6.51 +/- 1.02) were not significantly different (P > 0.01). Therefore, it was concluded that strenuous endurance training of 11 wk duration did not alter total hepatic cytochrome P-450 content or CYP1A or CYP2B activity.

Hepatic microsomal alterations during Dipetalonema viteae infection in Mastomys natalensis

The multimammate rat, Mastomys natalensis was used as a model system to evaluate the chronic effects of infection by Dipetalonema viteae on hepatic mixed function oxidase activity. Total hepatic cytochrome P450 content and related total tissue mixed function oxidase activity were decreased to about 50% of control levels at patent phase of infection. The decrease in total tissue mixed function oxidase activity was due to a large decrease in cytochrome P450 concentration in the endoplasmic reticulum. Although the decrease in total liver monooxygenase activity in two substrates aniline and aminopyrine roughly paralleled the loss in cytochrome P450 content, several other microsomal enzyme markers not related to cytochrome P450 monooxygenation were elevated in proportion to total liver microsomal protein content. These results suggest that in M. natalensis during experimental filariasis, there is proliferation of hepatic cells with normal content of endoplasmic reticulum. Furthermore, there appears to be selective toxicity for hepatic cytochrome P450 and related monooxygenase activities. This may compromise the animal's ability to metabolize and dispose of other drugs to which the animals may be exposed in the course of infection.

Hepatic microsomal alterations during chronic trypanosomiasis in the field vole, Microtus montanus

The field vole, Microtus montanus, was used as a model system to evaluate the chronic effects of infection by Trypanosoma brucei gambiense on hepatic mixed-function oxidase activity. At day 28 post inoculation there was a 97% increase in liver wet weight per g body weight. A portion of the increase (21%) was accounted for by tissue edema which occurred after day 14 of infection. Total hepatic cytochrome P-450 content and related total tissue mixed-function oxidase activities were decreased to about 60% of control levels at day 28 post inoculation. The decrease in total tissue mixed-function oxidase activity was partly due to a small decrease in microsomal protein per cell, and partly to a large decrease in cytochrome P-450 concentration in the endoplasmic reticulum. Although the decrease in total liver monooxygenase activity in several substrates roughly paralleled the loss in cytochrome P-450 content, several other microsomal enzyme markers not related to cytochrome P-450 monooxygenation were elevated in proportion to total liver microsomal protein content. The results suggest that in M. montanus during trypanosomiasis, there is proliferation of hepatic cells with normal content of endoplasmic reticulum. Furthermore, there appears to be selective toxicity for hepatic cytochrome P-450 and related monooxygenase activities. This may compromise the animals' ability to metabolize and dispose of other drugs to which the animal may be exposed in the course of infection.

Paradoxical effects of cobaltous chloride treatment on 2-methylnaphthalene disposition and hepatic cytochrome P-450 content in carp.

Effects of cobaltous chloride (CoCl2) treatment on the disposition of 2-methylnaphthalene (2-MeNap)-derived radioactivity was studied in adult carp. The fish were treated with CoCl2 (60 mg/kg, ip) or an equivalent volume of saline (control) 36 and 12 hr before 2-MeNap administration (5 mg/kg, ip). Carp were killed 12, 24, 48 and 72 hr later. CoCl2-treated carp had less 2-MeNap-derived radioactivity in gallbladder bile and a greater amount in the liver at all of these times. Solvent extraction of bile revealed that virtually all of the radioactivity was due to polar 2-MeNap metabolites in both groups. Solvent extraction of liver showed that the greater total amount of 2-MeNap-derived radioactivity in the CoCl2 treatment group was due to a greater amount of parent compound being present; amount of polar 2-MeNap metabolites in the liver of CoCl2-treated carp and control carp was similar. Inasmuch as 2-MeNap metabolism in carp is thought to involve monooxygenation in the liver, our results suggest that the altered hepatobiliary disposition of 2-MeNap in CoCl2-treated carp may be due to depressed 2-MeNap metabolism by liver microsomal enzymes. However, total hepatic cytochrome P-450 content and activity of an associated monooxygenase enzyme (7-ethoxycoumarin-O-deethylase) were similar in CoCl2-treated and control carp throughout the study. To resolve this paradox (apparent depression of hepatic 2-MeNap metabolism by CoCl2 but lack of effect on hepatic cytochrome P-450 content and 7-ethoxycoumarin-O-deethylase activity), it is suggested that a minor form of cytochrome P-450 may be responsible for hepatic 2-MeNap metabolism in carp and be selectively inhibited or degraded by CoCl2. Inherent in this postulate is the assumption that this minor form of cytochrome P-450 does not contribute significantly to total cytochrome P-450 content of carp liver or 7-ethoxycoumarin-O-deethylase activity.

Genetic differences in the metabolism of carcinogens and in the binding of benzo[a]pyrene metabolites to DNA

1. 1.|Aromatic hydrocarbon “responsiveness,” or the Ah locus, controls in the mouse the induction of cytochrome P 1-450 and more than ten associated monoxygenase activities by polycyclic hydrocarbons. One of the induced activities is aryl hydrocarbon hydroxylase (EC 1.14.14.2), which metabolizes BP and other chemical carcinogens. 2. 2.|For certain substrates, cytochrome P 1-450 is known to produce a predominance of reactive intermediates and products that differ from those formed by other forms of cytochrome P-450. The aromatic hydrocarbon-treated responsive mouse therefore may produce, in various tissues throughout the body, quantitative and qualitative differences in reactive intermediates of a given test compound. 3. 3.|Numerous conditions in the mouse, which include an increased susceptibility to chemical carcinogenesis and drug toxicity, are therefore directly associated with the Ah locus — considered to be a single gene or a small number of genes. 4. 4.|The various forms of cytochrome P-450 in mouse liver or skin microsomes produce reactive metabolites of BP that bind to DNA in vitro. These results confirm the work of other laboratories in which rat liver microsomes or cell cultures were used. 5. 5.|Nine peaks, reflecting BP metabolites bound to DNA nucleosides, can be reproducibly identified from C3H, C57BL/6, and DBA/2 mice. When either the C3H or C57BL/6 strain is compared with the DBA/2 strain, a positive general correlation is found between the carcinogenic index in vivo and each of three determinations in vitro: the size of eight of the nine peaks, the hepatic aryl hydrocarbon hydroxylase activity, and the total hepatic cytochrome P-450 content. 6. 6.|Peaks D, E, H, and I demonstrate relative differences among the three strains that are in the same direction as the carcinogenic index in vivo, i.e. the peaks are greater in C3H than in C57BL/6 and both are considerably greater than those in DBA/2 mice. When C3H mice are compared with C57BL/6 mice, however, no quantitative correlation is found between the carcinogenic index and the size or shape of any of the nine peaks, the hepatic hydroxylase activity, or the total hepatic cytochrome P-450 content. Hence, the total quantity of nucleoside-BP metabolites in each of the nine peaks is not necessarily associated with biological activity (i.e. cancer in vivo). It is suggested that a defect in the general immune competence of the C3H inbred strain may be an important factor in its markedly increased susceptibility to BP-initiated tumorigenesis, compared with the C57BL/6 strain. 7. 7.|With hepatic microsomes, peak G is associated, at least in part, with nucleoside-metabolite complexes involving BP-4,5-oxide, whereas the other eight peaks are associated predominantly with metabolites formed by cytochrome P 1-450 rather than other forms of P-450. With skin microsomes, all peaks including peak G appear to be associated with metabolites formed predominantly by cytochrome P 1-450. Peak E is associated predominantly with BP-7 8-diol-9,10-epoxide bound to one or more nucleosides. This reactive intermediate, oxygenated in the non-K region, is associated with genetically mediated increases in aromatic hydrocarbon-inducible cytochrome P 1-450. 8. 8.|Cultured human lymphocytes and microsomes from a human liver biopsy are both capable of metabolizing BP to intermediates that bind to DNA and cause peaks similar to those found with mouse liver or skin microsomes. (Permission to reprint the previously published material — Figs. 3, 4, 5, 6, 7, 8, and 9 and Tables 1 and 2 — has been obtained from the publishing house indicated in the references indicated).