Down-regulation Of P450 Research Articles

Nano-silica and magnetized-silica mitigated lead toxicity: Their efficacy on bioaccumulation risk, performance, and apoptotic targeted genes in Nile tilapia (Oreochromis niloticus).

Contamination of aquatic systems with heavy metals (HM) is of great concern owing to their deleterious impact on living organism. The current research is focused on application of silica particles with new functionalized properties (magnetic silica; SiMag or Nanoporous silica; SiNPs) and their efficacy to mitigate lead (pb) toxicity in Nile tilapia. One thousand fingerlings were distributed: two control groups (negative; without pb toxicity (NC) positive (with pb toxicity) and other four groups received two silica sources (SiMag or SiNPs) with two levels (400 and 600mg/kg diet) for 56 days then exposed to pb for 30 days. Before toxicity exposure, maximum growth, and most improved feed conversion ratio and biochemical parameters were noticed with higher SiMag or SiNPs levels. Serum antioxidant enzymes and their transcriptional levels in muscle and liver were boosted in groups received SiMag or SiNPs. After toxicity exposure, hematological and antioxidants biomarkers maintained at adequate levels in SiMag or SiNPs. Prominent reduction of residual pb in gills, liver, kidney, and muscle was observed in SiNPs then SiMag groups. Interestingly, the maximum down-regulation of P450, caspase-3 and HSP-70 and MT were observed in groups received 600mg/kg diet of SiMag or SiNPs. The higher level of P53 in liver and gills was detected in PC, inversely reduced in SiMag or SiNPs. Severity of the histopathological alterations in examined organs greatly reduced in groups received SiMag or SiNPs, unlike it were induced in PC group. In conclusion, higher SiMag or SiNPs levels not only mitigate negatives impact of pb toxicity in fish but also ensure its safety for human consumption.

Downregulation of cytochrome P450 2C8 by 3-methylcholanthrene in human hepatocellular carcinoma cell lines.

The marked induction of cytochromes P450 such as CYP1A1 caused by polycyclic aromatic hydrocarbons (PAHs) like 3-methylcholanthrene (MC) is often accompanied by suppression of other hepatic P450s. The molecular mechanisms, functional consequences, and human relevance of P450 downregulation by PAHs are poorly understood. MC suppresses mRNA levels for CYP2C8, an important human P450, in cultured human hepatocytes. To avoid hepatocyte lot-to-lot variability, we assessed CYP2C8 regulation by MC in HepaRG cells, a terminally differentiated human hepatocellular carcinoma cell line that maintains high P450 expression. MC strongly induced CYP1A1 mRNA levels and markedly downregulated CYP2C8 mRNA levels in HepaRG cells. Although MC also suppressed CYP2C8 mRNA levels in the HepG2 human hepatocellular carcinoma cell line, basal CYP2C8 expression was extremely low. HepaRG cells appear to be an appropriate model system for studying the mechanisms and functional consequences of CYP2C8 downregulation by PAHs.

Virtual Experiments Enable Exploring and Challenging Explanatory Mechanisms of Immune-Mediated P450 Down-Regulation.

Hepatic cytochrome P450 levels are down-regulated during inflammatory disease states, which can cause changes in downstream drug metabolism and hepatotoxicity. Long-term, we seek sufficient new insight into P450-regulating mechanisms to correctly anticipate how an individual’s P450 expressions will respond when health and/or therapeutic interventions change. To date, improving explanatory mechanistic insight relies on knowledge gleaned from in vitro, in vivo, and clinical experiments augmented by case reports. We are working to improve that reality by developing means to undertake scientifically useful virtual experiments. So doing requires translating an accepted theory of immune system influence on P450 regulation into a computational model, and then challenging the model via in silico experiments. We build upon two existing agent-based models—an in silico hepatocyte culture and an in silico liver—capable of exploring and challenging concrete mechanistic hypotheses. We instantiate an in silico version of this hypothesis: in response to lipopolysaccharide, Kupffer cells down-regulate hepatic P450 levels via inflammatory cytokines, thus leading to a reduction in metabolic capacity. We achieve multiple in vitro and in vivo validation targets gathered from five wet-lab experiments, including a lipopolysaccharide-cytokine dose-response curve, time-course P450 down-regulation, and changes in several different measures of drug clearance spanning three drugs: acetaminophen, antipyrine, and chlorzoxazone. Along the way to achieving validation targets, various aspects of each model are falsified and subsequently refined. This iterative process of falsification-refinement-validation leads to biomimetic yet parsimonious mechanisms, which can provide explanatory insight into how, where, and when various features are generated. We argue that as models such as these are incrementally improved through multiple rounds of mechanistic falsification and validation, we will generate virtual systems that embody deeper credible, actionable, explanatory insight into immune system-drug metabolism interactions within individuals.

Microplate Assay Measurement of Small Amount of P450

Cytochrome P450 (P450) has the feature of having an absorption spectrum in the 450 nm if it binds with carbon monoxide (CO). A spectrophotometrical assay of the P450 content was established through the use of this feature. We improved the analytical methodology to deal with a small sample and to become suitable for a high-throughput measurement. A microplate-based assay using a CO releasing agent, dimanganese decacarbonyl (DMDC), and an exposure to light, instead of a CO bubbling was the process utilized. The absorbance difference between the sample which added DMDC and did not add DMDC was measured after adding a sodium hydrosulfite solution to each sample. The absorbance index was determined on the basis of the optical path length of the microplate well. The P450 content was calculated using the absorbance difference and this absorbance index. A suitable concentration of DMDC solution, lighting time and luminescence intensity were determined. The P450 density of rats liver microsomes determined by this new method had a good correlativity to the traditional method. Analysis of P450 content in the microsome of the liver and intestine are necessary as a predictor of a drug-interaction which are referred from induction or down-regulation of P450 at an early phase of development of functional foods or drugs. The new assay we propose was suitable for a small sample and high-throughput analysis of multiple samples, and suitable to determine influences exerted by a test sample which was administered to small animals such a rodent.

Modulation of Hepatic Cytochrome P450s byCitrobacter rodentiumInfection in Interleukin-6- and Interferon-γ-Null Mice

After infection with Citrobacter rodentium, murine hepatic cytochrome P450 (P450) mRNAs are selectively regulated. Several serum proinflammatory cytokines are elevated, the most abundant being interleukin-6 (IL6). To elucidate the role of cytokines in the regulation of P450s during infection, we orally infected wild-type, IL6(-/-), or interferon-γ(-/-) [IFNγ(-/-)] female C57BL/6J mice with C. rodentium and analyzed hepatic P450 expression 7 days later. The majority of P450 mRNAs were equally affected by infection in each genotype, indicating that IL6 and IFNγ are not the primary mediators of P450 down-regulation in this disease model. The down-regulation of CYP3A11 and CYP3A13 and induction of CYP2D9 mRNAs were attenuated in the IL6(-/-) mice, suggesting a role of IL6 in the regulation of only these P450s. Similar evidence implicated IFNγ in the regulation of CYP2D9, CYP2D22, CYP3A11, CYP3A25, and CYP4F18 mRNAs in C. rodentium infection and CYP2B9, CYP2D22, and CYP2E1 in the bacterial lipopolysaccharide model of inflammation. This is the first indication of an in vivo role for IFNγ in hepatic P450 regulation in disease states. The deficiency of IL6 or IFNγ affected serum levels of the other cytokines. Moreover, experiments in cultured hepatocytes demonstrated that tumor necrosis factor α (TNFα) is the most potent and efficacious of the cytokines tested in the regulation of murine P450 expression. It is therefore possible that part of the IFNγ(-/-) and IL6(-/-) phenotypes could be attributed to the reduced levels of TNFα and part of the IFNγ(-/-) phenotype could be caused by reduced levels of IL6.

Polychlorinated biphenyls have inhibitory effect on testicular steroidogenesis by downregulation of P45017α and P450scc

Polychlorinated biphenyls (PCBs) are environmental pollutants that are quite toxic to biological systems. This study examined the inhibitory effect of PCB126 and PCB114 on testicular steroidogenesis in male rats. Male Sprague Dawley rats received weekly intraperitoneal injections of PCB126 (0.2 mg/kg) or PCB114 (20 mg/kg) or vehicle (corn oil). Animals from each group were sacrificed at 2, 5 and 8 weeks after the injections. Blood and testis tissue samples were collected for the hormone assay, Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR). The testosterone, luteinizing hormone (LH) and follicle-stimulating hormone (FSH) levels were assayed, and the expression levels of the mRNA and proteins associated with the testosterone biosynthesis pathway were measured to determine the effect of PCB126 and PCB114 on testicular steroidogenesis. The results showed that the testis weight was significantly higher in the PCB126-treated rats given eight shots. Moreover, the serum testosterone levels were significantly lower in the PCB126 and PCB114-treated groups than the control. The transcription and translation levels of P450(17alpha) and P450(scc) were significantly lower in the PCB126-treated groups than the control. These results suggest that PCB126 may affect testicular steroidogenesis by downregulating P450(17alpha), P450(scc) and have inhibitory effect on the testicular functions.

Abstract A105: Dietary quercetin, quercetin-gene interaction, metabolic gene expression in lung tissue, and lung cancer risk

Abstract Background: Quercetin is a flavonol found in fruits (apple, grape) and vegetables (onion, broccoli). Quercetin possesses anticarcinogenic properties that protect against cancer. However, the epidemiological evidence on the association of quercetin-rich food intake with lung cancer risk is inconclusive, and how quercetin affects lung carcinogenesis in humans is largely unknown. Laboratory analyses have shown that quercetin may inhibit Phase I cytochrome P450s enzymes responsible for the activation of tobacco-related procarcinogens, and induce Phase II glutathione S-transferase (GST) enzymes responsible for the removal of activated carcinogens. Methods: We investigated the role of dietary quercetin and the interaction between quercetin and P450 and GST polymorphisms on lung cancer risk in incident lung cancer cases and matched controls from Environment and Genetics in Lung cancer Etiology (EAGLE), a population-based case-control study including 2100 primary lung cancer cases and 2120 controls. Dietary information and genotype data were available for 1740 cases and 1919 controls. In fresh-frozen non-tumor lung tissue from 38 adenocarcinoma patients, we also assessed the correlation between dietary quercetin intake and mRNA expression of tobacco-related Phase I and Phase II metabolic genes. Multivariable odds ratios (ORs) and 95% confidence intervals (CIs) for sex-specific quintile of intake were calculated using unconditional logistic regression, adjusted for putative risk factors. Two sample t-test was used to assess difference in expression by quercetin-rich consumption status. Results: Frequent intake of quercetin-rich foods was inversely associated with lung cancer risk (ORQ5vsQ1=0.49;95% CI=0.37–0.67; p-trend: <0.001), and did not differ by genotype, gender, or histological subtypes. The association was stronger in subjects who smoked more than 20 cigarettes per day (ORQ5vsQ1=0.35;95% CI=0.19–0.66; p-trend=0.003). The comparison between gene expression and high-versus-low consumption of quercetin-rich foods showed an overall down-regulation of P450 and up-regulation of GST metabolic genes (p-values<0.05) in lung tissue. Conclusions: Quercetin-rich diet was associated with lower risk of lung cancer and with down-regulation of Phase I P450 genes and up-regulation of Phase II GST genes in lung tissue. These findings suggest a possible mechanism for quercetin-related protection against lung cancer risk through quercetin-induced changes in the expression of genes involved in the metabolism of tobacco-carcinogens. Further studies exploring the interplay between quercetin intake, tobacco smoking, and lung cancer risk are warranted. Citation Information: Cancer Prev Res 2010;3(1 Suppl):A105.

Regulation of Hepatic Cytochrome P450 Expression in Mice with Intestinal or Systemic Infections ofCitrobacter rodentium

We reported previously that infection of C3H/HeOuJ (HeOu) mice with the murine intestinal pathogen Citrobacter rodentium caused a selective modulation of hepatic cytochrome P450 (P450) gene expression in the liver that was independent of the Toll-like receptor 4. However, HeOu mice are much more sensitive to the pathogenic effects of C. rodentium infection, and the P450 down-regulation was associated with significant morbidity in the animals. Here, we report that oral infection of C57BL/6 mice with C. rodentium, which produced only mild clinical signs and symptoms, produced very similar effects on hepatic P450 expression in this strain. As in HeOu mice, CYP4A mRNAs and proteins were among the most sensitive to down-regulation, whereas CYP4F18 was induced. CYP2D9 mRNA was also induced 8- to 9-fold in the C57BL/6 mice. The time course of P450 regulation followed that of colonic inflammation and bacterial colonization, peaking at 7 to 10 days after infection and returning to normal at 15 to 24 days as the infection resolved. These changes also correlated with the time course of significant elevations in the serum of the proinflammatory cytokines interleukin (IL)-6 and tumor necrosis factor-alpha, as well as of interferon-gamma and IL-2, with serum levels of IL-6 being markedly higher than those of the other cytokines. Intraperitoneal administration of C. rodentium produced a rapid down-regulation of P450 enzymes that was quantitatively and qualitatively different from that of oral infection, although CYP2D9 was induced in both models, suggesting that the effects of oral infection on the liver are not due to bacterial translocation.

Impaired Microsomal Oxidation of the Atypical Antipsychotic Agent Clozapine in Hepatic Steatosis

Hepatic lipid infiltration (steatosis) is a complication of the metabolic syndrome and can progress to nonalcoholic steatohepatitis and severe liver injury. Microsomal cytochrome P450 (P450) drug oxidases are down-regulated in experimental steatosis. In this study we evaluated the separate and combined effects of lipid accumulation and P450 down-regulation on the microsomal oxidation of the antipsychotic agent clozapine (CLZ), the use of which is associated with an increased incidence of the metabolic syndrome. Several important drug oxidizing P450s were down-regulated, and the formation of N-desmethyl-CLZ (norCLZ) and CLZ N-oxide was decreased in microsomal fractions from orotic acid-induced early steatotic rat liver. Inclusion of lipids extracted from steatotic, but not control, liver decreased the free concentration of CLZ in microsomes and suppressed norCLZ formation; CLZ N-oxidation was unchanged. Triglycerides increased in steatotic liver to 15-fold of control, whereas increases in the monounsaturated oleic acid to 10-fold of control and total polyunsaturated and saturated fatty acids to 4- and 5-fold of control also occurred. Addition of triglycerides containing esterified omega-6 and omega-3 fatty acids inhibited the microsomal formation of norCLZ but not that of CLZ N-oxide; triglycerides esterified with unsaturated and monounsaturated fatty acids were inactive. Thus, drug oxidation may be suppressed in steatosis by P450 down-regulation and the accumulation of polyunsaturated fatty esters. In contrast, the activity of the flavin-containing monooxygenase that mediates CLZ N-oxidation was unimpaired. Lipid deposition in livers of patients with the metabolic syndrome may necessitate dosage adjustments for toxic drugs, including CLZ.

HEPATIC AND RENAL CYTOCHROME P450 GENE REGULATION DURINGCITROBACTER RODENTIUMINFECTION IN WILD-TYPE AND TOLL-LIKE RECEPTOR 4 MUTANT MICE

Citrobacter rodentium is the rodent equivalent of human enteropathogenic Escherichia coli infection. This study investigated regulation of hepatic and renal cytochrome P450 (P450) mRNAs, hepatic P450 proteins, cytokines, and acute phase proteins during C. rodentium infection. Female C3H/HeOuJ (HeOu) and C3H/HeJ (HeJ) mice [which lack functional toll-like receptor 4 (TLR4)] were infected with C. rodentium by oral gavage and sacrificed 6 days later. Hepatic CYP4A10 and 4A14 mRNAs were decreased in HeOu mice (<4% of control). CYP3A11, 2C29, 4F14, and 4F15 mRNAs were reduced to 16 to 55% of control levels, whereas CYP2A5, 4F16, and 4F18 mRNAs were induced (180, 190, and 600% of control, respectively). The pattern of P450 regulation in HeJ mice was similar to that in HeOu mice for most P450s, with the exception of the TLR4 dependence of CYP4F15. Hepatic CYP2C, 3A, and 4A proteins in both groups were decreased, whereas CYP2E protein was not. Renal CYP4A10 and 4A14 mRNAs were significantly down-regulated in HeOu mice, whereas other P450s were unaffected. Most renal P450 mRNAs in infected HeJ mice were increased, notably CYP4A10, 4A14, 4F18, 2A5, and 3A13. Hepatic levels of interleukin (IL)-1beta, IL-6, and tumor necrosis factor alpha (TNFalpha) mRNAs were significantly increased in infected HeOu mice, whereas only TNFalpha mRNA was significantly increased in HeJ mice. Hepatic alpha1-acid glycoprotein was induced in both groups, whereas alpha-fibrinogen and angiotensinogen were unchanged. These data indicate that hepatic inflammation induced by C. rodentium infection is mainly TLR4-independent and suggest that hepatic P450 down-regulation in this model may be cytokine-mediated.

Hepatic Cytochrome P450 Gene Regulation during Endotoxin-Induced Inflammation in Nuclear Receptor Knockout Mice

Inflammatory agents such as lipopolysaccharide (LPS) down-regulate the hepatic expression of many cytochrome P450 (P450) mRNAs and proteins. Previous studies suggested that suppression of some P450 mRNAs could involve the regulation or modulation of the nuclear receptors peroxisome proliferator-activated receptor alpha (PPARalpha) or pregnane X receptor (PXR). To determine the involvement of these receptors in P450 down-regulation, PPARalpha knockout (KO), PXR KO, and appropriate wild-type (WT) mice were administered either saline or 1 mg/kg LPS. Hepatic mRNA and protein expression of several P450 isoforms, interleukin (IL)-1beta, IL-6, tumor necrosis factor (TNF) alpha, alpha1-acid glycoprotein (AGP), and fibrinogen (FBG) were examined 16 h later. LPS administration significantly decreased the hepatic expression of CYP1A2, 2A5, 2C29, 2E1, 3A11, 4A10, and 4A14 mRNAs in both groups of PPARalpha and PXR mice, whereas CYP3A13 mRNA was increased slightly in PPARalpha WT and KO mice, but not in PXR mice. Effects of LPS administration on mouse hepatic P450 proteins (probed using rat P450 2C, 3A, 4A, and 2E antibodies) were consistent with mRNA results in most cases. LPS treatment significantly increased IL-1beta, IL-6, TNFalpha, AGP, and FBG mRNA in both PPARalpha and PXR mice, with the greatest effect observed with TNFalpha. Because decreases in P450 mRNA expression were essentially identical in both WT and KO mice for both nuclear receptors, these data indicate that down-regulation of P450 during inflammation does not require the nuclear receptors PPARalpha and PXR.

Hepatic cytochrome P450 down-regulation during aseptic inflammation in the mouse is interleukin 6 dependent.

Expression of cytochromes P450 (CYP) is markedly reduced during inflammatory processes. In vitro studies with hepatocytes have shown that cytokines generated during these processes down-regulate CYP. However, it is not clear to what extent each individual cytokine contributes to the overall reduced expression of the various CYP isoenzymes in vivo. Interleukin 6 (IL-6), a major player during inflammatory processes, is recognized as the most important cytokine modulating the hepatic expression of acute-phase protein (APP) genes. For this reason, we selected the IL-6(-/-) mouse as a model to investigate the role of IL-6 in the down-regulation of hepatic CYP during experimental inflammation. Our results show that the reduction in messenger RNA (mRNA) levels of CYP1A2, CYP2A5, and CYP3A11 during turpentine-induced inflammation was abrogated in IL-6-deficient mice, confirming that IL-6 is an indispensable player for the down-regulation of hepatic CYP during aseptic inflammation. Moreover, the different CYP isoenzymes showed a variable grade of dependence on IL-6, CYP2A5 being the most sensitive one. In the case of CYP2E1, differences between IL-6(-/-) and wild-type mice were no longer maintained after 24 hours, suggesting a delayed, rather than abrogated, CYP down-regulation in the absence of IL-6. As opposed to that, hepatic CYP repression took place in IL-6-deficient mice during lipopolysaccharide (LPS)-mediated inflammation. This contrasting behavior observed for CYP is surprisingly similar to the one seen for extracellular (serum amyloid A, beta-fibrinogen) and intracellular (metallothionein-1) APPs and points to the fact that, in the model of bacterial inflammation (LPS), the effects of IL-6 on CYP down-regulation are likely to be substituted by other cytokines or mediators.

Down-regulation of the hepatic cytochrome P450 by an acute inflammatory reaction: implication of mediators in human and animal serum and in the liver.

1. Infection and inflammation trigger a cascade of mediators that eventually will down-regulate the hepatic cytochrome P450 (P450). The present study aimed to characterize the mediators contained in the serum of rabbits with an acute inflammatory reaction (AIR) induced by the s.c. injection of turpentine (5 ml), and in the serum of humans with an acute upper respiratory tract viral infection. 2. Hepatocytes from control (H(CONT)) rabbits and rabbits with an AIR (H(INFLA)) were isolated and cultured. Compared with H(CONT) in H(INFLA) the production of theophylline metabolites, 3-methylxanthine (3MX), 1-methyluric acid (1MU), and 1,3-dimethyluric acid (1,3DMU) was reduced as was the amount of total P450, while lipid peroxidation was increased. Incubation of H(INFLA) with serum of rabbits with an AIR (RS(INFLA)) for 4 h further reduced the formation of the metabolites of theophylline as well as the amount of P450, and enhanced the lipid peroxidation. RS(INFLA) obtained 6, 12 and 24 h after the injection of turpentine showed the same ability to down-regulate hepatic P450 as the serum obtained at 48 h. 3. The efficacy (Emax) of RS(INFLA) to inhibit the formation of theophylline metabolites differed, i.e. 1,3DMU > 1MU > 3MX, and the potency of serum mediators (IC50) was similar for 3MX and 1MU, but lower for 1,3DMU. 4. Incubation of serum of human volunteers (HS(INFLA)) with a viral infection with H(CONT) or H(INFLA) reduced the production of theophylline metabolites, as well as the amount of P450, and increased the lipid peroxidation. HS(INFLA) depressed 1,3DMU more efficiently than 3MX and 1MU. HS(INFLA) reduced 3MX with greater efficacy than did RS(INFLA). Potency was very variable but not different from rabbits. 5. It is concluded that the serum of rabbits with an AIR or of humans with a viral infection contain several mediators that inhibit noncompetitively various isoenzymes of the hepatic P450. The decrease in P450 induced by HS(INFLA) or RS(INFLA) is closely associated with the increase in lipid peroxidation (r2= 0.8870) suggesting that lipid peroxidation could directly or indirectly be involved in the P450 down-regulation.

Cytochrome P4501A1 and cytochrome P4501A2 are downregulated at both transcriptional and post-transcriptional levels by conditions resulting in interferon-alpha/beta induction.

The interferon mediated downregulation of constitutive and inducible cytochrome P450 enzymes occurs through a pretranslational mechanism which depresses the mRNA encoding cytochrome P450s. We measured the transcription rates of CYP1A genes and the turnover of CYP1A mRNA in rats treated with the interferon-alpha/beta inducer polyinosinic acid-polycytidylic acid. The rate of transcription of CYP1A1 and CYP1A2 genes was significantly decreased in hepatic nuclei isolated from male rats treated with polyinosinic acid-polycytidylic acid (10 mg/kg). In addition the rate of degradation of hepatic CYP1A1 and CYP1A2 mRNA was examined following the inhibition of de novo transcription by actinomycin D (1 mg/kg). Messenger RNA levels were analysed by Northern and slot blotting with a 1.2 kb murine CYP1A1 cDNA probe. Interferon significantly augmented the rate of loss of CYP1A1 and CYP1A2 mRNAs suggesting that post-transcriptional degradation of mRNA contributes to the pre-translational events that cause cytochrome P450 downregulation. These results support the involvement of both transcriptional and post-transcriptional mechanisms in the loss of cytochrome P450s mediated by interferon inducers.

Differential effect of pentoxifylline on lipopolysaccharide-induced downregulation of cytochrome p450

It is now established that inflammatory stimuli such as lipopolysaccharides (LPS) and polyinosinic acid:polycytidylic (polyIC) suppress hepatic expression of cytochrome P450 (P450) genes in rat liver. Previous studies have suggested that LPS- or polyIC-induced downregulation of P450 was due to endogenously released inflammatory cytokines such as tumor necrosis factor-a (TNF-α), interleukin-1, interleukin-6, and interferons (IFNs). To improve our understanding of the role of inflammatory cytokines in mediating P450 depression, we investigated the possibility of preventing P450 downregulation with pentoxifylline. Pentoxifylline has been shown to inhibit LPS-induced TNF-a production by suppression of TNF-a gene expression. The present study shows that in uninduced male rats pentoxifylline selectively prevents the downregulation of microsomal P4501A2 and P4502B caused by LPS. No protective effect of pentoxifylline on the downregulation of P4502E1 and P4503A1/2 was observed. PolylC-induced downregulation of P4501A2, P4502B, P4502E1, and P4503A1/2 was not affected by pentoxifylline. These results suggest that the LPS-induced downregulation of P4501A2 and P4502B is mediated to a large extent by TNF-α. Other cytokines might be involved in the suppression of P4502E1 and P4503A1/2. The fact that polylC-induced downregulation is not protected by pentoxifylline is further evidence that this agent acts via a selective induction of IFNs.

The role of nitric oxide generation in interferon-evoked cytochrome P450 down-regulation

The down-regulation of cytochrome P450 during host defence involves one or more mediators of unknown source and identity. Owing to striking similarities in the features that are involved in the production of NO and the features involved in the down-regulation of cytochrome P450 we hypothesized that NO could be a mediator in the interferon-evoked loss of cytochrome P450. In these experiments the nitric oxide synthase inhibitor l-nitroarginine (5 mg/kg) failed to protect mice from the down-regulation of cytochrome P450, ethoxyresorufin dealkylase and para-nitrophenol hydroxylase in the liver following administration of the interferon inducer polyinosinic.polycytidylic acid. A higher dose of L-nitroarginine (30 mg/kg) given every 12 h for 3 days also did not protect against cytochrome P450 losses. In addition, no down-regulation of the enzyme was observed in animals treated with multiple doses of the NO-generating drugs glyceryl trinitrate (16 and 80 mg/kg given every 4 h for 16 h) and nitroprusside (20 and 100 mg/kg given every 4 h for 16 h). These results indicate that the down-regulation of cytochrome P450 by interferon or its inducers probably does not involve NO as a required intermediary.

Depression of hepatic cytochrome P450 monooxygenases after chronic environmental tobacco smoke exposure of young ferrets

Six-week-old ferrets were exposed head-only to clean air or environmental tobacco smoke (ETS) for 2 h/day, 5 days a week for a total of 8 weeks. Exposure to ETS caused a significant reduction in the levels of hepatic microsomal cytochrome P450 and P450 reductase activity in both the male and female ferrets. The content of cytochrome b 5 and the activity of its reductase were significantly reduced in the hepatic microsomes of female ferrets. 7-Ethoxycoumarin O- deethylase activity and cytochrome P450 (CYP) 1A protein were markedly decreased in the hepatic microsomes of both the male and female ferrets after ETS exposure. In accord with the downregulation of P450, total metabolites formed from benzo[ a] pyrene were significantly reduced in the liver homogenates of ETS-exposed animals. Similarly, sum total of free (+)- anti- benzo[ a] pyrene-7,8- dihydrodiol-9,10- epoxide, glutathione conjugates and DNA-bound metabolites formed from precursor (−)-7 R- trans- benzo[ a] pyrene-7,8- dihydrodiol showed marked reduction in both the male and female ferrets after ETS exposure in a dose-response manner. This is the first report showing downregulation of hepatic P450 and accompanying benzo[ a] pyrene metabolism after tobacco smoke exposure which apparently occurred after an initial upregulation of these parameters (Rasmussen et al. (1994) FASEB J. 8, A122).

Dissociation of xanthine oxidase induction and cytochrome P450 depression during interferon induction in the rat

Interferon (IFN) and IFN inducers down-regulate hepatic cytochrome P450 (P450) through a pretranslational mechanism involving depression of P450 mRNA levels and a subsequent decrease in P450 synthesis. Current evidence suggests that interferon induces the synthesis of a protein which subsequently mediates the down-regulation of P450. Xanthine oxidase (XO) activity is induced by interferons in rodents, and the XO inhibitor allopurinol (AP) inhibits the down-regulation of P450 by interferons in the mouse and hamster so it has been proposed as the putative intermediate protein. In studies undertaken in rats to further characterize the molecular basis of the protective effect of AP, we observed that AP (20 and 50 mg/kg) did not protect against down-regulation of P450 by the interferon inducer polyinosinic-polycytidylic acid (10 mg/kg). In fact, at 50 mg/kg AP had an additive effect on the depression of CYP2E1. Total XO induction in the rat was only 30–50% compared with 100–500% in mice and hamsters, and this induction was inhibited completely by AP. Therefore, XO does not mediate the down-regulation of hepatic cytochrome P450 by interferons in the rat.

Interferon suppresses the isoform-specific activities of hepatic cytochrome P450 in female rats

The results from the present study indicate that rat IFN-gamma decreases hepatic P450 levels and catalytic activities in female rats. The magnitude of the down-regulation of P450 is similar to that found in male rats. Furthermore, the activities of specific P450 isoforms (P4502A1 and P4502C6 and possibly P4502C12, and P4502B1 and 2) are decreased by IFN. Hence, the effect of IFN in female rats appears to involve several, but not all, hepatic P450 isoforms. Both hormone-dependent and hormone-independent isoforms are suppressed by IFN. The mechanism for this effect does not, however, appear to be mediated by changes in serum levels of sex steroids.

Age-dependence of the rat Leydig cell and Sertoli cell function.

The first part of this study compares peripheral testosterone levels with intratesticular levels of the cytochromes P-450 of two key enzymes of androgen biosynthesis, i.e. mitochondrial cholesterol monooxygenase (P-450(cscc)) and microsomal steroid-17 alpha-monooxygenase (P-450(C17 alpha)) during puberty and early adulthood of male Wistar rats. From 4 to 10 weeks of age, the testosterone level increases 8.7-fold, the P-450(C17 alpha) level 8.3-fold, but the P-450(cscc) level 24.5-fold as an indication of specific induction of this protein. From 13 to 50 weeks of age, the testosterone level remains constant, the P-450(cscc) level increases continuously by a factor of 1.4, but 62% of the P-450(C17 alpha) content are lost. This discrepancy is explained by a divergent regulation of the cytochromes P-450 of the two steroid monooxygenases: a persisting induction of P-450(cscc) and a concurrent down-regulation of P-450(C17 alpha) that may be a consequence of the high rate of Leydig cell steroid hydroxylation after puberty. Overlapping of both processes may (probably besides other developmental factors) result in a constant testosterone concentration in blood. The second part of the study compares testicular and epididymal levels of androgen-binding protein (ABP) with the peripheral testosterone level. The peripubertal increase in testicular ABP content is shown to be related only to the increase in testicular mass, whereas a specific accumulation of ABP occurs in the epididymis from 4 to 13 weeks of age. This pattern indicates an increasing secretory activity of the Sertoli cells that remains high during adulthood up to the 50th week.(ABSTRACT TRUNCATED AT 250 WORDS)