Developmental changes in the expression of enterocytic and hepatic cytochromes P4501A in rat

1. The development of CYP1A enzymes was studied in enterocytic and hepatic microsomes from 1-day-old to adult male and female rats. Microsomes were prepared by calcium precipitation. Enzyme expression was determined by Western blotting using a polyclonal CYP1A1 antibody. 2. The developmental expression of CYP1A in enterocytic and hepatic microsomes was similar for males and females. 3. Enterocytic CYP1A (CYP1A1) showed a sharp increase at weaning, plateauing at adult levels by 60 days. 4. Hepatic CYP1A (mostly CYP1A2) increased sharply just before weaning. However, in contrast to the enterocytic enzyme, there was a 4-fold decrease in enzyme expression down to adult levels by day 60.     

Developmental changes in the expression of enterocytic and hepatic cytochromes P4501A in rat

1. The development of CYP1A enzymes was studied in enterocytic and hepatic microsomes from 1-day-old to adult male and female rats. Microsomes were prepared by calcium precipitation. Enzyme expression was determined by Western blotting using a polyclonal CYP1A1 antibody. 2. The developmental expression of CYP1A in enterocytic and hepatic microsomes was similar for males and females. 3. Enterocytic CYP1A (CYP1A1) showed a sharp increase at weaning, plateauing at adult levels by 60 days. 4. Hepatic CYP1A (mostly CYP1A2) increased sharply just before weaning. However, in contrast to the enterocytic enzyme, there was a 4-fold decrease in enzyme expression down to adult levels by day 60.     

The effect of CYP1A induction on amiodarone disposition in the rat

In the treatment of cardiac arrhythmias, amiodarone (AM) has emerged as a primary therapeutic agent. In addition to other cytochrome P450 (CYP), 1A1 and 1A2 facilitate the biotransformation of AM to the pharmacologically and toxicologically active metabolite, desethylamiodarone (DEA). The exposure to polycyclic aromatic hydrocarbons can induce these isoforms. This study was aimed at investigating the effect of CYP1A induction on the disposition of AM, after single and multiple intravenous doses, using β-naphthoflavone (BNF) treated rats to model induction of CYP1A. After a single dose (25 mg/kg), the plasma AUC of DEA were significantly induced (～3-fold). With multiple doses, AM AUC_{0–24 h} was significantly reduced in the BNF plasma (30%), lung (35%), liver (48%), kidney (52%), heart (34%), and intestine (43%). In contrast the DEA AUC_{0–24 h} was increased significantly in the BNF plasma (36%), lung (56%), liver (101%), kidney (65%), and heart (73%). The DEA/AM ratios of the AUC_{0–24 h} were increased in the BNF plasma, lung, liver, kidney, and heart by 1.9-, 2.5-, 3.8-, 3.4-, and 2.7-fold, respectively. In both groups of rats, the highest concentrations of AM and DEA were in the lung. Exposure to BNF was shown to increase DEA concentrations in the rat. © 2009 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:539–548, 2010     

Inhibition of murine cytochrome P4501A by tacrine: in vitro studies.

Tacrine, a cholinesterase inhibitor, was approved for the treatment of Alzheimer's disease. Oxidative metabolism of tacrine occurs by CYP1A-catalyzed hydroxylation. In rats, it was observed that the area under the curve (AUC) of the second oral dose was consistently higher than the AUC after the first oral dose, which was not due to the accumulation of the drug in the plasma from the first dose. This finding suggested inhibition of the enzyme during metabolism or inhibition by a metabolite. The inhibitory mechanism was studied in liver and intestinal microsomes prepared from 3-methylcholanthrene-treated rats and with recombinant CYP1A1 and CYP1A2. Preincubation of CYP1A2 with tacrine and NADPH revealed a time-dependent inhibition of 7-ethoxyresorufin O-de-ethylation with a K(i) of 1.94 microM and a k(inact) of 0.091 min(-1). No time-dependent inhibition was observed with CYP1A1 or with 1-hydroxytacrine or 2-hydroxytacrine. Tacrine metabolism catalyzed by CYP1A was also carried out, and the partition ratio was estimated to be 22. A modified Michaelis-Menten equation involving mechanism-based inhibition was derived and used to analyze the data. Reasonable parameter fits were obtained indicating that this equation is suitable to describe metabolism data when the substrate is a mechanism-based inhibitor of the enzyme. The probable inactivation mechanism involves either hydrogen atom abstraction to produce a carbon-centered radical intermediate at the benzylic position or insertion of OH(+) into a C-H bond with subsequent loss of water to produce a carbocation. Rapid rearrangement of the carbocation or radical and subsequent covalent binding of the tacrine moiety would result in enzyme inactivation.     

A comparison of the ontogeny of enterocytic and hepatic cytochromes P450 3A in the rat

Enzymes of the cytochrome P450 3A (CYP3A) sub-family are abundant in adult liver and gut and contribute significantly to the first-pass metabolism of many orally administered drugs. The development of CYP3A enzymes with regard to their expression and activity in enterocytic and hepatic microsomes from 1-day-old through to adult male and female rats has been studied. Microsomes were prepared by calcium precipitation. Enzyme expression was assessed semi-quantitatively by Western blotting using rat polyclonal CYP3A2 and 2C11 antibodies and peptide antibodies specific to rat CYPs 3A1, 3A2, 2C12, and 2C13. The formation of 6beta-hydroxytestosterone (6OHT), determined by HPLC, was used as a measure of enzyme activity. Formation of 6OHT by enterocytic microsomes was similar for males and females and showed a sharp increase at weaning. This pattern was mirrored by levels of immunoquantifiable CYP3A2 (CYP3A9), but CYP3A1 followed a more gradual development. CYPs 2C11, 2C12, or 2C13 were not detected in gut microsomes. In contrast, CYPs 3A1, 3A2, 2C11, 2C12, and 2C13 were all expressed in hepatic microsomes. There was no surge in hepatic enzyme expression or hepatic 6OHT formation at weaning, and a marked sex difference in 6OHT formation was apparent from day 25. The surge in gut activity at weaning may be a protective mechanism against ingested toxins.     

Modulation of the induction of rat hepatic cytochromes P-450 by selenium deficiency

The induction by phenobarbital of liver microsomal cytochrome P-450 has been demonstrated to be impaired in rats fed a selenium-deficient diet. Cytochrome P-450 isozyme specific immunologic and molecular techniques were used in the present study to better define the role of selenium in the induction of cytochrome P-450 by phenobarbital. Phenobarbital treatment of the selenium-deficient rats resulted in an increase in the level of total cytochrome P-450 50% of that observed with control rats and in a 10-fold increase in microsomal heme oxygenase. Quantitative immunoblot analyses demonstrated that the levels of cytochromes P-450b + e and P-450p in the phenobarbital-treated selenium-deficient rats were approximately 50% of those found in the phenobarbital-treated control rats. Finally, RNA hybridization studies using cDNA probes to cytochromes P-450b + e or P-450p demonstrated that the accumulations of the RNAs encoding these cytochromes P-450 were unaffected by the selenium status of the rats. These studies suggest that the impaired phenobarbital induction of the cytochromes P-450 in the selenium-deficient rats is the result of an increase in the degradation of the cytochromes P-450 or a decrease in the translation of the mRNAs coding for them.     

加替沙星和环丙沙星对大鼠肝微粒体酶系的影响

目的研究加替沙星和环丙沙星对大鼠肝微粒体细胞色素P450酶系的影响.方法Wistar大鼠经加替沙星和环丙沙星400mg/kg灌胃给药,qd×7d后,应用差速离心法制备大鼠肝微粒体,采用Lowry法测定蛋白浓度,应用分光光度计法检测6种肝微粒体细胞色素P450酶含量及活性,并用单因素方差分析进行统计.结果加替沙星组中6种细胞色素P450酶测定结果与空白对照组相比差异无统计学意义,而环丙沙星能抑制6种细胞色素P450酶中的b5、NADPH-CytC还原酶、氨基比林N-脱甲基酶、红霉素N-脱甲基酶和7-乙氧基香豆素脱烃酶的活性,对CYP450酶系有选择性抑制作用.结论加替沙星对大鼠肝微粒体CYP450酶系无显著影响,而环丙沙星对CYP450酶系有选择性抑制作用.     

Cytochrome P4501A(CYP1A) induction in rat and man by the benzodioxino derivative,fluparoxan

1. Fluparoxan is an α₂-adrenoceptor antagonist that has a relatively planar, tricyclic structure and was considered a potential substrate and inducer of cytochrome P4501A (CYP1A) enzymes. 2. Structure-activity analysis indicated some potential for CYP1A interaction, although its greater log P and molecular depth, compared with many CYP1A inducers, suggested fluparoxan would be a weak ligand for the aryl hydrocarbon (Ah) receptor and only a weak inducer. 3. In vitro, fluparoxan showed little affinity for the CYP1A enzymes. The compound was not metabolized by human CYP1A1 or 1A2 heterologously expressed in yeast and its rate of metabolism in rat and human microsomes was unaffected by the addition of the 1A inhibitor α-naphthoflavone. Furthermore, K_i's for fluparoxan against EROD activity were >4000-fold higher than those of α-naphthoflavone. 4. In vivo, however, fluparoxan did show some capacity for CYP1A induction. In rat, hepatic EROD activity increased approximately 40- fold with seven once-daily oral doses of fluparoxan (50?mg kg, solution), and immunoblotting studies confirmed induction of CYP1A2, though not of 1A1. In man, administration of 11 twice-daily oral doses of fluparoxan (8?mg tablet) produced some reduction in plasma levels of orally administered phenacetin and in the ratio of phenacetin AUC urinary paracetamol, consistent with increased O-deethylation.     

糖肝康对糖尿病慢性肝损伤模型大鼠肝脏结构和细胞色素P450的影响

目的：观察糖肝康对糖尿病慢性肝损伤大鼠肝脏组织光镜病理结构、电镜超微结构和肝功能、细胞色素P450（ CYP450）的影响。方法将50只大鼠分为正常组、模型组、糖肝康小剂量组、糖肝康大剂量组、降糖甲组5组,每组10只。用链脲佐菌素和四氯化碳腹腔注射建立糖尿病慢性肝损伤大鼠模型。干预12周后,应用光镜和电镜观察各组大鼠肝脏病理结构和超微结构改变,比较各组大鼠肝功能,比较各组大鼠CYP4501A1,CYP4502E1,CYP4503A1指标。结果光镜下,模型组大鼠肝细胞大量脂肪变性、增生有异型性,可见核分裂象,肝小叶间有少量纤维组织增生;糖肝康大剂量组肝脏结构基本正常。电镜下,模型组大鼠肝细胞体积小,核变形、皱缩,有切迹,核仁明显,常染色质团块状,异染色质边聚,线粒体肿胀,亦可见线粒体空泡化,线粒体嵴模糊;内质网肿胀;可见大量脂滴。糖肝康大剂量组肝细胞结构基本正常。糖肝康小剂量组与模型组相比,ALT明显降低（P＜0．05）;糖肝康大剂量组与模型组相比,ALT明显降低（P＜0．01）;降糖甲组与模型组相比,ALT明显降低（P＜0．01）;糖肝康大剂量组与降糖甲组相比,ALT明显降低（P＜0．05）。糖肝康小剂量组与模型组相比,AST明显降低（ P＜0．01）;糖肝康大剂量组与模型组相比,AST明显降低（P＜0．01）;降糖甲组与模型组相比,AST明显降低（P＜0．05）;糖肝康大剂量组与降糖甲组相比,AST明显降低（P＜0．01）。糖肝康小剂量组、糖肝康大剂量组分别与模型组相比,CYP4501A1,CYP4502E1,CYP4503A1的表达均降低（P＜0．05）;降糖甲组与模型组相比,CYP4501A1, CYP4502E1,CYP4503A1的表达均降低（P＜0．05）;糖肝康大剂量组与降糖甲组相比,CYP4501A1,CYP4502E1,CYP4503A1的表达均明显降低（ P＜0．05）。结论糖肝康对肝脏的病理改变起到一定改善作用,还可以通过调节肝功能,CYP4501A1,CYP4502E1,CYP4503A1的表达,起到对肝脏的保护作用。     

细胞色素P4501A2活性与奥氮平代谢的相关性研究

目的 研究奥氮平(抗精神病药)体内代谢与细胞色素P450酶亚型1A2(CYP1A2)活性的相关性.方法 15例男性健康志愿者单次口服咖啡因150mg,第5 h末采血;2天后单次口服奥氮平10 mg,收集血样;用高效液相色谱电化学法测定奥氮平血浆浓度,紫外法测定咖啡因及其代谢产物次黄嘌呤的血浆浓度.结果 次黄嘌呤与咖啡因的比值与奥氮平清除的奥氮平AUC0.96的倒数不相关(γ=0.057,P=0.84).结论 CYP1A2酶活性与奥氮平体内代谢不相关.     

The influence of ageing on the induction of the mRNAs of rat liver cytochromes P450IIB1 and P450IIB2

To investigate the influence of age on the regulation of the cytochromes P450IIB1 and P450IIB2 the levels of the messenger RNAs for these two cytochromes were determined in liver cytoplasmic RNA of rats of various ages after maximal induction with either phenobarbital or isosafrole and in untreated rats. The levels of these mRNAs were determined by solution hybridization with a RNA-probe (riboprobe system) complementary to both mRNAs. This study showed a marked decrease in the maximal induction levels of these mRNAs between the ages of 12 and 36 months irrespective of the type of inducer used. To assess whether this age-related decrease could be found for both individual mRNAs also solution hybridization experiments were performed with deoxyoligonucleotide probes of a defined sequence. The data presented in this paper show that ageing influences the levels of both the cytochrome P450IIB1 and P450IIB2 mRNA in a similar way. After induction the amount of mRNA for P450IIB1 was in all age groups measured four- to five-fold higher than that of P450IIB2. These data indicate that previously observed age-related changes in the cytochrome P450 system could be related to a lower accumulation of its mRNAs.     

Effect of natural analogues of trans-resveratrol on cytochromes P4501A2 and 2E1 catalytic activities

The aim was to assess the inhibitory effect of a series of naturally occurring trans-resveratrol analogues on cytochromes P450, namely CYP1A2 and CYP2E1, in vitro in order to analyse any structure-activity relationships. 3,5-Dimethoxy-4'-hydroxy-trans-stilbene (pterostilbene), 3,4',5-trimethoxy-trans-stilbene (TMS), 3,4'-dihydroxy-5-methoxy-trans-stilbene (3,4'-DH-5-MS) and 3,5-dihydroxy-4'-methoxy-trans-stilbene (3,5-DH-4'-MS) inhibited the activity of CYP1A2, with K(i) = 0.39, 0.79, 0.94 and 1.04 microM, respectively. Piceatannol (3,3',4,5'-tetrahydroxy-trans-stilbene) was the least potent inhibitor of CYP1A2 with a K(i) = 9.67 microM. Piceatannol and TMS in the concentration range 1-100 microM did not inhibit CYP2E1 activity. The activity of this enzyme likewise was not significantly influenced by pterostilbene and 3,5-DH-4'-MS with IC(50) > 100 microM, whereas 3,4'-DH-5-MS appeared to be a moderately potent, competitive inhibitor of CYP2E1 (K(i) = 42.6 microM). Structure-activity relationship analysis leads to the conclusion that the substitution of hydroxy groups of resveratrol with methoxy groups increases the inhibition of CYP1A2, yet the number and position of methylation are not essential. However, the 4'-hydroxy group in trans-resveratrol and its analogues may play an important role in the interaction with a binding site of CYP2E1.     

Effect of natural analogues of trans-resveratrol on cytochromes P4501A2 and 2E1 catalytic activities

The aim was to assess the inhibitory effect of a series of naturally occurring trans-resveratrol analogues on cytochromes P450, namely CYP1A2 and CYP2E1, in vitro in order to analyse any structure–activity relationships. 3,5-Dimethoxy-4′-hydroxy-trans-stilbene (pterostilbene), 3,4′,5-trimethoxy-trans-stilbene (TMS), 3,4′-dihydroxy-5-methoxy-trans-stilbene (3,4′-DH-5-MS) and 3,5-dihydroxy-4′-methoxy-trans-stilbene (3,5-DH-4′-MS) inhibited the activity of CYP1A2, with K_i?=?0.39, 0.79, 0.94 and 1.04?µM, respectively. Piceatannol (3,3′,4,5′-tetrahydroxy-trans-stilbene) was the least potent inhibitor of CYP1A2 with a K_i?=?9.67?µM. Piceatannol and TMS in the concentration range 1–100?µM did not inhibit CYP2E1 activity. The activity of this enzyme likewise was not significantly influenced by pterostilbene and 3,5-DH-4′-MS with IC₅₀?>?100?µM, whereas 3,4′-DH-5-MS appeared to be a moderately potent, competitive inhibitor of CYP2E1 (K_i?=?42.6?µM). Structure–activity relationship analysis leads to the conclusion that the substitution of hydroxy groups of resveratrol with methoxy groups increases the inhibition of CYP1A2, yet the number and position of methylation are not essential. However, the 4′-hydroxy group in trans-resveratrol and its analogues may play an important role in the interaction with a binding site of CYP2E1.     

The effect of domperidone on theophylline disposition in the rat

The effect of domperidone (2 mg kg-1) on the pharmacokinetics of a single oral dose of theophylline (25 mg kg-1) was studied in the rat. Theophylline concentrations were measured serially for 12 h using an HPLC technique. Domperidone did not have any significant effect on any of the four parameters studied: peak plasma levels (Cpmax), the time these were attained (tmax), elimination half-life (t1/2) and area under the plasma concentration-time curve (AUC). Our data preliminarily suggests that domperidone may be safely coadministered with theophylline but clearly further studies in patients or relevant animal models of gastric motility disturbances are needed to reliably rule out any potential interaction between these agents.     

Regulation of pulmonary and hepatic cytochrome P4501A expression in the rat by hyperoxia: implications for hyperoxic lung injury

Supplemental oxygen therapy is frequently used in the treatment of pulmonary insufficiency, as is encountered in premature infants, and in patients with acute respiratory distress syndrome. However, hyperoxia causes lung damage in experimental animals and may do so in humans. Cytochrome P4501A enzymes have been implicated in hyperoxic lung injury. In this study, we investigated the mechanisms of CYP1A1 regulation by hyperoxia and tested the hypothesis that aryl hydrocarbon receptor (AHR)-dependent mechanisms contribute to induction of CYP1A1 and that modulation of CYP1A by hyperoxia may have implications for lung injury. Exposure of adult male Sprague-Dawley rats to hyperoxia for 24 to 48 h led to increased expression of pulmonary CYP1A1 enzyme, which was preceded by enhancement of the corresponding mRNA, followed by decline of induction at 60 h, when the animals displayed severe respiratory distress and lung inflammation. Similarly, hepatic CYP1A1/1A2 mRNAs were markedly induced between 24 and 48 h of hyperoxia, with induction declining by 60 h. Electrophoretic mobility shift assays (EMSA) and experiments with AHR (-/-) mice indicated that AHR-dependent mechanisms contributed to CYP1A induction. The AHR (-/-) mice were refractory to CYP1A1 induction by hyperoxia and were more sensitive to lung injury than wild-type mice. Lungs of hyperoxic rats showed increase in the expression of CYP1A1 in airway epithelial cells, type II pneumocytes, and endothelial cells. In conclusion, our results suggest that induction of CYP1A1 by hyperoxia is mediated by AHR-dependent mechanisms and that modulation of CYP1A enzymes by hyperoxia may have implications for hyperoxic lung injury.     

Piperine effects on the expression of P4502E1, P4502B and P4501A in rat

Treatment of rat with piperine (PIP) (1·4 mmol/kg, 3 days ip injections) resulted in an approximate two-fold increase in total liver microsomal P450 content relative to that in uninduced animals.

2. 4-Nitrophenol and aniline hyroxylase activities in the hepatic microsomes prepared from rat treated with PIP decreased by 30 and 28% respectively as compared with control. Immunoblot analyses also revealed decreased P4502E1 levels in hepatic microsomes from PIP-treated animals.

3. In contrast with P4502E1 suppression, hepatic 2B1 and 2B2 levels were significantly increased in PIP-induced animals, as evidenced by both metabolic activity and immunoblot analysis of the liver microsomal fractions. The rate of hexobarbital hydroxylase activity in microsomes from PIP-treated animals was markedly elevated and was inhibited by approximately 62% in the presence of monoclonal anti-P4502B IgG. Immunoblot analyses demonstrated that P4502B1 ana 2B2 levels in hepatic microsomes from PIP-treated animals were comparable with those from phenobarbital-treated animals.

4. 7-Ethoxycoumarin deethylase activity was elevated approximately two-fold in PIP-induced animals and was 17% of that derived from 3-methylcholanthrene-induced animals. 7-ethoxycoumarin deethylase activity in PIP-induced hepatic microsomes was inhibited 63% in the presence of monoclonal anti-P4501 A antibody. Immunoblot analysis confirmed the increase in P4501A levels by PIP, which was 15% of that in hepatic microsomes from 3-methylcholanthrene-induced animals.

5. PIP treatment failed to affect microsomal epoxide hydrolase (mEH) and glutathione S-transferases (GST) expression, as indicated by immunoblot analyses using polyclonal antibodies toward mEH and GST subunits Ya, Yb1, Yb2 and Yc.

6. These results demonstrate that PIP treatment suppressed P4502E1 expression and enhanced 2B and 1A expression, whereas this agent failed to affect hepatic mEH and GST expression.     

The effect of hepatic disease on the disposition of moricizine in humans

The pharmacokinetics of moricizine and two of its metabolites, moricizine sulfoxide and phenothiazine-2-carbamic acid ethyl ester sulfoxide, were studied in healthy control subjects and in patients with chronic liver disease (cirrhosis). Moricizine disposition was significantly altered by hepatic cirrhosis. Compared to healthy subjects, the hepatic disease patients had an increased C_max (59%), an increased t_1/2 (141%), and a reduced plasma clearance (71%). Additionally, small but statistically significant increases were observed for t_max and the fraction of moricizine not bound to plasma proteins in patients with hepatic disease. The elimination of both moricizine metabolites was also altered by hepatic dysfunction as indicated by significantly prolonged terminal half-lives. Furthermore, there was a reduction in the conversion of moricizine to moricizine sulfoxide. Both hepatic blood flow and hepatic metabolizing capacity were assessed in all subjects and patients by administration of indocyanine green and antipyrine, respectively. Indocyanine green and antipyrine plasma clearances were decreased by 38 and 51%, respectively, indicating that both functions were diminished by hepatic cirrhosis. We conclude that the moricizine dose required for arrhythmia patients with hepatic disease should be lower, and perhaps, the dosing frequency should be less than in patients with normal liver function.     

Dose-response studies on the induction of liver cytochromes P4501A1 and 1B1 by polycyclic aromatic hydrocarbons in arylhydrocarbon-responsive C57BL/6J mice

1. To determine the biological effects of 23 polycyclic aromatic hydrocarbons (PAHs) and 3,4,30,40-tetrachlorobiphenyl, the dose–response studies of the induction of CYP1- dependent xenobiotic oxidation activities by these chemicals in liver microsomes of C57BL/6J mice were studied. 2. In arylhydrocarbon-responsive C57BL/6J mice, the liver microsomal xenobiotic oxidation with substrates of 7-ethoxyresoru.n, 7-ethoxycoumarin, (±)-benzo[a]pyrene- 7,8-diol, dibenzo[a,l] pyrene-11,12-diol and 2-amino-3,5-dimethylimidazo[4,5-f]quinoline increased by increasing the doses of PAHs to mice, particularly when the PAHs that have been reported to be carcinogenic in experimental animals were used. In arylhydrocarbon receptor-knockout mice, there were no increases in liver microsomal 7-ethoxyresoru.n O-deethylation activities nor in liver mRNA levels of CYP1A1, 1A2 and 1B1 by these chemicals. 3. Of the chemicals examined, benzo[k].uoranthene, benzo[b].uoranthene, benzo [j] -. uoranthene, 3-methylcholanthrene, dibenz[a,h]anthracene, dibenz[a,c]anthracene and 3,4,30,40-tetrachlorobiphenyl were potent inducers of the induction of liver microsomal 7-ethoxyresoru.n O-deethylation in mice. 4. Other PAHs such as 5-methylchrysene, benzo[a]pyrene, dibenzo[a, l] pyrene, dibenz[a, j]acridine, benzo[a]anthracene and 7,12-dimethylbenz[a]anthracene moderately induced 7-ethoxyresoru.n O-deethylation activities in mice. PAHs reported to be weak or less carcinogenic in experimental animals did not induce the xenobiotic oxidation activities of CYP1A1 and 1B1 in the mice. 5. The results suggest that induction of liver microsomal CYP1-dependent xenobiotic oxidation activities is a good tool in determining the potencies of carcinogenic PAHs in arylhydrocarbon-responsive C57BL/6J mice.     

Comparison of hepatic and extra hepatic induction of cytochrome P4501A by graded doses of aryl hydrocarbon receptor agonists in Atlantic tomcod from two populations

Atlantic tomcod Microgadus tomcod from the Hudson River, New York, are exposed to high levels of polycyclic aromatic hydrocarbons (PAHs) and bioaccumulate mixtures of polychlorinated biphenyls (PCBs), polychlorinated dibenzo-p-dioxins and polychlorinatedfurans (PCDD/Fs). Previous studies demonstrated that hepatic cytochrome P4501A (CYP1A) mRNA was not inducible in tomcod from the Hudson River treated with single doses of PCB77 or 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), but was inducible with PAHs. In this study, we sought to determine if CYP1A mRNA was inducible with higher doses of these and other halogenated aromatic hydrocarbons (HAHs) in Hudson River tomcod and if decreased sensitivity to gene inducibility occurs across all tissues. Tomcod from the Hudson River and the cleaner Miramichi River, New Brunswick, were treated individually with graded doses of TCDD and coplanar PCBs (PCB77, PCB81, PCB126, PCB169) and profiles of hepatic CYP1A mRNA expression were compared between the two populations. CYP1A mRNA inducibility was also compared in multiple tissues of tomcod from the two rivers that were treated with PCB77. Additionally, hepatic CYP1A mRNA was characterized in Miramichi River tomcod treated with pairs of PCB congeners that included aryl hydrocarbon receptor (AHR) agonists and antagonists. Hepatic CYP1A mRNA was significantly inducible by all agonists in tomcod from the Miramichi River and TCDD and two of four PCBs in tomcod from the Hudson River. CYP1A mRNA was also significantly inducible in four of five tissues of tomcod from the Miramichi River but only in liver of Hudson River tomcod. In summary, CYP1A mRNA inducibility was approximately two orders of magnitude less sensitive in tomcod from the Hudson River than in those from the Miramichi River. But when achieved, maximum levels of CYP1A expression were similar in tomcod from the two populations. Co-administration of PCB126 and PCB77 did not produce significantly greater CYP1A mRNA induction than administration of PCB126 alone and co-administration of mono-ortho-substituted PCB105 significantly decreased CYP1A mRNA inducibility by PCB77. These results indicate that CYP1A mRNA expression is significantly inducible by HAHs in tomcod from the Hudson River and suggest that all components of the AHR pathway are present and functional, but that the pathway is less sensitive to activation than in tomcod from the Miramichi River. Our results also indicate that CYP1A mRNA levels in environmentally exposed fish may not reflect additive tissue burdens of PCB congeners.