Induction of CYP3A and associated terfenadineN-dealkylation in rat hepatocytes cocultured with 3T3 cells

Long-term culture of hepatocytes has been challenged by the loss of differentiated functions. In particular, there is a rapid decline in cytochrome P450 (CYP). In this study, we cocultured rat hepatocytes with 3T3 fibroblasts for 10 days, and examined hepatocyte viability, morphology, and expression of CYP3A. Terfenadine was incubated with the cultures, and its biotransformation was quantitatively analyzed by HPLC. Terfenadine is metabolized by two major pathways:C-hydroxylation to an alcohol metabolite which is further oxidized to a carboxylic acid, andN-dealkylation to azacyclonol. In rat liver, only theN-dealkylation pathway appears to be mediated by CYP3A since anti-rat CYP3A antibody inhibited azacyclonol but not alcohol metabolite formation in incubations of terfenadine with liver microsomes. Freshly isolated rat hepatocytes were seeded on top of confluent 3T3 cells. Cultures were maintained in Williams' E medium supplemented with 10% fetal bovine serum and either 0.1 mol/L or 5 mol/L dexamethasone. In pure hepatocyte cultures, viability, as determined by lactate dehydrogenase (LDH) activity, decreased steadily to less than 30% of initial levels by day 10. In cocultures, LDH activity remained high and was 70% of initial levels on day 10. The half-life of terfenadine disappearance was optimally maintained in cocultures treated with 5 mol/L dexamethasone, and was associated with the increased formation of azacyclonol. On day 5, nearly 50% of added 5 mol/L terfenadine was converted to azacyclonol within 6 h, whereas the conversion was only 4% on day 1. Western and RNA-slot blot analyses confirmed that treatment with 5 mol/L dexamethasone induced CYP3A mRNA expression and CYP3A protein expression. This coculture system could offer a useful approach in the study of drugs and xenobiotics metabolized by CYP3A.Abbreviations BSA bovine serum albumin - CYP cytochrome P450 - DMSO dimethyl sulfoxide - LDH lactate dehydrogenase - PCN pregnenolone-16-carbonitrile - SDS sodium dodecyl sulfate - SSC saline sodium citrate     

Regulation of phenobarbital-induction of CYP2B and CYP3A genes in rat cultured hepatocytes: involvement of several serine/threonine protein kinases and phosphatases

We investigated the involvement of diverse protein kinases and phosphatases in the transduction pathways elicited by phenobarbital (PB), a well-known inducer of some hepatic cytochromes P450 (CYP). Different inhibitors or activators of protein kinases or phosphatases were assessed for their ability to modulate PB-induction of CYP2B and CYP3A mRNA expression. Rat hepatocytes in primary culture were treated with the test compounds one hour prior to, and then continuously, in the absence or presence of 1 mmol/L PB for 24 h. By northern blot analysis of CYP2B1/2 and 3A1/2 gene expression, we first confirmed the negative role of the adenosine 3′:5′ cyclic monophosphate (cAMP)/protein kinase A pathway and the positive role of some serine/threonine protein phosphatases in the mechanism of PB-induction. The present data further suggested that Ca²⁺/calmodulin-dependent protein kinases II (independently of Ca²⁺) and extracellular signal-regulated kinases 1/2 (ERK1/2) might function respectively as positive and negative regulator in the PB-induction of CYP2B and CYP3A. In contrast, protein kinases C and phosphatidylinositol-3-kinase did not appear to be involved, while the role of tyrosine kinases remained unclear. We conclude that a complex network of phosphorylation/dephosphorylation events might be crucial for PB-induction of rat CYP2B and CYP3A. This revised version was published online in July 2006 with corrections to the Cover Date.     

Regulation and induction of CYP3A11, CYP3A13 and CYP3A25 in C57BL/6J mouse liver

This study reports that dexamethasone (DEX) significantly induces CYP3A11, CYP3A13 and CYP3A25 mRNA expression in male and female 4 days, 3 weeks and 18 weeks old C57BL/6J mice. Furthermore, CYP3A activity, as measured by erythromycin-N-demethylation, is also significantly increased. PXR, RXRalpha and CAR are known to be involved in the induction of CYP3As. Here we report nuclear receptors PXR and RXRalpha but not CAR demonstrate gender- and age-dependent expression. Also, treatment of C57BL/6J mice with DEX induces PXR but not RXRalpha or CAR. In summary, we demonstrate DEX is not only able to up-regulate CYP3A expression and activity, but also the nuclear receptor PXR through which it may exert this effect. Furthermore, the gender- and age-dependent pattern of basal PXR and RXRalpha expression is similar to the 3 CYP3As analysed.     

Expression and induction of CYP3As in human fetal hepatocytes

CYP3A4 and CYP3A7 mRNA expression levels were markedly up-regulated by dexamethasone (DEX), but not by rifampicin (RIF). CYP3A5 mRNA level was not increased significantly by DEX, RIF, or phenobarbital. Testosterone 6beta-hydroxylase activity was induced to about 2-fold of control by DEX. However, concomitant treatment with RIF did not alter DEX-mediated induction of CYP3A mRNA expression and testosterone 6beta-hydroxylase activity. DEX-mediated induction of CYP3A mRNA was suppressed in a dose-dependent manner by RU486, a glucocorticoid receptor (GR) antagonist. At 5microM RU486, DEX-mediated induction of CYP3A4, CYP3A5, and CYP3A7 mRNA expression was inhibited almost completely. These results suggest that, in human fetal hepatocytes, PXR is not involved in DEX-mediated induction of CYP3A4 and CYP3A7, and that the induction is mediated directly by GR.     

氯吡格雷对大鼠肝药物酶CYP3A4 和CYP1A2 表达的影响

目的：氯吡格雷主要由CYP3A4 催化使其激活,CYP1A2 也参与氯吡格雷活化。关于氯吡格雷对肝微粒体酶的影响国内外 文献报道不多,因此本实验通过检测肝细胞色素氧化酶CYP3A4 和CYP1A2 的表达,探讨氯吡格雷对大鼠肝药物酶的影 响。方法：生理盐水为对照组,氯吡格雷设高、中、低三个剂量组（27,13.5,6.75mg/kg/d）,雄性健康大鼠连续灌胃给药7天,脱臼处 死,取肝组织,通过western blot法检测大鼠肝脏CYP3A4 和CYP1A2 蛋白表达情况。结果：1）、氯吡格雷抑制大鼠CYP3A4 蛋白 表达,氯吡格雷高中低剂量组分别比生理盐水组大鼠CYP3A4 蛋白表达量降低（P<0.05）;氯吡格雷低中高剂量组间进行比较,大 鼠CYP3A4 蛋白表达量呈梯度减少（P<0.05）;2）、氯吡格雷抑制大鼠CYP1A2 蛋白表达,氯吡格雷高中低剂量组分别比生理盐水 组大鼠CYP1A2 蛋白表达量降低（P<0.05）,氯吡格雷低中高剂量组间进行比较,大鼠CYP1A2 蛋白表达量呈梯度减少（P<0.05）。 结论：氯吡格雷使肝细胞色素氧化酶CYP3A4 和CYP1A2 的表达量减少,因此氯吡格雷高、中、低3 个剂量组均不同程度的抑制 大鼠肝脏CYP3A4 和CYP1A2 的表达,提示当氯吡格雷与某些主要经CYP3A4 和CYP1A2 代谢的药物合用时,发生代谢性相关 作用的可能性大。     

氯吡格雷对大鼠肝药物酶CYP3A4和CYP1A2表达的影响

目的：氯吡格雷主要由CYP3A4催化使其激活,CYPlA2也参与氯吡格雷活化。关于氯吡格雷对肝微粒体酶的影响国内外文献报道不多,因此本实验通过检测肝细胞色素氧化酶CYP3A4和CYPlA2的表达,探讨氯吡格雷对大鼠肝药物酶的影响。方法：生理盐水为对照组,氯吡格雷设高、中、低三个剂量组（27,13．5,6．75mg／kg／d）,雄性健康大鼠连续灌胃给药7天,脱臼处死,取肝组织,通过westernblot法检测大鼠肝脏CYP3A4和CYPlA2蛋白表达情况。结果：1）、氯吡格雷抑制大鼠CYP3A4蛋白表达,氯吡格雷高中低剂量组分别比生理盐水组大鼠CYP3A4蛋白表达量降低（P〈0．05）;氯吡格雷低中高剂量组间进行比较,大鼠CYP3A4蛋白表达量呈梯度减少（P〈0．05）;2）、氯吡格雷抑制大鼠CYPlA2蛋白表达,氯吡格雷高中低剂量组分别比生理盐水组大鼠CYPlA2蛋白表达量降低（P〈0．05）,氯吡格雷低中高剂量组间进行比较,大鼠CYPlA2蛋白表达量呈梯度减少（P〈0．05）。结论：氯吡格雷使肝细胞色素氧化酶CYP3A4和CYPlA2的表达量减少,因此氯吡格雷高、中、低3个剂量组均不同程度的抑制大鼠肝脏CYP3A4和CYPlA2的表达,提示当氯吡格雷与某些主要经CYP3A4和CYPlA2代谢的药物合用时,发生代谢性相关作用的可能性大。     

CYP3A4基因原核表达质粒构建与诱导表达

以质粒pCWNF14为模板,采用PCR技术扩增出CYP3A4基因,并将其接入pET-22b（＋）、pET-28b（＋）和pET-32a（＋）载体中,经PCR测序鉴定后,转化Rosetta（DE3）2pLysS细菌,并用IPTG诱导表达．采用考马斯亮蓝染色的方法检测重组蛋白表达,3个表达重组质粒中,只有pET-32a（＋）-CYP3A4可以表达目的蛋白;在低浓度IPTG（50μmol／L）诱导6h,所表达的CYP3A4蛋白约占细菌总蛋白的40％。且该蛋白不溶于8mol／L的尿素,而溶于去污剂10mmol／L 3-（（3-Cholamidopropyl）dimethvlammonio propanesulfonic acid（CHAPS））和0．3％ lauroyl sarcosine（SKL）．成功地使CYP3A4基因在原核系统中高效表达．     

Aldrin Epoxidation in Flathead Mullet (Mugil cephalus): Possible Involvement of CYP1A and CYP3A

The primary objective of this study was to determine specific cytochrome P450 isozyme(s) involved in the metabolism of aldrin to its toxic metabolite dieldrin in flathead mullet (Mugil cephalus) liver microsomes. To identify the cytochrome P450 isozyme responsible for the aldrin metabolism in mullet liver, the effects of mammalian‐specific cytochrome P450 inhibitors and substrates were determined in the epoxidation reaction of aldrin. CYP3A‐related inhibitors, ketoconazole, SKF‐525A, and cimetidine, inhibited the metabolism of aldrin. The contribution of CYP1A to the aldrin metabolism was shown by the inhibition of 7‐ethoxyresorufin‐O‐deethylase activity in the presence of aldrin. The results indicate that CY1A and CYP3A are the cytochrome P450s involved in aldrin epoxidase activity in mullet. In addition, the suitability of aldrin epoxidase activity for monitoring of environmental pollution was also assessed in the fish samples caught from four different locations of the West Black Sea coast of Turkey.     

Regulation of the expression of two female-predominant CYP3A mRNAs (CYP3A41 and CYP3A44) in mouse liver by sex and growth hormones

A second female-predominant murine CYP3A, CYP3A44, was isolated from liver and its mRNA expression was compared with that of the previously described CYP3A41. The expression of CYP3A44 was relatively constant after birth in females, whereas it gradually declined in males after 5 weeks of age. The expression of CYP3A41 increased with age in females after 3 weeks of age, whereas it gradually declined in males after 5 weeks of age. Hypophysectomy and growth hormone replacement indicated that expression of both CYP3A mRNAs in females was dependent on the feminine plasma growth hormone profile. Estradiol induced the expression of both mRNAs and the effect was dependent on the presence of the pituitary gland. These observations suggest that endocrine control of expression might be similar, but not identical, for two female-predominant CYP3A mRNAs.     

CYP3A4基础与临床研究进展

细胞色素P4503A4(CYP3A4)是存在人类肝脏及肠道中的一种主要的细胞色素CYP450酶,约占成人肝脏CYP450酶总量的25%左右。临床中约有50%的药物是通过其代谢,并且其基因位点突变也与其多种疾病相关,知晓CYP3A4的表达水平和不同功能的遗传学基础,无论是对疾病的发病基础、临床药物的应用,会带来前所未有的启发,在药物应用过程中,通过对基因组学的认识,从而可以在基因层面了解个体代谢差异产生的原因,调整药物用量,提高疗效,最终使药物副作用降到最低限。目前对CYP3A4的研究渐趋于成熟,已逐渐阐明了其药物间相互作用的机制,它能够被多种药物竞争性抑制或者诱导,并受到某些蛋白受体的调控影响,可改变药物的药代动力学,增强或降低药效,造成个体用药差异,这也是造成药物间相互作用的重要原因。然而CYP3A4基因多态性与基因导向治疗关系,还有待进一步深入研究。该文对CYP3A4基因多态性、分布以及与临床疾病及用药的研究现状作一综述。     

Cholesterol 25-hydroxylation activity of CYP3A

To date, many studies have been conducted using 25-hydroxycholesterol, which is a potent regulator of lipid metabolism. However, the origins of this oxysterol have not been entirely elucidated. Cholesterol 25-hydroxylase is one of the enzymes responsible for the metabolism of 25-hydroxycholesterol, but the expression of this enzyme is very low in humans. This oxysterol is also synthesized by sterol 27-hydroxylase (CYP27A1) and cholesterol 24-hydroxylase(CYP46A1), but it is only a minor product of these enzymes. We now report that CYP3A synthesizes a significant amount of 25-hydroxycholesterol and may participate in the regulation of lipid metabolism. Induction of CYP3A by pregnenolone-16α-carbonitrile caused the accumulation of 25-hydroxycholesterol in a cell line derived from mouse liver. Furthermore, treatment of the cells with troleandomycin, a specific inhibitor of CYP3A, significantly reduced cellular 25-hydroxycholesterol concentrations. In cells that overexpressed human recombinant CYP3A4, the activity of cholesterol 25-hydroxylation was found to be higher than that of cholesterol 4β-hydroxylation, a known marker activity of CYP3A4. In addition, 25-hydroxycholesterol concentrations in normal human sera correlated positively with the levels of 4β-hydroxycholesterol (r = 0.650, P < 0.0001, n = 78), but did not significantly correlate with the levels of 27-hydroxycholesterol or 24S-hydroxycholesterol. These results demonstrate the significance of CYP3A on the production of 25-hydroxycholesterol.     

Modelling atypical CYP3A4 kinetics: principles and pragmatism

The Michaelis-Menten model, and the existence of a single active site for the interaction of substrate with drug metabolizing enzyme, adequately describes a substantial number of in vitro metabolite kinetic data sets for both clearance and inhibition determination. However, in an increasing number of cases (involving most notably, but not exclusively, CYP3A4), atypical kinetic features are observed, e.g., auto- and heteroactivation; partial, cooperative, and substrate inhibition; concentration-dependent effector responses (activation/inhibition); limited substrate substitution and inhibitory reciprocity necessitating sub-group classification. The phenomena listed above cannot be readily interpreted using single active site models and the literature indicates that three types of approaches have been adopted. First the 'nai ve' approach of using the Michaelis-Menten model regardless of the kinetic behaviour, second the 'empirical' approach (e.g., employing the Hill or uncompetitive inhibition equations to model homotropic phenomena of sigmoidicity and substrate inhibition, respectively) and finally, the 'mechanistic' approach. The later includes multisite kinetic models derived using the same rapid equilibrium/steady-state assumptions as the single-site model. These models indicate that 2 or 3 binding sites exist for a given CYP3A4 substrate and/or effector. Multisite kinetic models share common features, depending on the substrate kinetics and the nature of the effector response observed in vitro, which allow a generic model to be proposed. Thus although more complex than the other two approaches, they show more utility and can be comprehensively applied in relatively simple versions that can be readily generated from generic model. Multisite kinetic features, observed in isolated hepatocytes as well as in microsomes from hepatic tissue and heterologous expression systems, may be evident in substrate depletion-time profiles as well as in metabolite formation rates. Failure to adequately account for multisite kinetic phenomena will compromise any attempts to predict human drug clearance and drug-drug interaction potential from in vitro data.     

CYP3A4 and CYP2D6 inhibitory activities of Indonesian medicinal plants

Thirty samples of Indonesian medicinal plants were analyzed for their capacity to inhibit in vitro metabolism by human cytochrome P450 3A4 (CYP3A4) and CYP2D6 with a radiometric assay. The MeOH-soluble fractions of 25 samples, prepared from water extracts, demonstrated inhibitory activity more than 50% on the metabolism mediated by CYP3A4, and 21 samples on the metabolism mediated by CYP2D6. Among the MeOH-soluble fractions, Piper nigrum leaf showed the highest inhibitory activity against CYP3A4 (91.7%), and Punica granatum against CYP2D6 (98.1%). The water extracts of which MeOH-soluble fraction showed inhibitory activity more than 70% were fractionated with EtOAc. From the EtOAc-soluble fractions, Curcuma heyneana (67.0%), Pi. cubeba (75.0%), Pi. nigrum fruit (84.0%), Pi. nigrum leaf (85.8%), and Zingiber aromaticum (75.3%) demonstrated inhibitory activity more than 50% on the metabolism mediated by CYP3A4, but only Pi. nigrum fruit (72.8%) and Pi. nigrum leaf (69.1%) showed strong inhibitory activity against CYP2D6. For samples that showed more than 70% inhibition, their IC(50) values were determined. The most potent inhibitory activity against CYP3A4 (IC(50) value of 25 microg/ml) was found for the extract of Pi. nigrum leaf, while that of Catharanthus roseus showed the most potent inhibitory effect against CYP2D6 (IC(50) value of 11 microg/ml). These results should indicate once more the possibility of potential medicinal plant-drug interactions.     

Inhibition of CYP450 1A and 3A by berberine in crucian carp Carassius auratus gibelio

Berberine has long been considered as an antibiotic candidate in aquaculture. However, studies regarding its effects on drug-metabolizing enzymes in fish are still limited. In the present study, the effects of berberine on cytochrome P4501A (CYP1A) and CYP3A in crucian carp were investigated. Injection of different concentrations of berberine (0, 5, 25, 50, and 100 mg/kg) inhibited the CYP1A mRNA expression, thereby inhibiting further the catalytic activity of CYP1A-related ethoxyresorufin-O-deethylase (EROD). Furthermore, both CYP1A expression and EROD activity were further inhibited with increasing berberine concentrations. In addition, the CYP3A expressions at both the mRNA and the protein levels were downregulated by higher berberine concentrations. The catalytic activity of CYP3A-related erythromycin N-demethylase (ERND) was also inhibited by berberine at a dose of no less than 25 mg/kg. Moreover, at the berberine concentration exceeding 25 mg/kg, the inhibition of CYP3A expression and ERND activity increased with increasing berberine concentrations. In vitro experiments were also performed. When berberine was pre-incubated with the crucian carp liver microsomes, it competitively inhibited the corresponding EROD activity with the IC₅₀ of 11.7 μM. However, the ERND activity was slightly inhibited by berberine with the IC₅₀ of 206.4 μM. These results suggest that, in crucian carp, berberine may be a potent inhibitor to CYP1A, whereas the CYP3A inhibition needs a higher concentration of berberine.     

The role of protein tyrosine kinases in CYP1A1 induction by omeprazole and thiabendazole in rat hepatocytes

Benzimidazoles compounds like omeprazole (OME) and thiabendazole (TBZ) mediate CYP1A1 induction differently from classical aryl hydrocarbon receptor (AhR) ligands, 3-methylcholanthrene (3-MC) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). To clarify the involvement of an intracellular signal pathway in CYP1A1 induction by OME and TBZ, the TBZ, OME and 3-MC signal-transducing pathways were compared by using specific protein tyrosine kinase inhibitors in primary culture of rat hepatocytes. The effect of OME and TBZ (75-250 microM) on cytochrome P450 1A1 (CYP1A1) expression was therefore studied in primary cultures of rat hepatocytes after 24 h, 48 h and 72 h of exposure. Both compounds provoked a dose- and time-dependent increase in CYP1A1 (EROD activity, protein and mRNA levels), but OME was less effective at all the concentrations and times tested. The mechanism of benzimidazole-mediated induction of CYP1A1 was investigated by comparison with 3-MC, a prototypical AhR ligand. As expected, OME and TBZ were unable to displace [(3)H]-TCDD from its binding sites to the AhR in competitive binding studies. Moreover, classic tyrosine kinase inhibitor herbimycin A (HA) inhibited the two benzimidazoles-mediated CYP1A1 inductions, but only partially inhibited the 3-MC-mediated one. Another two tyrosine kinase inhibitors, Lavendustin A (LA) and genistein (GEN), had no effect on CYP1A1 induction by benzimidazoles and 3-MC. These results are consistent with the implication of a tyrosine kinase, most probably the Src tyrosine kinase, in the mechanism of CYP1A1 induction in rat hepatocytes.     

Association of CYP3A5*3 polymorphism with development of acute leukemia

BACKGROUND:

CYP3A5 was observed to be an important genetic contributor to inter individual differences in CYP3A-dependent drug metabolism in acute leukemic patients. Loss of CYP3A5 expression was mainly conferred by a single nucleotide polymorphism at 6986A>G (CYP3A5*3). We investigated the association between CYP3A5*3 polymorphism and acute leukemia.

MATERIALS AND METHODS:

Two hundred and eighty nine acute leukemia cases comprising of 145 acute lymphocytic leukemia (ALL), 144 acute myeloid leukemia and 241 control samples were analyzed for CYP3A5*3 polymorphism using PCR-RFLP method. Statistical analysis was performed with SPSS version (15.0) to detect the association between CYP3A5*3 polymorphism and acute leukemia.

RESULTS:

The CYP3A5*3 polymorphism 3/3 genotype was significantly associated with acute leukemia development (χ²- 133.53; df-2, P 0.000). When the data was analyzed with respect to clinical variables, mean WBC, blast % and LDH levels were increased in both ALL and AML cases with 3/3 genotype. The epidemiological variables did not contribute to the genotype risk to develop either AML or ALL.

CONCLUSION:

The results suggest that the CYP3A5*3 polymorphism might confer the risk to develop ALL or AML emphasizing the significance of effective phase I detoxification in carcinogenesis. Association of the polymorphism with clinical variables indicate that the 3/3 genotype might also contribute to poorer survival of the patients.     

In vitro 3-D culture demonstrates incompetence in improving maintenance ability of primary hepatocytes

Primary hepatocytes (PHs) are considered the ‘gold standard’ in drug screening owing to their ability to express many drug-metabolizing enzymes and transporters. Culturing hepatocytes and maintaining their fate in vitro is a major issue since last decade. The main problem with in vitro hepatocytes culture is that they rapidly lose their hepatic morphology and liver-specific functions in culture. Herein, we isolated rat PHs, and cultured them in monolayers (2-D) and spheroids (3-D). The 2-D-cultured PHs exhibited elongated morphology, whereas the 3-D-cultured PHs exhibited spheroid morphology with gradual diameter decrease until 7 days. After 7 days of in vitro culture, PHs were analyzed for the expression of hepatic (Alb, Tf, and Afp) and apoptotic markers (Bax and Bcl2), and co-expression of CYP3A1 and Abumin after 2 and 7 days. Furthermore, in both cultures, PHs were induced with 3-methylcholanthrene (3-MC, Cyp1a-specific inducer) and dexamethasone (Cyp3a-specific inducer) for 48 and 72?h, respectively. The mRNA levels of Cyp1a and Cyp3a were analyzed in induced (3-MC, dexamethasone) and non-induced PHs. After 7 days of in vitro culture, PHs exhibited dramatic downregulation of hepatic marker expression in both cultures. Furthermore, apoptotic marker expression was higher in the 2-D-cultured PHs than 3-D-cultured PHs. The mRNA levels of Cyp1a and Cyp3a indicated higher RNA content in the 2-D-cultured PHs after 48?h of induction. Therefore, we concluded that there was no significant difference between the culture systems, and further studies are required to identify the essential components for in vitro PH culture rather than culture systems.     

Regulation of CYP1A2 by histone deacetylase inhibitors in mouse hepatocytes

Cytochrome P450 1A2 (CYP1A2) is constitutively expressed in the mouse liver, but the constitutive expression progressively declines to an undetectable level in isolated hepatocytes. In this study, CYP1A2 was induced in hepatocytes exposed to the histone deacetylase inhibitors trichostatin A (TSA) and sodium butyrate (SB), but only well after constitutive CYP1A2 expression was silenced. However, cotreatment with the arylhydrocarbon receptor (AhR) ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and either TSA or SB reduced the induction of CYP1A2 with the same time course as TSA or SB increased its induction. These results suggest that histone modification is involved in CYP1A2 regulation in hepatocytes through pathways that are independent of AhR.