Ethanol and Isopentanol Increase CYP3A and CYP2E in Primary Cultures of Human Hepatocytes

Kostrubsky, Vsevolod E.; Strom, Stephen C.; Wood, Sheryl G.; Wrighton, Steven; Sinclair, Peter R.; Sinclair, Jacqueline F.

doi:10.1006/abbi.1995.1495

Cited by 90 publications

(50 citation statements)

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“…CYP3A4 but not CYP3A5 expression appeared to be upregulated in the frontal cortex of human alcoholic brain. Previous studies suggested that CYP3A expression is induced by ethanol in liver in vivo and ex vivo in primary hepatocytes (Hoshino and Kawasaki, 1995;Kostrubsky et al, 1995) and in the intestine of moderate alcohol consumers (Liangpunsakul et al, 2005). However, to our knowledge, this is the first report of induction of CYP3A by alcohol in the human brain.…”

Section: Discussionmentioning

confidence: 58%

Differential Expression of Human Cytochrome P450 Enzymes from the CYP3A Subfamily in the Brains of Alcoholic Subjects and Drug-Free Controls

et al. 2013

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Cytochrome P450 enzymes are responsible for the metabolism of most commonly used drugs. Among these enzymes, CYP3A forms mediate the clearance of around 40-50% of drugs and may also play roles in the biotransformation of endogenous compounds. CYP3A forms are expressed both in the liver and extrahepatically. However, little is known about the expression of CYP3A proteins in specific regions of the human brain. In this study, form-selective antibodies raised to CYP3A4 and CYP3A5 were used to characterize the expression of these forms in the human brain. Both CYP3A4 and CYP3A5 immunoreactivity were found to varying extents in the microsomal fractions of cortex, hippocampus, basal ganglia, amygdala, and cerebellum. However, only CYP3A4 expression was observed in the mitochondrial fractions of these brain regions. Nterminal sequencing confirmed the principal antigen detected by the anti-CYP3A4 antibody in cortical microsomes to be CYP3A4. Immunohistochemical analysis revealed that CYP3A4 and CYP3A5 expression was primarily localized in the soma and axonal hillock of neurons and varied according to cell type and cell layer within brain regions. Finally, analysis of the frontal cortex of chronic alcohol abusers revealed elevated expression of CYP3A4 in microsomal but not mitochondrial fractions; CYP3A5 expression was unchanged. The site-specific expression of CYP3A4 and CYP3A5 in the human brain may have implications for the role of these enzymes in both normal brain physiology and the response to drugs.

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Section: Discussionmentioning

confidence: 58%

Differential Expression of Human Cytochrome P450 Enzymes from the CYP3A Subfamily in the Brains of Alcoholic Subjects and Drug-Free Controls

et al. 2013

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“…Although CYP2E1 is widely accepted to be the sole form of cytochrome P450 responsible for APAP hepatotoxicity (for review, see Bromer and Black, 2003), we previously found that EIP increased APAP hepatotoxicity in Cyp2e1(Ϫ/Ϫ) mice, showing that CYP2E1 is not essential in alcohol-mediated increases in APAP hepatotoxicity (Sinclair et al, 2000c). Alcohol has been shown to induce CYP3A along with CYP2E1 in rats (Louis et al, 1994;Roberts et al, 1995) and in primary cultures of human and rat hepatocytes (Sinclair et al, 1991;Kostrubsky et al, 1995a). TAO, an inhibitor of CYP3A, protected rats from alcohol-mediated increases in APAP hepatotoxicity, suggesting a role of CYP3A (Kostrubsky et al, 1997;Sinclair et al, 2000b).…”

Section: Discussionmentioning

confidence: 94%

Role of CYP3A and CYP2E1 in Alcohol-Mediated Increases in Acetaminophen Hepatotoxicity: Comparison of Wild-Type andCyp2e1(–/–) Mice

Wolf¹,

Wood²,

Allard³

et al. 2007

Drug Metab Dispos

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ABSTRACT:CYP2E1 is widely accepted as the sole form of cytochrome P450 responsible for alcohol-mediated increases in acetaminophen (APAP) hepatotoxicity. However, we previously found that alcohol [ethanol and isopentanol (EIP)] causes increases in APAP hepatotoxicity in Cyp2e1(؊/؊) mice, indicating that CYP2E1 is not essential. Here, using wild-type and Cyp2e1(؊/؊) mice, we investigated the relative roles of CYP2E1 and CYP3A in EIP-mediated increases in APAP hepatotoxicity. We found that EIP-mediated increases in APAP hepatotoxicity occurred at lower APAP doses in wild-type mice (300 mg/kg) than in Cyp2e1(؊/؊) mice (600 mg/kg). Although this result suggests that CYP2E1 has a role in the different susceptibilities of these mouse lines, our findings that EIP-mediated increases in CYP3A activities were greater in wild-type mice compared with Cyp2e1(؊/؊) mice raises the possibility that differential increases in CYP3A may also contribute to the greater APAP sensitivity in EIP-pretreated wild-type mice. At the time of APAP administration, which followed an 11 h withdrawal from the alcohols, alcohol-induced levels of CYP3A were sustained in both mouse lines, whereas CYP2E1 was decreased to constitutive levels in wild-type mice. The CYP3A inhibitor triacetyloleandomycin (TAO) decreased APAP hepatotoxicity in EIP-pretreated wild-type and Cyp2e1(؊/؊) mice. TAO treatment in vivo resulted in inhibition of microsomal CYP3A-catalyzed activity, measured in vitro, with no inhibition of CYP1A2 and CYP2E1 activities. In conclusion, these findings suggest that both CYP3A and CYP2E1 contribute to APAP hepatotoxicity in alcohol-treated mice.Acetaminophen (APAP) is considered to be a relatively safe drug and is widely prescribed and used for its analgesic and antipyretic properties. The first reports of hepatotoxicity in humans resulting from APAP overdose were in 1966 ( Thomson and Prescott, 1966). Today, APAP is a frequent cause of liver failure and the leading cause of death due to accidental and deliberate overdose (for review, see Bromer and Black, 2003). APAP is mainly glucuronidated and sulfated in the liver by phase II metabolic pathways, leading to excretion of the drug. However, APAP is also metabolized by certain forms of cytochromes P450 (P450s) to N-acetyl-p-benzoquinone imine (NAPQI), a highly electrophilic metabolite that is considered to be responsible for triggering the ensuing liver damage. CYP1A2, CYP2E1, and CYP3A are three forms of rat and human P450s active in the conversion of APAP to NAPQI, with CYP2E1 and CYP3A having greater intrinsic activity than CYP1A2 (Patten et al., 1993;Thummel et al., 1993). Reduced glutathione (GSH) reacts with NAPQI, resulting in the inactivation of this reactive metabolite. When GSH levels become depleted, liver damage can occur as a result of the

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“…After removing the medium, cells were harvested in a phosphate buffer (0.1 M, pH 7.4) and stored at Ϫ80°C for protein determination (Lowry et al, 1951) and detection of immunoreactive P450 protein. Immunochemical detection of all P450 isoforms was performed as previously described (Kostrubsky et al, 1995) using 18 g of sonicated pooled proteins. CYP3A4 protein was detected with a rabbit antihuman antibody, CYP2C9 protein was detected with a rabbit anti-human CYP2C9 antibody, and CYP1A2 and CYP2D6 were detected with a goat antihuman antibody.…”

Section: St Johnјs Wort and Drug-metabolizing Enzymesmentioning

confidence: 99%

Induction and Inhibition of Cytochromes P450 by the St. John's Wort Constituent Hyperforin in Human Hepatocyte Cultures

Komoroski

Zhang²,

Cai³

et al. 2004

Drug Metab Dispos

158

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ABSTRACT:St. John's wort extract (SJW) (Hypericum perforatum L.) is among the most commonly used herbal medications in the United States. The predominance of clinical reports indicates that SJW increases the activity of cytochrome P450 3A4 (CYP3A4) enzyme and reduces plasma concentrations of certain drugs. Although the inductive effect of SJW on CYP3A4 is clear, other reports indicate that SJW constituents may have, to a small degree, some enzyme inhibitory effects. Therefore, we sought to study the induction and inhibition effects of the constituents of SJW on CYP3A4 in the human hepatocyte model. Moreover, most research has focused on the induction of CYP3A4 by SJW with little attention paid to other prominent drug-metabolizing enzymes such as CYP1A2, CYP2C9, and CYP2D6. To examine the effects of SJW on CYP1A2, CYP2C9, CYP2D6, as well as CYP3A4, hepatocytes were exposed to hyperforin and hypericin, the primary constituents of SJW extract. Hepatocytes treated with hypericin or hyperforin were exposed to probe substrates to determine enzyme activity and protein and RNA harvested. Hyperforin treatment resulted in significant increases in mRNA, protein, and activity of CYP3A4 and CYP2C9, but had no effect on CYP1A2 or CYP2D6. Acute administration of hyperforin at 5 and 10 M 1 h before and along with probe substrate inhibited CYP3A4 activity. Hypericin had no effect on any of the enzymes tested. These results demonstrate that with chronic exposure, the inductive effect of SJW on drug-metabolizing enzymes predominates, and human hepatocyte cultures are a versatile in vitro tool for screening the effect of herbal products on cytochrome P450 enzymes.In 2002, sales of botanical supplements in the United States reached nearly $293 million dollars. St. John's wort accounted for 15 million U.S. dollars in sales, making it the fourth highest grossing botanical supplement (Blumenthal, 2003). Several clinical studies have demonstrated the effectiveness of St. John's wort compared with conventional therapy in the treatment of mild to moderate depression (Linde et al., 1996;Wheatley, 1997).Marketed St. John's wort, an extract of the flowering portion of the plant Hypericum perforatum L., is a mixture of a number of biologically active, complex compounds. At 0.3 mg per capsule, the naphthodianthrone hypericin is used as a means of standardization of the marketed product. The phloroglucinol hyperforin, the most plentiful lipophilic compound in the extract, is a potent reuptake inhibitor of serotonin, norepinephrine, and dopamine (Muller et al., 1998).Several recent reports have documented decreased blood/plasma levels of cytochrome P450 3A4 (CYP3A4) substrates, such as indinavir and cyclosporin A, in patients concomitantly taking St. John's wort (Piscitelli et al., 2000;Ahmed et al., 2001). Similar observations have been documented for digoxin, a substrate of the intestinal transporter P-glycoprotein (P-gp 4 ). Additional in vivo evidence has demonstrated that St. John's wort increased CYP3A4 and P-gp protein levels in rats (Dur...

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Ethanol and Isopentanol Increase CYP3A and CYP2E in Primary Cultures of Human Hepatocytes

Cited by 90 publications

References 0 publications

Differential Expression of Human Cytochrome P450 Enzymes from the CYP3A Subfamily in the Brains of Alcoholic Subjects and Drug-Free Controls

Differential Expression of Human Cytochrome P450 Enzymes from the CYP3A Subfamily in the Brains of Alcoholic Subjects and Drug-Free Controls

Role of CYP3A and CYP2E1 in Alcohol-Mediated Increases in Acetaminophen Hepatotoxicity: Comparison of Wild-Type andCyp2e1(–/–) Mice

Induction and Inhibition of Cytochromes P450 by the St. John's Wort Constituent Hyperforin in Human Hepatocyte Cultures

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