Quantification of the expression and inducibility of 12 rat cytochrome P450 isoforms by quantitative RT–PCR

✍ Scribed by Etienne Caron; Nathalie Rioux; Olivier Nicolas; Hugo Lebel-Talbot; Bettina A. Hamelin

• Publisher: John Wiley and Sons
• Year: 2006
• Tongue: English
• Weight: 823 KB
• Volume: 19
• Category: Article
• ISSN: 1095-6670
• DOI: 10.1002/jbt.20103

✦ Synopsis

Abstract

The administration of xenobiotics may significantly alter the expression of cytochromes P450 (CYPs), thereby leading to potentially toxic cellular, physiologic, and pharmacologic responses. Indeed, an important task in the development of new therapeutic entities is to evaluate efficiently and quantitatively their potential effects on the expression level of different CYPs. In this report, reverse transcriptase polymerase chain reaction (RT–PCR) was used to measure basal and induced mRNA of a wide range of rat CYP isoforms. Rats (n = 3 per treatment) were treated with five prototype inducers of CYP isoforms or with vehicle only. RT and PCR efficiencies were determined using appropriate RNA and DNA standards. Messenger RNA was quantified by PicoGreen standard curves and normalized to cyclophilin. Quantitative RT–PCR was used successfully to demonstrate that CYP isoforms were induced at the mRNA level following drug administration. Notably, phenobarbital resulted in significant induction of CYP2B1, CYP2B2, CYP2C6, CYP2C13, CYP2E1, CYP3A1, and CYP3A2. 3‐Methylcholanthrene induced CYP1A1, CYP1A2, and CYP1B1. CYP2C11 expression was highly variable and suppressed by pyridine, whereas the expression of CYP2E1 was suppressed by dexamethasone. We demonstrated that quantitative RT–PCR can be used to evaluate efficiently the effect of compounds on the expression of a wide range of CYP isoforms. The technique is advantageous over others in that it is very sensitive, efficient and applicable to highly homologous CYP isoforms. © 2005 Wiley Periodicals, Inc. J Biochem Mol Toxicol 19:368–378, 2005; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20103