3-METHYLCHOLANTHRENE-MEDIATED INDUCTION OF CYTOCHROME P4501A2 IN HUMAN HEPATOMA HEPG2 CELLS AS QUANTIFIED BY THE REVERSE TRANSCRIPTION-POLYMERASE CHAIN-REACTION

The feasibility of using HepG2 human hepatoma cells to study the regul ation of the expression of the cytochrome P4501A2 gene (CYP1A2) was ex amined. The reverse transcription-polymerase chain reaction (RT-PCR) a ssay revealed that HepG2 cells constitutively express CYP1A2 and are a ble to respond to 3-methylcholanthrene (3MC) by an induction of CYP1A2 mRNA. In these studies, selected sequences from intron-exon junctions were used as mRNA-specific primers for both CYP1A2 and the beta-actin gene. The level of induction was quantitated based on two parameters within the exponential phase of the amplification: the difference in t he number of cycles that yields the same level of amplification and th e efficiency of the PCR reaction. Using this method, it was estimated that the CYP1A2 steady-state mRNA level increased to a maximum of 12-f old at 24 h after exposure of the cells to 3MC. Both a reduction in ba sal expression as well as an increased accumulation of CYP1A2 mRNA app eared responsible for the overall induction at 24 and 48 h. These resu lts suggested that the HepG2 cell line would be appropriate for studyi ng the regulation of CYP1A2 expression. (C) 1994 Academic Press, Inc.