Expression, induction and regulation of the cytochrome P450 monooxygenase system in the rat glioma C6 cell line

Abstract

The cytochrome P450 monooxygenase system consists of NADPH-cytochrome P450 reductase (P450 reductase) and cytochromes P450, which can catalyze the oxidation of a wide variety of endogenous and exogenous compounds. P450 reductase transfers reducing equivalents from NADPH to P450, which in turn catalyzes metabolic reactions. In previous studies, we have used the rat glioma C6 cell line as an in vitro model system and identified the presence of P450 reductase and of cytochrome P450 1A1, 1A2, 2A1, 2B1/2, 2C7, 2D1-5 and 2E1 by reverse transcription followed by polymerase chain reaction (RT–PCR). In C6 cells, the induction of P450 1A1 and 2B1/2 at mRNA level after BA (benzo(a)anthracene) or PB (phenobarbital) treatments was detected. In this study, analysis of microsomal preparations of glioma C6 cells was utilized to demonstrate the presence of P450 2B and P450 reductase at the protein level. ELISAs showed that PB induced P450 2B proteins 12-fold. These experiments further establish that the rat glioma C6 cell line contains an active cytochrome P450 monooxygenase system that can be induced by P450 inducers. We also found that the mRNAs of P450 1A1 and 2B1/2 from glioma C6 cells do not bind to the oligo(dT)-based separation techniques efficiently, suggesting that they may have very short poly(A) tails. The half-lives of P450 1A1 and 2B1/2 mRNA in glioma C6 cells are 1/10 and 1/3 of that in liver, respectively. This may partly contribute to the low expression level of P450s in glial cells. The induction of P450s by BA or PB did not change their mRNA half-lives, indicating the induction may be due to transcriptional regulation. In summary of this study, we believe the presence of the cytochrome P450 monooxygenase system in glial cells of the brain may be important in chemotherapy and carcinogenesis of brain tumors.