Induction of cytochrome P-450s by 2-acetylaminofluorene in primary cultures of adult rat hepatocytes

Abstract

The induction of cytochrome P-450 by 2-acetylaminofluorene, a potent hepatocarcinogen, was studied in primary cultures of adult rat hepatocytes. Addition of 2-acetylaminofluorene to the hepatocyte cultures induced the P-450IA 1-specific ethoxyresorufin O-deethylase activity in dose- and time-dependent manner. The P-450IIB-specific pentoxyresorufin O-dealkylase activity was also induced, though to an even lesser degree, at high concentration of 2-acetylaminofluorene. To elucidate the mechanism of the induction of ethoxyresorufin O-deethylase activity by 2-acetylaminofluorene, we performed Northern blot analysis using a specific oligonucleotide probe recognizing P-450IA 1/2 mRNA and found that the induction is not mediated by the accumulation of the specific mRNA. The result of experiment using cycloheximide, an inhibitor of protein biosynthesis, in which 0.5$\mu$M cycloheximide completely blocked the induction of P-450IA1-specific ethoxyresorufin O-deethylase activity indicated that protein biosynthesis is required for the induction in response to 2-acetylaminofluorene and this was supported by the enzyme-linked immunosorbent assay showing actual increase in P-450IA1 protein.