Relationship between mRNA levels quantified by reverse transcription-competitive PCR and metabolic activity of CYP3A4 and CYP2E1 in human liver

Reverse transcription-competitive polymerase chain reaction is a powerful a nd sensitive tool for quantifying the absolute amount of mRNA. Using this m ethod with beta-actin as the standard, we measured the mRNA level of CYP3A and CYP2E1 isoforms in human livers. We also determined the metabolic activ ities for both CYP isoforms. The absolute amounts of CYP3A4 and CYP2E1 mRNA in 15 liver tissues ranged from 15 to 5127 and 15163 to 69289 copies/10(4) copies of beta-actin (341- & 3.6-fold), respectively. The testosterone 6 b eta-hydroxylation for CYP3A4 and chlorzoxazone 6-hydroxylation activity for CYP2E1 ranged from 30 to 505 pmol/mg/min (16-fold) and from 0.59 to 2.73 n mol/mg/ml (3.6-fold), respectively. The correlation between the mRNA level and activity of CYP3A4 was significant (r = 0.94), while there was no signi ficant correlation for CYP2E1 (r = 0.04). In conclusion, we observed a sign ificant correlation between enzyme activity and mRNA expression for CYP3A4 but not for CYP2E1. This fact indicates that CYP2E1, in addition to being l ess variable between individuals than CYP3A4, differs in its regulation mec hanism. (C) 1999 Academic Press.