COMPARATIVE-ANALYSIS OF CYTOCHROME P4503A INDUCTION IN PRIMARY CULTURES OF RAT, RABBIT, AND HUMAN HEPATOCYTES

We previously demonstrated that induction of hepatic cytochrome P4503A (CYP3A) immunoreactive protein is a response in rats, but not rabbits , treated with the antiglucocorticoid, pregnenolone 16 alpha-carbonitr ile and in rabbits, but not rats, treated with rifampicin. These strik ing interspecies differences in response to CYP3A inducers prompted us to compare the effects of a variety of agents on CYP3A expression in primary cultures of hepatocytes from rats, rabbits, and humans, mainta ined under nearly identical conditions on Matrigel. We used treatment with dexamethasone, the most effective inducer of CYP3A mRNA and CYP3A immunoreactive protein in cultures of rat hepatocytes, to define the 100% response. As expected from their effects in vivo, incubations of cultures with medium containing pregnenolone 16 alpha-carbonitrile or rifampicin induced CYP3A mRNA to high levels exclusively in rat hepato cytes or rabbit hepatocytes, respectively. Pregnenolone 16 alpha-carbo nitrile treatment also did not induce CYP3A immunoreactive protein in rabbit hepatocytes, although rifampicin treatment did increase CYP3A i mmunoreactive protein levels in rat hepatocyte cultures. Additions of phenobarbital to the cultures induced CYP3A mRNA and CYP3A immunoreact ive protein to a greater extent in rabbit hepatocytes (94-108% of the dexamethasone response) than in rat hepatocytes (38-57% of the dexamet hasone response). In primary cultures of human hepatocytes, dexamethas one and phenobarbital treatments induced CYP3A mRNA (greater than or e qual to 4.4- and 1.9-fold, respectively, over the amounts of CYP3A mRN A in incubated control cultures). In each preparation of cultured huma n hepatocytes examined, rifampicin treatment increased the amount of C YP3A mRNA (1.5- to 8.5-fold over untreated controls), whereas pregneno lone 16 alpha-carbonitrile treatment induced CYP3A mRNA in only 2 of 4 human cultures tested, raising the possibility of heterogeneity among humans in this response. Incubation with RU-486 or trans-nonachlor (e ach at 10(-5) M) induced CYP3A gene products in cultured hepatocytes f rom all three species. Likewise, treatment of the cultures with lovast atin, a hypocholesterolemic agent reported to elevate CYP3A mRNA level s in cultured human hepatocytes, increased the amounts of CYP3A mRNA a nd CYP3A immunoreactive protein in rabbit (39-58% of the dexamethasone response) and in rat (11-35% of the dexamethasone response) hepatocyt es. In contrast, incubation of cultures with spironolactone, cyprotero ne acetate, or clotrimazole resulted in species-specific effects, indu cing CYP3A gene products to high levels in hepatocytes from rats and h umans, but not rabbits. We conclude that there are dramatic interspeci es differences in CYP3A induction, intrinsic to the hepatocyte, that l ikely preclude the use of a single type of laboratory animal from reli ably predicting the response in humans.