Phenobarbital induction of CYP2B1/2 in primary hepatocytes: Endocrine regulation and evidence for a single pathway for multiple inducers

Phenobarbital (PB) and many structurally unrelated chemicals induce the pro tein and mRNA of P450 cytochromes CYP2B1, CYP2B2, CYP3A1, and specific phas e II enzymes to a greater extent in Fischer 344 CF344) than in Wistar Furth (WF) female rats. This sex- and strain-dependent polymorphism can be partl y attributed to suppressive effects of thyroid hormone (TH) on WF but not F 344 females. We show here that this strain difference was largely retained in primary hepatocyte cultures and could be resolved into two components; ( 1) Expression of PB-inducible genes-WE hepatocytes had inherently lower bas al and PB-induced levels of CYP2B1/2B2 protein and mRNA and UDPGT mRNA; and (2) TH sensitivity-in WF hepatocytes, PB induction, but not basal expressi on, of CYP2B1/2B2 was three- to fivefold more susceptible to inhibition by TH when the hormone was added to the medium. This second component explains the selective effect of in vivo treatment with methimazole, which lowers c irculating TH and partially improves PB induction in WF female rats. Follow ing transfection of a reporter construct containing a PB-responsive unit (P BRU), the plasmid was activated by PB to similar extents in hepatocytes fro m both rat strains. TH treatment did not inhibit PB-mediated induction of t he plasmid in either cell type, Thus, neither of the components determining the strain polymorphism are linked to trans-activating factors contributin g to this PBRU activity. The PB-like inducers, 2,2',4,4',5,5'-hexachlorobip henyl (HCB) and 1,1-dichloro-2,2-bis(p-chlorophenyl)ethane (o,p-DDD), propo rtionally induced the CYP2B1/2B2 and UDPGT genes and activated the plasmid (HCB = PB > DDD). CYP2B1/2B2 expression following induction by PB and HCB w as subject to identical patterns of inhibition by okadaic acid, cAMP, and G H. Together, these data suggest that PB-like inducers utilize the same poly morphic pathway to affect the same PBRU-activating factors. (C) 1999 Academ ic Press.