Activity and Inducibility of Drug-Metabolizing Enzymes in Immortalized Hepatocyte-like Cells (mhPKT) Derived from al-PK/Tag1 Transgenic Mouse
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文摘
This report describes the establishment and characterization of the mhPKT cell line derived from the liver of a transgenic mouse harboring the simian virus (SV40) large T and small t antigens placed under the control of the 5′ regulatory sequence of the ratl-type pyruvate kinase (l-PK) gene. mhPKT cells had a prolonged life span, expressed the SV40-encoded nuclear large T antigen when grown in glucose-enriched medium, and induced tumors when injected subcutaneously into athymic (nu-nu) mice. Growth on petri dishes or filters yielded multiple layers of cuboid cells, with numerous spaces between adjacent cells that were closed by junctional complexes. These bile canaliculi-like structures exhibited numerous microvilli in which villin, an actin-binding brush-border protein, colocalized with actin. These bile canaliculi-like structures appeared to be functional as they accumulated fluorescein. mhPKT cells conserved the expression of the liver-specific transcription factors HNF1, HNF3, HNF4, and DBP together with substantial levels ofl-PK and albumin but not α-fetoprotein mRNA transcripts. mhPKT cells mainly metabolized testosterone into androstenedione and 6β-hydroxytestosterone, asin vivo.3-Methylcholanthrene and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) markedly increased ethoxyresorufin-O-deethylase activity and the related cytochrome P450 (CYP) 1A1/2 protein, whereas α-naphtoflavone antagonized the TCDD-elicited induction. Phenobarbital slightly increased the CYP2B-mediated activities of pentoxyresorufin-O-depentylase, 2β- and 16β-testosterone hydroxylase. mhPKT cells also had substantial sulfotransferase, UDP-glucuronyltransferase, and glutathioneS-transferase activities. This model may serve as a tool for long-termin vitrostudies of xenobiotic metabolism, potent CYP inducers, and hepatocyte damage due to drugs and other factors.