2,3,7,8 tetrachlorodibenzo-p-dioxin induction of cytochrome P4501A in cultured rat and human hepatocytes

We report here a novel observation that 2,3,7,8-tetracholorodibenzo-p-dioxi n (TCDD) induced predominantly cytochrome P4501A1 (CYP1A1) in rat-hepatocyt es and predominantly CYP1A2 in human hepatocytes. As part of our research p rogram to evaluate species-differences in response to CYP inducers, We Stud ied the effects of TCDD on CYP1A activity, protein, and gene expression in primary cultures of rat and human hepatocytes. TCDD was found to induce CYP 1A activity, measured as ethoxyresorufin-O-deethylase (EROD) activity, in b oth rat and human hepatocytes, TCDD induction of EROD activity in human hep atocytes (2-5 fold of concurrent solvent control), was significantly lower than that found in rat hepatocytes (approximate to 20-fold of concurrent so lvent control). Two structural analogs of TCDD, 2,3,7,8-tetrachlorodibenzof uran (TCDE) and 6-nitro-1,3,8-trichlorodibenzofuran (6-NCDF), were also eva luated. As observed for TCDD, human hepatocytes consistently showed a lower response than rat hepatocytes. As most TCDD-related effects are believed t o be mediated via binding of the TCDD-Ah receptor (AhR) complex to DNA, nuc lear AhR levels were measured in rat and human hepatocytes after TCDD treat ment. We found that the nuclear AhR levels in TCDD-treated rat hepatocytes were approximate to 4 times higher than found in TCDD-treated human hepatoc ytes. However, the estimated binding affinity of [H-3]TCDD to nuclear AhR f rom rat hepatocytes was similar. The species difference in response to TCDD was further evaluated by analysis of CYP1A1 and CYP1A2 mRNA levels using N orthern analysis, and P4501A1 and 1A2 protein levels using Western immunobl otting. Results showed that, at both gene expression and protein levels, TC DD induced predominantly CYP1A1 in rat hepatocytes and CYP1A2 in human hepa tocytes. (C) 2000 Elsevier Science Ireland Ltd. All rights reserved.