The Ah receptor, cytochrome P450IA1 mRNA induction, and aryl hydrocarbon hydroxylase in a human lymphoblastoid cell line

Abstract

The immunosuppressive and carcinogenic effects of aryl hydrocarbons such as 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) and 3-methylcholanthrene (MC) on B lymphocytes of adult rodents and the induction of cytochrome P450IA1 and aryl hydrocarbon hydroxylase (AHH) in human mitogenactivated lymphocytes and B-lymphoblastoid cell lines are believed to be mediated by the Ah receptor. However, there has not been a direct demonstration or characterization of the Ah receptor in denned populations of any of these cells. We report here the detection and characterization of an abundant, high-affinity B lymphocyte Ah receptor in the AHH-inducible human B lymphoblastoid cell line BCR-5. Our results represent the first characterization of a human lymphocyte receptor in a well-defined lymphocyte population. Sucrose density gradient analysis of BCR-5 cytosols incubated with [ 3H]TCDD revealed a characteristic 9 S specific binding peak. The maximum concentration of Ah receptor was about 200 fmol/mg protein. Specific binding to the Ah receptor was also detected with [ 3H]MC and, to a lesser extent, with [ 3H]benzolalpyrene. The apparent binding affinity ( K d ) for [ 3H]TCDD (determined by saturation analyses) was about 5 nM. A specific [ 3 H] TCDD-Ah receptor complex which sedimented at 5 S was extracted from nuclei of BCR-5 cells incubated at 37° with [ 3H]TCDD. The Ah receptor of BCR-5 cells is thus similar in characteristics to that identified in other cell lines. When BCR-5 cells were exposed in culture for 24 hr to increasing concentrations of benz[ a]anthracene there was a concentrationdependent increase in induction and a good correlation ( r = 0.98) between the level of induced AHH activity and the relative abundance of cytochrome P450IA1 mRNA. The human B lymphoblastoid cell line BCR-5, therefore, has a complete regulatory mechanism for Ah receptor-mediated induction of cytochrome P450IA1 that is essentially the same as that which has been well established in many rodent species. The accessibility of human blood lymphocytes and the ease of establishment of B lymphoblastoid cell lines from any donor provide a source of pure cultures of human B lymphocytes which can be grown continuously in vitro for the study of mechanisms related to Ah receptor-mediated cytochrome P450IA1 induction, immunosuppression and carcinogenesis.