Objective: To study the application of different metabolic activation systems in benzo(a)pyrene [B(a)P]-induced human bronchial epithelium cell HBETR transformation.
Methods: In vitro metabolic activations of B(a)P were compared with rat liver S9 fraction mix, overexpression of a key enzyme (P450 CYP1A1), and prior low dose B(a)P (1 micromol/L) induction. Using soft agar assay and tumorigenicity assay, the different metabolic activation systems were compared to the influence on transformation of human bronchial epithelium cell HBETR.
Results: Both immunoblotting and enzyme activity showed that cells overexpressing CYP1A1 (HBETR-1A1) and 48 h after low dose B(a)P induction (HBETR-IN) had high-level expression of CYP1A1. There were no obvious changes in the biology characteristic of these cells. The latencies of cell transformation in HBETR-1A1 and HBETR-IN cells were 11 weeks when cells were treated with B(a)P at concentration of 20 micromol/L, while it took 14 weeks to achieve cell transformation in their control cells. The latencies of malignant transformation in HBETR cells in presence or absence of S9-mix were 14 weeks and 20 weeks, respectively. The efficiencies of cell transformation were in consonance with the protein level of endogenous CYP1A1 enzyme and its enzyme activity.
Conclusion: The three metabolic conditions of the addition of rat liver S9 fraction mix, overexpression of a key enzyme (CYP1A1), and low dose B(a) P induction could enhance the B(a)P metabolic activation and shorten the latency of malignant transformation. In terms of the feasibility, difficulty of manipulation, stability, and reliability, low dose B(a)P induction could seem to be a prospective system used in metabolic activation in comparison with rat liver S9 fraction mix addition.