A BIOMARKER, P450 RGS, FOR ASSESSING THE INDUCTION POTENTIAL OF ENVIRONMENTAL-SAMPLES

In this study we describe the use of a transgenic cell line for the id entification of potentially toxic compounds in test solutions and envi ronmental samples. The reporter gene system (RGS), derived from a huma n liver cancer cell line, has been engineered such that the CYP1A1 gen e, when activated by an inducer compound, will produce luciferase inst ead of P450. Eighteen hours after application of an inducer the reacti on is stopped by rinsing, the cells lysed, and the cytoplasm measured for luminescence and protein content (for normalization). Induction by such compounds as dioxin, dioxin-like PCB congeners (coplanars), and polycyclic aromatic hydrocarbons (PAHs) infers these xenobiotics are p resent at levels that are potentially toxic, carcinogenic, or mutageni c to organisms. Multiple wells with attached cells in 2 ml of media we re inoculated with various concentrations of toxicants in organic solv ents (including DMSO, toluene, and dichloromethane). Volumes tested su ccessfully thus far are 2 to 20 mu l of solvent. Solvent extracts (usi ng standard extraction methods) of aquatic sediments were directly app lied to the assay system. Test results show significant RGS induction from concentrations of inducer compound that, if present in a typical 40-g sediment sample, would be (in ng/g or ppb): 0.05 for dioxin; 10 t o 1,000 for a range of coplanar PCB congeners; about 2,000 for several Aroclor mixtures; and 300 for a mixture of PAHs. A mixture of pestici des at concentrations about four times the PAH levels did not produce an induction response. Results presented here indicate that testing fo r the presence of potentially toxic chemicals in environmental samples could be performed rapidly, with sensitivity and specificity using th is RGS cell line.