Development and characterization of a stable epithelial cell line from Muta™Mouse lung

We have isolated and characterized a stable epithelial cell line from Muta™Mouse lung that is a suitable complement to the in vivo assay system. The cells are contact inhibited, forming a flat monolayer, and retain several epithelial/pulmonary characteristics. The genome is stable across more than 50 generations, with a modal chromosome number of 78. Spontaneous rates of micronuclei (19.2 ± 1.4 per 1,000), sister chromatid exchanges (0.25 ± 0.004 per chromosome), and chromosome aberrations (∼ 4%) are lower than, or comparable to, other transgenic cell lines currently used in mutagenicity research. Fluorescence in situ hybridization analyses showed that 80% of cells contain three λgt10 lacZ loci. Slot‐blot analyses indicated that the average cell contains ∼17 transgene monomers. Spontaneous mutant frequency at the lacZ transgene is stable (39.8 ± 1.1 × 10 −5 ), and the direct‐acting mutagens N ‐ethyl‐ N ‐nitrosourea and ICR‐191 yielded increases in mutant frequency of 6.3‐ and 3.2‐fold above control, respectively. Benzo[ a ]pyrene (BaP) exposure increased mutant frequency more than 25‐fold above control and did not require an exogenous metabolic activation mixture. Inhibition of Cyp1A1 by 5 μM α‐naphthoflavone eliminated BaP mutagenesis. Activation and mutation induction by the heterocyclic amine 2‐amino‐1‐methyl‐6‐phenylimidazo[4,5‐ b ]pyridine required a low concentration (0.05% v/v) of exogenous rat liver S9. High activity of α, μ, and π glutathione‐ S ‐transferase isozymes appears to confer resistance to the cytotoxic effects of xenobiotics. The cell line is a suitable complement to the in vivo Muta™Mouse assay, and provides an opportunity for routine in vitro mutagenicity testing using an endpoint that is identical to that employed in vivo. Environ. Mol. Mutagen. 42:166–184, 2003. © 2003 Wiley‐Liss, Inc.