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Different sensitivity of BALB/c 3T3 cell clones in the response to carcinogens.

Toxicology in vitro : an international journal published in association with BIBRA (2011-06-16)
Annamaria Colacci, Maria Grazia Mascolo, Stefania Perdichizzi, Daniele Quercioli, Antonio Gazzilli, Francesca Rotondo, Elena Morandi, Angela Guerrini, Paola Silingardi, Sandro Grilli, Monica Vaccari
ABSTRACT

Cell transformation assays (CTAs) are currently regarded as the only possible in vitro alternative to animal testing for carcinogenesis studies. CTAs have been proposed as screening tests for the carcinogenic potential of compounds that have no evidence of genotoxicity but present structural alerts for carcinogenicity. We have extensively used the BALB/c 3T3 model based on the A31 cell clone to test single chemicals, complex mixtures and environmental pollutants. In the prevalidation study carried out by ECVAM, the improved protocol is based on BALB/c 3T3 A31-1-1 cells, a clone derived by A31 cells, that is very sensitive to PAH-induced transformation. The present study was performed in the aim to compare the results obtained with the two different clones exposed to different classes of carcinogens. Cells were treated with PAHs (3-methylcholanthrene, benzo(a)pyrene), alkylating agents (melphalan) and aloethanes (1,2-dibromoethane). The induction of cytotoxicity and the onset of chemically transformed foci were evaluated by two experimental protocols, differing for cell seeding density and chemical treatment duration. The A31-1-1 cells showed higher inherent transformation rate after PAHs treatment, but they were insensitive to 1,2-dibromoethane at concentrations that usually induced transformation in A31 cells. As 1,2-dibromoethane is bioactivated to reactive forms able to bind DNA mainly through the conjugation with intracellular glutathione, these results suggested a reduced activity of phase-2 enzymes involved in glutathione conjugation in A31-1-1 cells. Our results give evidence that inherent metabolic capacity of cells may play a critical role in in vitro cell transformation, cautioning against possible misclassification of chemicals.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
1,2-Dibromoethane, 98%
Sigma-Aldrich
1,2-Dibromoethane, ≥99%
Sigma-Aldrich
1,2-Dibromoethane, purum, ≥98.0% (GC)
Supelco
1,2-Dibromoethane solution, certified reference material, 200 μg/mL in methanol
Supelco
1,2-Dibromoethane, PESTANAL®, analytical standard