Permanent embryonic mouse germ cell-lines, an in vitro alternative to in vivo germ cell mutagenicity tests

Permanent embryonic mouse germ cell-lines, an in vitro alternative to in vi vo germ cell mutagenicity tests Germ cell mutagenesis is required by the 7( th) amendment of the directive 67/548 EEC into the national regulations on existing chemicals. Officially accepted in vivo test systems for stage spec ific mutagenicity al-e the dominant lethal (DL) test and the specific locus test (SLT) in mice. An acceptable in vitro alter-native designed to addres s germ cell mutagenesis and discriminate between male and female specific e ffects is not available at present. In order to offer a sensitive and predictive in vitro method to assess the genotoxic potential of chemical agents on male and female reproduction, we established primordial germ (PG) cell-derived permanent embryonic germ (EG) cell lines of the mouse (strain BALB/cJ). The differences in developmental sensitivity of the EG, cell line and differentiated fibroblast cells 3T3 w ere comparatively tested with cytotoxicity assay (MTT test) and genotoxic s tudies (SCE-assay) under identical rest conditions. The concentration-respo nse curves reflected the female cell line EG, to be extremely sensitive con cerning cytotoxic and genotoxic endpoints Therefore this cell line was used to classify in vivo genotoxic and non-genotoxic test substances with diffe rent potential endpoints. Applying linear discriminant analysis three endpo ints were identified for the con ect classification (100%) of all test chem icals, namely the SCE200 value (increase of 200% in the mean number of SCEs per metaphase spread) for EG(3) (3hrs and 24 hrs assay) and the IC50 value for EG(3) after 3hrs of exposure to test chemicals.