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

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Martina Klemm, Elke Genschow, Inge Pohl, Christa Barrabas, Manfred Liebsch and Horst Spielmann
ZEBET/ Bundesinstitut für gesundheitlichen Verbraucherschutz und Veterinärmedizin (BgVV), D-Berlin

Germ cell mutagenesis is required by the 7th amendment of the directive 67/548 EEC into the national regulations on existing chemicals. Officially accepted in vivo test systems for stage specific mutagenicity are the dominant lethal (DL) test and the specific locus test (SLT) in mice. An acceptable in vitro alternative designed to address germ cell mutagenesis and discriminate between male and female specific effects 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 EG3 cell line and differentiated fibroblast cells 3T3 were comparatively tested with cytotoxicity assay (MTT test ) and genotoxic studies (SCE-assay) under identical test conditions. The concentration-response curves reflected the female cell line EG3 to be extremely sensitive concerning cytotoxic and genotoxic endpoints. Therefore this cell line was used to classify in vivo genotoxic and non-genotoxic test substances with different potential endpoints. Applying linear discriminant analysis three endpoints were identified for the correct classification (100%) of all test chemicals, namely the SCE200 value (increase of 200% in the mean number of SCEs per metaphase spread) for EG3 (3hrs and 24hrs assay) and the IC50 value for EG3 after 3hrs of exposure to test chemicals.

ALTEX 18(2), 127-130

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