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Embryotoxicity testing is either performed in vivo using pregnant animals or in vitro on cultured embryos or embryonic tissues and cells from pregnant animals. Both for in vivo and for in vitro testing, pregnant animals have to be sacrified (Doetschmann et al, 1985). Taking advantage of the potential of embryonic stem cells to differentiate in culture, a new in vitro embryotoxicity test with permanent cell lines from the mouse was proposed, the embryonic stem cell test (Spielmann et al, 1996). This test is based on the determination of the most important embryotoxic parameters. The inhibition of differentiation is combined with the study of differences in sensitivity between embryonic and adult tissues to cytotoxic damage.
Two permanent mouse cell lines are used, ES cells (D3), to represent embryonic tissue, and fibroblasts (3T3 cells), to represent adult tissue. The test has been developed only after it was found that ES cells can be maintained in the undifferentiated stage in the presence of the cytokine leukemia inhibiting factor (LIF). When released from the undifferentiated stage, ES cells will form embryo bodies (Ebs) and differentiate under appropriate conditions into the major embryonic tissues.
Cytotoxicity data show that ES cells are more sensitive to toxic agents than adult cells (Laschinski et al, 1991). The inhibition of the differentiation of ES cells and the inhibition of the growth of ES cells and 3T3 cells are the selected endpoints in the EST for predicting the embryotoxic potential of chemicals.
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