Abstract: OBJECTIVE: To explore the developmental toxicity of genistein (GEN) by micromass cultures of rats limb bud (LB) and midbrain (MB) cells, and investigate its possible mechanisms. METHODS: Micromass cultures of LB and MB were exposed to GEN with a series of concentrations (0, 0.94, 1.875, 3.75, 7.5 and 15 μg/ml). The effect of GEN on cell proliferation was detected by neutral red uptake; the effect of GEN on LB and MB differentiation was assessed by Alcian Blue Staining and Image Analysis, respectively. Cell cultures were pretreated by ICI182780 (0.1, 0.5 and 1 μmol/L), and then with GEN (7.5 μg/ml) added, in order to observe the role of estrogen receptor pathway in the developmental toxicity induced by GEN. RESULTS: For micromass cultures of LB, IC50-P(cell proliferation)and IC50-D (cell differentiation) of GEN were 5.4 μg/ml and 4.8 μg/ml, respectively. For micromass cultures of MB, they were 6.2 μg/ml and 7.1 μg/ml (differentiation judged by number of foci)/5.3 μg/ml (differentiation judged by area of foci), respectively. The ratio IC50-P/IC50-D all approximated to 1. The developmental toxicity induced by GEN with 7.5 μg/ml could not be changed by ICI182780 pre-treatment. CONCLUSION: According to the discrimination rules of Flint and European Center for the Validation of Alternative Methods (ECVAM), and in the light of the human exposure level, GEN was regarded as a strong teratogen, with potential risk toward human. The results indicated that the developmental toxicity GEN may not be mediated by estrogen receptor (ER) pathway.

Key words: genistein, micromass cultures, developmental toxicity, estrogen receptor