Induction of micronuclei by stilbene-type and steroidal estrogens in Syrian hamster embryo and ovine seminal vesicle cells in vitro

Abstract

The induction of micronuclei (MN) is a known effect of the carcinogenic estrogen diethylstilbestrol (DES). We have now tested the time course and dose dependence of MN induction by DES and its analogs 3',3"-DES, indenestrol A (IA), indenestrol B (IB) or by the steroidal estrogen 17 beta-estradiol (E2) and by the clastogenic compound 4-nitroquinoline-N-oxide (NQO) in two primary mammalian cell culture systems. All compounds induced MN in Syrian hamster embryo and ovine seminal vesicle cells with compound-specific time courses and dose dependences. DES induced a maximum MN frequency 12 h post treatment, whereas with all other estrogens the highest MN frequency was observed 3-6 h after removal of the compound. The maximum MN frequency after NQO treatment occurred at 24 h or later. Of the stilbene estrogens tested, only DES caused an increase of the mitotic index. Further characterization of the MN by indirect immunofluorescence microscopy using CREST antikinetochore antibodies revealed that 92-99% of the DES-induced MN but only 0-2% of the NQO-induced MN contained CREST-reactive kinetochores. Since kinetochore-positive MN are indicative of whole chromosomes/chromatids and kinetochore-negative MN of acentric chromosomal fragments, our findings support the view that DES acts as a pure aneuploidogen and NQO as a pure clastogen in the two cell systems. In the case of 3',3"-DES, IA, IB and E2, 41-68% of the induced MN contained CREST-reactive kinetochores. As the time courses of MN induction are not compatible with those of clastogenic agents, it is proposed that these estrogens induce MN containing chromatids/chromosomes with altered kinetochore structures.