17-alpha-ethinylestradiol and norgestrel in combination induce micronucleus increases and aneuploidy in human lymphocyte and fibroblast cultures

Oral contraceptives are highly efficient and easily administered drugs; however, it must not be forgotten that they are composed of chemical substances which can be classified as potential carcinogens. Testing of a substance for genotoxicity represents a reliable approach both to evaluate the genetic hazard and to obtain information on its possible tumorigenic (cancerogenic) properties. The present study was undertaken to evaluate through carefully planned and controlled investigations the in vitro cytogenetic effects of oral contraceptives (ethynilestradiol and norgestrel mixed in the proportion 1:5) using three different concentrations, with two different durations of treatment (48 and 72 h), on two types of human cells (lymphocytes and fibroblasts) and a series of short-term test procedures: sister chromatid exchange (SCE), micronucleus test (MN), and chromosome aberrations (CA). In addition, the FISH procedure and in vitro anaphase and metaphase preparation analyses were performed. In contrast to CA and SCE frequencies, the frequency of MN in treated blood lymphocytes showed higher values by comparison with the controls, although the difference was statistically significant only for the lowest concentration (P = 0.016). When using pancentromeric alphoid probes, the FISH procedure gave positive signals in more than 85% of micronuclei, clearly indicating that MN may contain whole chromosomes rather than acentric fragments. Unlike the lymphocytes, the fibroblasts showed dose-dependent effects, although those treated with the highest hormone concentrations showed an increased number of highly damaged cells (cytoplasmatic vacuolization, nuclear fragmentation, etc.), a decreased number of anaphase cells, a large number of which were abnormal, and a reduction of mitotic index. In conclusion, our data confirm that hormones do not induce structural chromosome aberrations in lymphocytes and indicate that ethynilestradiol and norgestrel have an aneugenic effect on fibroblast and lymphocyte cultures; FISH analysis on micronuclei from lymphocyte cultures and anaphase preparations from fibroblast cultures support this hypothesis.