Extruded micronuclei induced by colchicine or acrylamide contain mostly lagging chromosomes identified in paintbrush smears by minor and major mouse DNA probes

Abstract

In the mouse bone marrow micronucleus assay, it was studied whether micronuclei (MN) could be expelled from polychromatic erythrocytes (PCE) in a similar way to the main nucleus. To avoid the disrupting centrifugation step of the conventional bone marrow preparation procedure, the paintbrush technique was used in the present experiments. With May-Grunwald-Giemsa staining of paintbrush slides, 5% of the colchicine (COL)-induced MN were found attached to the outside membranes of PCE and were regarded as extruded. Of the acrylamide (AA)-induced MN, 22% were extruded. After fluorescence in situ hybridization (FISH) of a total of 300 MN per chemical treatment with the mouse minor and major satellite DNA probes, 9.7% MN were extruded in the COL group and 8.3% MN were extruded in the AA group. FISH showed that 76% of the retained COL-induced MN were signal-positive, indicating that they contained entire chromosomes. With AA, 29% minor-positive and 28.3% major-positive retained MN were found, confirming its known clastogenicity. However, the observed frequency of signal-positive MN (1.7 MNPCE(pos)/1000 PCE) in the AA group was about three times higher than in the control (0.5 MNPCE(pos)/1000 PCE) which indicates that AA has aneugenic potential. FISH analysis of the extruded MN showed 72-100% major as well as minor signals. It is concluded that expelled MN contain mostly entire chromosomes.