Aneugen‐induced micronuclei (MN) in human lymphocytes may be discerned using image analysis techniques when cell‐cycle stage is taken into account

Abstract

We show that for the in vitro cytochalasin-B human lymphocyte micronucleus (MN) test, the quantification of the DNA content of MN and the difference in DNA content between the two macronuclei in the binucleate cells without MN, as measured by image analysis, gives a first estimation of the aneugenic potential of a test compound. Cultures of isolated human lymphocytes were exposed either to gamma-rays as a clastogen or to carbendazim (MBC) as an aneugen. The lymphocytes were stained with Feulgen stain and the MN were analyzed for DNA content with a Magiscan 2A image analyzer. The mean DNA content of MN induced by MBC were statistically higher than gamma-irradiation-induced MN. It was demonstrated that in culture the lymphocytes, as well as the MN, are in different stages of the cell cycle, but this will not affect the discriminating power of the MN DNA content when only G1 cells are considered, or when DNA content of the MN is expressed relative to the total genome. The identification of G1 and G2 cell populations from image analysis data was performed by extrapolation of DNA content data from G1- and G2-sorted lymphocytes with a FacStar plus flow sorter. It was demonstrated that in MBC-treated cells the DNA rearrangement between the macronuclei in binucleates without MN was on the average higher than in gamma-irradiated and untreated cells, which points to aneugenic effects of MBC without the formation of MN. In contrast to DNA content measurements, the area of the MN is not a reliable measure for discriminating clastogens from aneugens.