ANEUGEN-REDUCED MICRONUCLEI (MN) IN HUMAN-LYMPHOCYTES MAY BE DISCERNED USING IMAGE-ANALYSIS TECHNIQUES WHEN CELL-CYCLE STAGE IS TAKEN INTO ACCOUNT

We show that for the in vitro cytochalasin-B human lymphocyte micronuc leus (MN) test, the quantification of the DNA content of MN and the di fference in DNA content between the two macronuclei in the binucleate cells without MN, as measured by image analysis, gives ct first estima tion of the aneugenic potential of a test compound. Cultures of isolat ed human lymphocytes were exposed either to gamma-rays os a clastogen or to carbendazim (MBC) as an aneugen. The lymphocytes were stained wi th Feulgen stain and the MN were analyzed for DNA content with a Magis can 2A image analyzer. The mean DNA content of MN induced by MBC were statistically higher than gamma-irradiation-induced MN. It was demonst rated that in culture the lymphocytes, as well os the MN, are in diffe rent stages of the cell cycle, but this will not affect the discrimina ting power of the MN DNA content when only G1 cells ore considered, or when DNA content of the MN is expressed relative to the total genome. The identification of GT and G2 cell populations from image analysis data was performed by extrapolation of DNA content data from G1- and G 2-sorted lymphocytes with a FacStar plus flow sorter. It was demonstra ted that in MBC-treated cells the DNA rearrangement between the macron uclei in binucleates without MN was on the overage higher than in gamm a-irradiated and untreated cells, which points to aneugenic effects of MBC without the formation of MN. In contrast to DNA content measureme nts, the area of the MN is not a reliable measure For discriminating c lastogens from aneugens. (C) 1995 Wiley-Liss, Inc.