Cytogenetic clonality analysis of megakaryocytes in myelodysplastic syndrome by dual-color fluorescence in situ hybridization and confocal laser scanning microscopy

In the myelodysplastic syndrome (MDS), cytogenetic abnormalities are often present and can be used as markers in studies for cell lineage involvement. Little is known of the involvement of the megakaryocytic lineage due to th e variable ploidy of these cells. We applied dual-color fluorescence in sit u hybridization (FISH) to routinely prepared bone marrow (BM) smears of cyt ogenetically normal patients and seven patients with MDS and monosomy 7 or trisomy 8. Probes specific for the centromeric regions of chromosomes 7 and 8 were detected with fluorescein isothiocyanate (FITC) and Texas Red, resp ectively. This enabled us to assess the ratio between the numbers of chromo somes 7 and 8 in the polyploid cells. We utilized confocal laser scanning m icroscopy to count the FITC and Texas Red FISH signals in the different foc al layers of the megakaryocytes. Fifty-six megakaryocytes in six normal BM smears were analyzed, giving a mean ratio of 1.0, a standard deviation (SD) of 0.12, and a range of 0.8-1.33. This ratio was applied to evaluation of clonal involvement of individual megakaryocytes in the patients with MDS. I n two patients with monosomy 7, the majority of the megakaryocytes were mon osomic. In the five patients with trisomy 8, all or a majority of the analy zed megakaryocytes were trisomic. These results add direct evidence that in MDS megakaryocytes are involved in the malignant clone. Genes Chromosomes Cancer 25:332-338, 1999. (C) 1999 Wiley-Liss, Inc.