Abstract
Histological examination of bone marrow biopsies shows that about one-third of chronic myeloid leukaemia (CML) patients exhibit an increase of megakaryocytes. The megakaryocytic predominance may be so striking that differentiation from other chronic myeloproliferative disorders (CMPD) may be difficult in some CML patients. Megakaryocytes in CML are clonal as demonstrated by loss of glucose-6-phosphate dehydrogenase isoenzymes. The Ph translocation, fusing the abl and bcr genes on chromosomes 9 and 22, however, obviously occurs as a second step in tumour development. So far, the Ph translocation has not been assigned explicitly to megakaryocytes. The question is whether the megakaryocytic cell lineage could harbour the bcr/abl fusion in those CML cases with striking proliferation of megakaryocytes but lack this genetic defect in cases with normal or decreased megakaryocyte counts. We therefore performed triple-colour fluorescence in situ hybridization (FISH) for portions of the bcr and abl genes flanking the breakpoint in CML in paraffin sections of CML cases with normal and with increased numbers of megakaryocytes. This method allows identification of the bcr/abl fusion in single, morphologically intact cells, whereas conventional cytogenetics requires lysis and thus destruction of the cell. Among the 21 CML patients examined by FISH, 10 were informative for bcr and abl genes and displayed distinct hybridization signals within nuclei of bone marrow cells. Besides the granulopoietic cells, megakaryocytes of all those patients (4 without and 6 with varying grades of megakaryocytic increase) displayed bcr/abl fusion signals indiciative of a Ph translocation. The lack of hybridization signals in the remaining 11 cases indicates that this technique is not of value diagnostically and should be reserved for scientific questions. Positive controls consisted of conventional chromosome preparations from bone marrow aspirates demonstrating the Ph chromosome in all patients examined, and negative controls of paraffin sections of bone marrow biopsies from non-CML patients. These showed no fusion signals in bone marrow cells, including megakaryocytes, using FISH. Our results demonstrate clearly that not only the transforming event but also the Ph translocation leading to the bcr/abl fusion happens prior to the differentiation of the pluripotent stem cell into different myeloid lineages. The megakaryocytic proliferation evident in some CML cases is probably a consequence of the disease progress.
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Buesche G, Buhr T, Georgii A (1995) Histopathology of chronic myeloid leukemia in diagnostic bone marrow biopsies. Pathologe 16:70–74
Buhr T, Choritz H, Georgii A (1992) The impact of megakaryocyte proliferation for the evolution of myelofibrosis. Virchows Arch [A] 420:473–478
Castro-Malaspina H (1984) Pathogenesis of myelofibrosis: role of ineffective megakaryopoiesis and megakaryocyte components. Prog Clin Biol Res 154:427–454
Dekmezian R, Kantarjian HM, Keating M, Talpaz M, McCredie KB, Freireich EJ (1987) The relevance of reticulin stain-measured fibrosis at diagnosis in chronic myelogenous leukemia. Cancer 59:1734–1743
Dubé ID, Kalousek DK, Coulombel L, Gupta CM, Eaves CJ, Eaves AC (1984) Cytogenetic studies of early myeloid progenitor compartments in Ph-positive chronic myeloid leukemia. II. Long-term culture reveals the persistence of Ph-negative progenitors in treated as well as newly diagnosed patients. Blood 63:1172–1177
Fialkow PJ, Jacobson RJ, Papayannopoulou T (1977) Chronic myelocytic leukemia: clonal origin in a stem cell common to the granulocyte, erythrocyte, platelet and monocyte/macrophage. Am J Med 3:125–130
Fialkow PJ, Martin PJ, Najfeld V, Penfold GK, Jacobson RJ, Hansen JA (1984) Evidence for a multistep pathogenesis of chronic myelogenous leukemia. Blood 63:1318–1323
Georgii A, Vykoupil KF, Buhr T, Choritz H, Döhler U, Kaloutsi V, Werner M (1990) Chronic myeloproliferative disorders in bone marrow biopsies. Path Res Pract 186:3–27
Georgii A, Bernhards J, Werner M (1995) The new Hannover method for plastic embedding of bone marrow biopsies. Pathologe 16:28–33
Iwamasa K, Yasukawa M, Fujita S (1989) A case of chronic myelogenous leukemia with T lymphoblastic and megakaryoblastic mixed crisis. Jpn J Med 28:89–93
Keating A (1993) Investigation of bcr/abl transcription by Phpositive chronic myeloid leukemia progenitors. Stem Cells 11 [Suppl 3]:31–33
Knuutila S, Larramendy M, Ruutu T, Paetau A, Heinonen K, Mahlamäki E (1993) Analysis of phenotype and genotype of individual cells in neoplasms. Cancer Genet Cytogenet 68:104–113
Larramendy ML, Nylund SJ, Wessman M, Ruutu T, Knuutila S (1994) Ploidy in bone marrow cells from healthy donors: a MAC (morphology antibody chromosomes) study. Br J Haematol 86:203–206
Lazzarino M, Morra E, Castello A, Inverardi D, Coci A, Pagnucco G, Magrini U, Zei G, Bernasconi C (1986) Myelofibrosis in chronic granulocytic leukaemia: clinico-pathologic correlations and prognostic significance. Br J Haematol 64:227–240
Martin PJ, Najfeld V, Hansen JA, Penfold GK, Jacobson RJ, Fialkow PJ (1995) Involvement of the B- lymphoid system in chronic myelogenous leukemia. Nature 237:49–50
McGhee JD, Hippel PH von (1977) Formaldehyde as a probe of DNA structure. 4. Mechanism of the initial reaction of formaldehyde with DNA. Biochem 16:3276–3293
Misawa M, Maeda H, Hara H, Yamamoto Y, Furuyama JI (1993) Absence of bcr/abl gene in single hemopoietic progenitors in some patients with chronic myelogenous leukemia. Stem Cells 11:536–542
Neumann MP, DeSolas I, Parkin JL, Gajl-Peczalska K, McKenna RW, Berkley CK, Nichols WL, Brunning RD (1986) Monoclonal antibody study of Philadelphia chromosome-positive blastic leukemias using the alkaline phosphatase anti-alkaline phosphatase (APAAP) technique. Am J Clin Pathol 85:564–572
Nolte M, Werner M, Ewig M, Wasielewski R von, Wilkens L, Georgii A (1995) of the Philadelphia translocation by fluorescence in situ hybridization (FISH) in paraffin sections and identification of aberrant cells by a combined FISH/immunophenotyping approach. Histopathology 26:433–437
Raskind WH, Ferraris AM, Najfeld V, Jacobson RJ, Moohr JW, Fialkow PJ (1993) Further evidence for the existence of a clonal Ph-negative stage in some cases of Ph-positive chronic myelocytic leukemia. Leukemia 7:1163–1167
Rowley JD (1973) A new consistent chromosomal abnormality in chronic myelogenous leukemia identified by quinacrine fluorescence and Giemsa staining. Nature 243:290–293
Schuh AC, Sutherland DR, Horsfall W, Mills GB, Dubé I, Baker MA, Siminovitch K, Bailey D, Keating A (1990) Chronic myeloid leukemia arising in a progenitor common to T cells and myeloid cells. Leukemia 4:631–636
Shtivelman E, Lifshitz B, Gale RP, Canaani E (1985) Fused transcript of abl and bcr genes in chronic myelogenous leukemia. Nature 315:550–554
Thiele J, Wagner S, Weuste R, Dienemann D, Wienhold S, Zankovich R, Fischer R, Stein H (1990) An immunomorphometric study on megakaryocyte precursor cells in bone marrow tissue from patients with chronic myeloid leukemia (CML). Eur J Haematol 44:63–70
Thiele J, Kvasnicka HM, Titus BR, Parpert U, Nebel R, Zankovich R, Dienemann D, Stein H, Diehl V, Fischer R (1993) Histological features of prognostic significance in CML — an immunohistochemical and morphometric study (multivariate regression analysis) on trephine biopsies of the bone marrow. Ann Hematol 66:291–302
Tkachuk DC, Westbrook CA, Andreef M, et al (1990) Detection of bcr-abl fusion in chronic myelogeneous leukemias by in situ hybridization. Science 250:559–562
Weber-Matthiesen K, Deerberg J, Müller-Hermelink K, Schlegelberger B, Grote W (1993) Rapid immunophenotypic characterization of chromosomally aberrant cells by the new FICTION method. Cytogenet Cell Genet 63:123–125
Werner M, Kaloutsi V, Buhr T, Delventhal S, Vykoupil KF, Georgii A (1991) Cytogenetics of chronic myelogenous leukemia (CML) correlated to the histopathology of bone marrow biopsies. Ann Hematol 63:201–205
Werner M, Nolte M, Ewig M, Kaloutsi V, Kausche F, Georgii A (1995) Cytogenetics and molecular genetics as complements to histopathology in the diagnosis of chronic myeloproliferative disorders. Pathologe 16:41–45
Westbrook CA, Rubin CM, Carrino JJ, LeBeau MM, Bernhards A, Rowley JD (1988) Long-range mapping of the Philadelphia chromosome by pulsed-field gel electrophoresis. Blood 71:697–702
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Nolte, M., Werner, M., Ewig, M. et al. Megakaryocytes carry the fused bcr-abl gene in chronic myeloid leukaemia: a fluorescence in situ hybridization analysis from bone marrow biopsies. Vichows Archiv A Pathol Anat 427, 561–565 (1996). https://doi.org/10.1007/BF00202886
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DOI: https://doi.org/10.1007/BF00202886