Determination of Monosomy 7 among In Vitro Generated CD1a Positive Cells in Patients with Juvenile Myelomonocytic Leukemia

To study whether the generated CD1a positive cells belong to the leukemic cells among patients with juvenile myelomonocytic leukemia. We used mononuclear cells from 3 patients with juvenile myelomonocytic leukemia, from which two had monosomy 7. The mononuclear cells from these patients were cultured in RPMI/10%FCS without adding exogeneous growth factors for 7 days. At day 7 the cultured cells were harvested and analyzed using antibodies against Ki 67, CD20 and CD1a. Additionally the cultured cells were analyzed using antibodies against CD1a and CD20 and chromosome 7 specific DNA probe, using combined fluorescence immunophenotyping and interphase cytogenetic techniques. The immunocytochemistry assay demonstrated that a high number of cells were in proliferation status, which was determined by antibody against proliferation associated nuclear protein ki 67. The percentage of Ki 67 positive cells was between 24% and 38% respectively. The percentage of CD1a positive cells was between 8% and 31% and the percentage of CD20 positive cells was between 5% and 12% respectively. The fluorescence immunophenotyping and interphase cytogenetic analysis showed that nearly all CD1a positive cells of one patient with monosomy 7 had one chromosome 7, whereas in other patient with monosomy 7, the amount of CD1a positive cells having only one chromosome 7 was approximately 11%. Furthermore, the combined immunophenotyping and cytogenetic analysis showed that the CD20 positive cells in all patients had normal karyotype. Our results suggest that CD1a positive cells generated by mononuclear cells from patients with juvenile myelomonocytic leukemia in vitro most probably belong to the leukemic cells. Since monosomy 7 could not be detected in all CD1a positive cells in juvenile myelomonocytic leukemia patients with monosomy 7, it is to assume that monosomy 7 is a secondary event in the pathophysiology of juvenile myelomonocytic leukemia.