Abstract
To preliminarily identify the existence of CD34- leukemia stem cell (LSC) in t(8;21) acute myeloid leukemia (AML) by in vitro test. Bone marrow samples collected from newly diagnosed t(8;21) AML patients were tested. Lin-CD34+ CD38-(abbreviation, CD34+CD38-), Lin-CD34+CD38+ (abbreviation, CD34+CD38+) and Lin-CD34-CD38-CD45dimSSClow(abbreviation, CD34-"LSC") cell fractions were gated by flow cytometry after staining with fluorescent antibodies. Cells in G0 phase were identified through Hoechst 33342 and pyronin Y staining. Aldefluor reagent was used to test aldehyde dehydrogenase (ALDH) activity. The above-mentioned 3 cell fractions were sorted, and mRNA levels of AML1-ETO and WT1 were measured by real-time quantitative PCR. The 3 tested samples displayed the same tendency in ratio of the cells in G0 phase: CD34-"LSC">CD34+ CD38->CD34+CD38+. The paired t-test of 53 patients showed that frequency of ALDHbr cells of both CD34+CD38- and CD34-"LSC" cell fractions was significantly higher than that of CD34+CD38+ (P<0.01), furthermore, the ALDHbr cell frequency was significantly higher in CD34-"LSC" than that in CD34+ CD38- (P<0.01). AML1-ETO mRNA levels of cells sorted from 3 patients were similar among the 3 cell fractions within each patient, whereas WT1 mRNA levels were significantly higher in CD34-"LSC" than that in other 2 cell fractions. CD34- LSC may exist in t(8;21) AML, and may be more primitive than CD34+ LSC. These results promote the necessity to perform in vivo xenogeneic transplantation mice.
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