Role of the tumor suppressor gene WT1 in human leukemogenesis.

为了阐明白血病细胞中WT 1基因的表达是异常的还是仅仅反映了正常对应物中的表达，对正常造血祖细胞中WT 1基因的表达水平进行了定量。根据CD 34、CD 33、CD 38、HLA-DR和c-kit的表达水平，将骨髓（BM）和脐带血（CB）细胞荧光激活细胞分选（FACS）分选为CD 34 ^+和CD 34 ^-细胞群，并将CD 34 ^+细胞分选为9个亚群（CD 34 ^+ CD 33 ^-、CD 34 ^+ CD 33 ^+、CD 34 ^+ CD 38 ^-、CD 34 ^+ CD 38 ^+、CD 34 ^+ HLA-DR^-、CD 34 ^+ HLA-DR^+、CD 34 ^+ c-kit^<hight>、CD 34 ^+c-kit-^<low>和CD 34 ^+c-kit^-）。此外，急性髓性白血病细胞也被FACS分选为四个群体（CD 34 ^+ CD 33 ^-、CD 34 ^+ CD 33 ^+、CD 34 ^-CD 33 ^+和CD 34 ^-CD 33 ^-）。流式细胞仪分选的正常造血祖细胞和白血病细胞和流式细胞仪未分选的白血病细胞的WT 1表达进行了检查，通过定量逆转录酶-聚合酶链反应。在CD 34 ^+和CD 34 ^-细胞群以及BM和CB的9个CD 34 ^+亚群中，WT 1的表达水平要么非常低（1.0至2.4 × 10 ~ 3<-2>），要么检测不到（&lt;10 ~ 3<-2>）（K562细胞的WT 1表达水平定义为1.0），而在经FACS分选和未经分选的白血病细胞中，WT 1表达的平均水平为2.4至9.3 × 10 ~ 3<-1>。因此，正常造血祖细胞中的WT 1表达水平比白血病细胞中的WT 1表达水平低至少10倍。因此，我们无法找到任何正常的BM或CB的WT 1表达水平与白血病细胞中的水平相当。这些结果表明白血病细胞中WT 1基因的异常过表达，并暗示该基因参与人类白血病的发生。

To clarify whether the expression of the WT1 gene in leukemic cells is aberrant or merely reflects that in normal counterparts, the expression levels of the WT1 gene were quantitated for normal hematopoietic progenitor cells. Bone marrow (BM) and umbilical cord blood (CB) cells were fluorescence-activated cell sorting (FACS)-sorted into CD34^+ and CD34^- cell populations, and the CD34^+ cells into nine subsets (CD34^+CD33^-, CD34^+CD33^+.CD34^+CD38^-, CD34^+CD38^+, CD34^+HLA-DR^-, CD34^+HLA-DR^+, CD34^+c-kit^<hight>, CD34^+c-kit-^<low>, and CD34^+c-kit^-) according to the expression levels of CD34, CD33, CD38, HLA-DR,and c-kit. Moreover, acute myeloid leukemic cells were also FACS-sorted into four populations (CD34^+CD33^-, CD34^+CD33^+, CD34^-CD33^+, and CD34^-CD33^-). FACS-sorted normal hematopoietic progenitor and leukemic cells and FACS-unsorted leukemic cells were examined for the WT1 expression by quantitative reverse transcriptase-polymerase chain reaction. The WT1 expression in the CD34^+ and CD34^- cell populations and in the nine CD34^+ subsets of BM and CB was at either very low (1.0 to 2.4 x 10^<-2>) or undetectable (<10^<-2>) levels (the WT1 expression level of K562 cells was defined as 1.0), whereas the average levels of WT1 expression in FACS-sorted and-unsorted leukemic cells were 2.4 to 9.3 x 10^<-1>. Thus, the WT1 expression levels in normal hematopoietic progenitor cells were at least 10 times less than those in leukemic cells. Therefore, we could not find any normal counterparts of BM or CB that expressed the WT1 at levels comparable with those in leukemic cells. These results indicate an aberrant overexpression of the WT1 gene in leukemic cells and imply the involvement of this gene in human leukemogenesis.