The Wilms' tumor gene is expressed in a subset of CD34+ progenitors and downregulated early in the course of differentiation in vitro.
The Wilms' tumor gene (wt1) is strongly expressed in malignant blasts of acute myeloid leukemia (AML) in approximately 80% of all cases. However, the role of wt1 expression in non malignant hematopoietic cells remains unclear. To characterize the expression of wt1 in differentiating hematopoietic progenitors, we isolated and cultured CD34+ progenitor cells from four healthy bone marrow donors with stem cell factor (SCF) and granulocyte colony stimulating-factor (G-CSF) to induce differentiation into granulocytes. Four different cultures were carried out for 12 days. During culture, wt1 mRNA expression was analyzed by defining its ratio relative to beta-actin using reverse transcriptase polymerase chain reaction (RT-PCR). To monitor the stage of differentiation, expression of cell surface markers and peroxidase was analyzed daily. The initial purity of CD34+ cells ranged between 80% and 90%; after 12 days, the frequency of neutrophil bands and segmented neutrophils was approximately 60%. Using RT-PCR to determine the ratio of wt1 to beta-actin expression, we reproducibly detected maximum expression of wt1 mRNA at day 0 in two cultures and at day 1 in two other CD34+ cell cultures; at both these time points nearly all cells fulfilled the morphological and immunephenotypical criteria of early hematopoietic blast cells. Wt1 expression dropped rapidly at day 1 and 2, respectively, in these two pairs of cultures, and was accompanied by an increase of cells expressing CD33 surface antigen. Our data suggest that wt1 expression is restricted to a subset of CD34+ progenitors and downregulated in later stages of differentiation in vitro.
Maurer
U
U
Medical Clinic III, University Hospital, J.W. Goethe University, Frankfurt/M., Germany.
Brieger
J
J
Weidmann
E
E
Mitrou
P S
PS
Hoelzer
D
D
Bergmann
L
L
D018952:Antigens, CD34 / Q000032:analysis*; D015139:Blotting, Southern; D001854:Bone Marrow Cells; D002454:Cell Differentiation; D015536:Down-Regulation; D005786:Gene Expression Regulation; D016162:Genes, Wilms Tumor; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis; D012194:RNA-Directed DNA Polymerase; D019089:Stem Cell Factor / Q000494:pharmacology
D018952:Antigens, CD34; D012333:RNA, Messenger; D019089:Stem Cell Factor; D016179:Granulocyte Colony-Stimulating Factor; D012194:RNA-Directed DNA Polymerase