title: Transforming growth factor beta3 inhibits chronic myelogenous leukemia hematopoiesis by inducing Fas-independent apoptosis.
abstract: OBJECTIVE
  Transforming growth factor beta3 (TGF-beta3) is a potent suppressor of human hematopoietic progenitor cells. In this article, we compare the activity of TGF-beta3 on highly purified CD34+ cells and more immature CD34-DR(-) cells from chronic myelogenous leukemia (CML) patients in chronic phase and normal donors.
  
  
  METHODS
  Primitive hematopoietic progenitors were stimulated in liquid cultures and clonogenic assays by early-acting growth factors such as stem cell factor (SCF) and interleukin 11 (IL-11) and the intermediate-late-acting stimulating factors IL-3, granulocyte-macrophage colony-stimulating factor, and erythropoietin. Molecular analysis of bcr/abl mRNA was performed on single CML colonies by nested reverse transcriptase polymerase chain reaction. Moreover, cell cycle analysis and assessment of apoptosis of normal and leukemic CD34+ cells were performed by propidium iodide (PI) alone and simultaneous staining with annexin V and PI, respectively.
  
  
  RESULTS
  The colony-forming efficiency of CML CD34+ cells was generally inhibited by more than 90% regardless of whether the colony-stimulating factors were used alone or combined. When compared to normal CD34+ cells, leukemic cells were significantly more suppressed in 6 of 8 culture conditions. The inhibitory effect of TGF-beta3 on CD34+ cells was exerted within the first 24 hours of incubation as demonstrated by short-term preincubation followed by IL-3-and SCF-stimulated colony assays. Evaluation of bcr/abl transcript on residual CML colonies incubated with TGF-beta3 demonstrated a small subset of neoplastic CD34+ cells unresponsive to the inhibitory effect of the study cytokine. TGF-beta3 demonstrated a greater inhibitory activity on primitive CD34+DR cells than on more mature CD34+ cells. Again, CML CD34+DR(-) cells were significantly more inhibited by TGF-beta3 than their normal counterparts in 3 of 8 culture conditions. Kinetic analysis performed on CD34+ cells showed that TGF-beta induces cell cycle arrest in G(1) phase. However, this mechanism of action is shared by normal and leukemic cells. Conversely, TGF-beta3 preferentially triggered the programmed cell death of CML CD34-cells without increasing the proportion of leukemic cells coexpressing CD95 (Fas receptor), and this effect was not reversed by functional blockade of Fas receptor. Conclusion. We demonstrate that TGF-beta3 exerts a potent suppressive effect on CML cells that is partly mediated by Fas-independent apoptosis.
authors.affiliation: Institute of Hematology and Medical Oncology L. & A. Seràgnoli, University of Bologna, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Fogli
authors.forename: C
authors.initials: C
authors.lastname: Carlo-Stella
authors.forename: A
authors.initials: A
authors.lastname: Curti
authors.forename: M
authors.initials: M
authors.lastname: Ratta
authors.forename: P L
authors.initials: PL
authors.lastname: Tazzari
authors.forename: E
authors.initials: E
authors.lastname: Ragazzi
authors.forename: S
authors.initials: S
authors.lastname: Colla
authors.forename: A M
authors.initials: AM
authors.lastname: Santucci
authors.forename: S
authors.initials: S
authors.lastname: Tura
authors.forename: R M
authors.initials: RM
authors.lastname: Lemoli
mesh_terms: D018952:Antigens, CD34 / Q000032:analysis; D017209:Apoptosis / Q000187:drug effects*; D002453:Cell Cycle; D002478:Cells, Cultured; D002999:Clone Cells / Q000378:metabolism; D016207:Cytokines / Q000494:pharmacology; D006410:Hematopoiesis / Q000187:drug effects*; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000503:physiopathology*; D016212:Transforming Growth Factor beta / Q000494:pharmacology*; D015854:Up-Regulation; D019014:fas Receptor / Q000502:physiology*
chemical_list: D018952:Antigens, CD34; D016207:Cytokines; D016212:Transforming Growth Factor beta; D019014:fas Receptor