c-kit Ligand가 Transforming Growth-Factor-β1(TGF-β1)에 의한 CD34^(+) 급성 골수성백혈병세포의 Apoptosis에 미치는 영향

Abstract

Background: Transforming growth factor-β1(TGF-β1) induces a inhibition of blast clonogenic cells in suspension and methylcellulose cultures. But the mechanisms of inhibition of leukemic clonogenic cells are not known exactly. We evaluated quantitatively the TGF-β1-induced apoptosis of CD34-positive(CD34+) acute myeloid leukemic(AML) cells because TGF-β1 is one of the representative cytokines that induce cell death by apoptotic mechanism. And we investigated the role of c-kit ligand(stem cell factor; SCF) with or without granulocyte-macrophage colony-stimulating factor(GM-CSF) and/or interleukin-3(IL-3) in the inhibition of TGF-β1-induced apoptosis of CD34+ AML cells. Materials and Methods: CD34+ AML cells were isolated from the bone marrow of 14 patients with AML using immunomagnectic microbead method. The various combination of cytokines including TGF-β1, SCF, GM-CSF, and IL-β1, was used to study the quantitation of colony formation in methylcellulose and of apoptosis of CD34+ AML cells in vitro. Analysis of apoptosis was done by ELISA method using Cell Death Detection kit. Results: SCF alone resulted in no significant colony formation, while the combination of SCF plus GM-CSF and/or IL-β1 stimulated significant colony formation. The addition of TGF-β1 alone to culture of CD34+ AML cells resulted in total inhibition of colony formation. The combination of SCF plus TGF-β1 also did not result in colony formation. But no significant inhibition of colony formation was obtained for the combination of SCF plus GM-CSF by TGF-β1. Our study demonstrated that TGF-β1 induces apoptosis in CD34+ AML cells. This TGF-β1-induced apoptosis was not inhibited by SCF alone. The combination of SCF and IL-3 showed a marginal effect. But the TGF-β1-induced apoptosis of CD34+ AML cells was significantly inhibited by SCF plus GM-CSF (P<0.05) or SCF and GM-CSF plus IL-3(p<0.05). This pattern of inhibition of TGF-β1-induced apoptosis by costimulating effect of SCF was also observed for the CD34+ normal bone marrow cells, but not for CD34- AML cells. Conclusion: This study showed that costimulatory effect of c-kit ligand, with GM-CSF or GM-CSF plus IL-3, can inhibit the TGF-β1-induced apoptosis preferentially in CD34+ AML cells rather than the CD34- AML cells. And we suggest that the TGF-β1-induced growth inhibition of clonogenic leukemic cells may be associated with the TGF-β1-induced apoptosis of clonogenic leukemic cells.