Bone marrow adherent layers inhibit apoptosis of acute myeloid leukemia cells.
Human acute myeloid leukemia (AML) cells, like normal hematopoietic progenitors, die rapidly by apoptosis when cultured under serum-free conditions. Apoptosis was demonstrated by electron microscopy and agarose gel electrophoresis and quantified by flow cytometry. Culturing AML blasts in the presence of a bone marrow fibroblast (BMF) monolayer reduced the percentage of AML blasts undergoing apoptosis in the majority of cases studied. The effect was more pronounced when AML cells were cultured in the presence of an adherent long-term bone marrow (LTBM) stroma rather than BMF. Overall, the mean percentage of AML cells with fragmented DNA fell from 85 +/- 8% in control cultures to 20 +/- 9% in cultures with adherent stroma (p = 0.0004, n = 7). Supplementation of serum-free medium with recombinant cytokines, including stem cell factor, granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor (TNF)-alpha or with human placenta-conditioned medium (HPCM) matched the degree of inhibition of apoptosis induced by BMF in only 50% of cases. Granulocyte colony-stimulating factor (G-CSF), interleukin-1 beta (IL-1 beta), and IL-6 were completely ineffective. Consistent with this observation, direct contact between leukemic cells and adherent layers was essential for maximum inhibition of leukemic-cell DNA fragmentation. Separation by a porous membrane allowing passage of soluble growth factors, but interrupting direct cell contact, was associated with significantly greater DNA fragmentation and cell death. Inhibition of leukemic-cell apoptosis correlated with improved survival and growth of malignant clonogenic cells. Colonies grown in cultures were identified as leukemic by morphology and by fluorescence in in situ hybridization to demonstrate numerical chromosomal abnormalities identified at diagnosis. Close contact between leukemic cells and bone marrow inhibits blast cell apoptosis and directly promotes survival of clonogenic AML cells.
Department of Haematology, Westmead Hospital, Sydney, Australia.
L J
LJ
Bendall
A
A
Daniel
K
K
Kortlepel
D J
DJ
Gottlieb
D000208:Acute Disease; D000293:Adolescent; D000328:Adult; D000368:Aged; D017209:Apoptosis; D001854:Bone Marrow Cells; D002448:Cell Adhesion; D002999:Clone Cells; D003114:Colony-Forming Units Assay; D004249:DNA Damage; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D007223:Infant; D007951:Leukemia, Myeloid / Q000473:pathology*; D008297:Male; D008875:Middle Aged; D014407:Tumor Cells, Cultured
D006133:Growth Substances