Jl. Liesveld et al., STEM-CELL FACTOR AND STROMAL CELL COCULTURE PREVENT APOPTOSIS IN A SUBCULTURE OF THE MEGAKARYOBLASTIC CELL-LINE, UT-7, Leukemia research, 20(7), 1996, pp. 591-600
The megakaryoblastic cell line, UT-7, is dependent for its growth upon
interleukin-3 (IL-3), erythropoietin, or granulocyte-macrophage colon
y stimulating factor (GM-CSF). A subculture of this line can be mainta
ined in recombinant human c-kit ligand [stem cell factor (SCF)] at 100
ng/ml without requirement for other growth factors. Removal of this s
ubculture from SCF results in rapid loss of viability and decreased pr
oliferation. Cells grown in SCF also can be maintained in GM-CSF but n
ot vice versa. In this work, we have characterized the SCF dependence
of this UT-7 subculture. Stem cell factor removal results in apoptosis
and a decline in viability which can be restored partially by re-addi
tion of SCF, GM-CSF, or co-culture with adherent marrow stromal cells.
Apoptosis in the factor-starved UT-7 population has been documented b
y light microscopy, electron microscopy and DNA analysis, showing the
typical 180 base pair laddering characteristic of apoptosis. To quanti
tate the degree of apoptosis in the cell populations, and to assess wh
ether apoptosis decreased with re-exposure of starved cells to growth
factors or stroma, we utilized flow cytometry. This confirmed that exp
osure of previously factor-starved cells to stroma decreased the perce
ntage of cells undergoing apoptosis. Co-culture with an SCF-deficient
murine stromal cell line was also able to prevent apoptosis, suggestin
g contribution of other stromal cell factors. Experiments performed us
ing trans-well inserts which do not allow cell passage, showed greates
t viability of cells in contact with stroma, but viability was also im
proved in cells cultured in the presence of, but not in contact with,
stromal cells compared to those cultured above plastic, suggesting a r
ole for soluble stroma-produced substances. These data demonstrate tha
t SCF alone can prevent apoptosis in cells dependent upon its presence
for proliferation. Also, marrow stromal cells can serve as a partial
substitute for growth factor in the prevention of apoptosis in these c
ells, probably due to constitutive presentation of SCF and other hemat
opoietic growth factors in both soluble and surface-bound forms. Copyr
ight (C) 1996 Elsevier Science Ltd.