Kinetics of Appearance of Differentiation-associated Characteristics in ML-1, a Line of Human Myeloblastic Leukemia Cells, after Treatment with 12-O-Tetradecanoylphorbol-13-acetate, Dimethyl Sulfoxide, or 1-β-d-Arabinofuranosylcytosine

Abstract

The process of drug-induced cell differentiation was studied by comparing the effects 12-O-tetradecanoylphorbol-13-acetate (TPA), dimethyl sulfoxide (DMSO), and 1-β-d-arabinofuranosylcytosine (ara-C) exert on the in vitro growth and viability of a human myeloblastic leukemia cell line, ML-1, and by monitoring the kinetics with which these agents induce the appearance of various differentiation-associated characteristics.

An increase in Fc surface receptors became detectable within 6 hr after exposure of the cells of 5 × 10^-10 m TPA or 1.6% DMSO, and this marker was found present in 25 and 40%, respectively, of the viable cells incubated for 24 hr in the presence of these agents. This level remained essentially constant during the additional 6-day incubation period. After treatment with 5 × 10^-8 or 1 × 10^-7 m ara-C, approximately 2 days were required for significant levels of Fc receptors to be expressed. The number of cells demonstrating this marker increased steadily during further incubation, amounting, on Day 7, to approximately 60 and 70%, respectively, of the viable cell population.

The ability of the cells to reduce nitroblue tetrazolium dye and to engulf latex particles also appeared within one day after their exposure to TPA but was delayed for an additional 24 or 48 hr after treatment with ara-C or DMSO, respectively. Once initiated, these marker activities increased as a function of time. On Day 5, approximately 50% of the viable cells exposed to TPA demonstrated nitroblue tetrazolium dye-reducing ability, whereas the number of viable cells displaying this marker after treatment with DMSO or ara-C for seven days amounted to 20 and 35 to 40%, respectively. The capacity to engulf latex particles was observed in approximately 50% of the viable cells treated for seven days with TPA and in approximately 30 and 40%, respectively, of the cells exposed to DMSO or to ara-C.

The rates at which these functional markers appeared was paralleled by the rates at which morphological maturation of the treated cells occurred, lineage specificity being monitored by α-naphthyl acetate esterase and AS-D chloroacetate esterase activity. TPA and ara-C gave rise to cells of the monocytemacrophage complex; DMSO stimulated the appearance of granulocytic intermediates. On Day 7, after exposure to TPA, approximately 95% of the viable cells were present at various stages of differentiation. After treatment with 5 × 10^-8 or 1 × 10^-7 m ara-C, differentiation to various stages occurred in approximately 60 and 80%, respectively, of the viable cell fraction. This fraction decreased with increasing drug concentrations. DMSO stimulated differentiation in approximately 45% of the viable cells.

These data show that the same differentiation-associated characteristics were caused to be expressed by the three chemical agents at different rates and to different extents, reflecting the differences in their ability to induce morphological maturation. They also suggest that an optimal differentiation-inducing dose can be defined at which the agents cause maximal differentiation at minimal cell kill. The implications of these findings for chemotherapy are discussed.