Analysis of commitment of human leukemia HL-60 cells to terminal granulocytic maturation

Analysis of commitment of human promyelocytic leukemia HL-60 cells to terminal granulocytic maturation induced by dimethyl sulfoxide (DMSO) or retinoic acid (RA) was accomplished using biochemical measurements and a plasma clot clonal assay system that permits a high plating efficiency of 40 to 60%. Commitment to granulocytic maturation occurs very rapidly. When cells are exposed to these inducers for only 8 to 18 h, an interval much shorter than a single generation time, and are then subcultured in inducer-free plasma clots, they demonstrate a decrease in proliferative capacity and form colonies composed of mature nitroblue tetrazolium (NBT)-positive cells along with occasional colonies containing both NBT-positive and NBT-negative cells; in both types of colony, the NBT-positive cells are widely dispersed. Undifferentiated HL-60 cells give rise to compact NBT-negative colonies of large size without cell migration. HL-60 cell differentiation induced by either DMSO or RA is associated with a progressive decline in both DNA and RNA synthesis; this includes transcriptional inactivation of ribosomal DNA sequences. In contrast to DMSO, which induces development primarily of metamyelocytes, RA treatment leads to the accumulation of more mature band and segmented neutrophils; sequential exposure of cells pretreated with DMSO to RA alone fails to cause rapid appearance of segmented neutrophils. From these studies, we conclude that HL-60 cells become very rapidly committed to terminal maturation and that DMSO and RA appear to induce granulocytic maturation via two different mechanisms.