Control of normal differentiation of myeloid leukemic cells. XII. Isolation of normal myeloid colony‐forming cells from bone marrow and the sequence of differentiation to mature granulocytes in normal and D+ myeloid leukemic cells

Abstract An enriched population of early myeloid cells has been obtained from normal mouse bone marrow by injection of mice with sodium caseinate and the removal of cells with C3 (EAC) rosettes by Ficoll‐Hypaque density centrifugation. This enriched population had no EAC or Fc (EA) rosettes and contained 87% early myeloid cells stained for myeloperoxidase and/or AS‐D‐chloroacetate esterase, 7% cells in later stages (ring forms) of myeloid differentiation and 6% unstained cells, 2% of which were small lymphocytes. After seeding in agar with the macrophage and granulocyte inducer MGI, the enriched population showed a cloning efficiency of 14% when removed from the animal and of 24% after one day in mass culture. Both the enriched and the unfractionated bone marrow cells gave the same proportion of macrophage and granulocyte colonies. The normal early myeloid cells were induced to differentiate by MGI in mass culture in liquid medium to mature granulocytes and macrophages. The sequence of granulocyte differentiation was the formation of EA and EAC rosettes followed by the synthesis and secretion of lysozyme and morphological differentiation to mature cells. D + myeloid leukemic cells with no EA or EAC rosettes had a similar morphology to normal early myeloid cells and showed the same sequence of differentiation. The induction of EA and EAC rosettes occurred at the same time in both the normal and D + leukemic cells, but lysozyme synthesis and the formation of mature granulocytes was induced later in the leukemic than in the normal cells. The results indicate that selection for non‐rosette‐forming normal early myeloid cells also selected for myeloid colony forming cells, that these normal early myeloid cells can form colonies with differentiation to macrophages and granulocytes, that normal and D + myeloid leukemic cells have a similar sequence of differentiation and that the normal cells had a greater sensitivity for the formation of mature cells by MGI.