The relative population sizes of megakaryocytic cells in mouse bone marrow as determined by mpl ligand responsiveness

The relative population sizes of mpl ligand-responsive megakaryocytic cells were investigated, and all megakaryocytes grown in culture were assessed. Three groups were analyzed: 1) immature cells with the ability to form single mature megakaryocytes; 2) cells with the ability to divide once before forming megakaryocytes (doublets); and 3) progenitor cells with the ability to form colonies, i.e., to undergo both cytokinesis and maturation. Immature cells forming single megakaryocytes proved most sensitive to the mpl ligand. Committed megakaryocyte progenitors required approximately 30 times more mpl ligand to achieve maximum growth than did the immature megakaryocyte population. Similar numbers of committed megakaryocyte progenitors responded to interleukin (IL)-3 and to mpl ligand. The amplification potential of these progenitor cells responding to each growth factor was assessed by measuring the number of megakaryocytes per colony. In response to mpl ligand progenitor, cells generated smaller colonies, with most cell divisions completed at a signifcantly earlier time point compared with progenitor cells responding to IL-3. The growth of more primitive megakaryocyte progenitors was best achieved in combination with other growth factors, notably IL-3; mpl ligand alone was ineffective in this regard. A novel finding was the significant number of megakaryocytes that grew in culture as closely coupled pairs (doublets). Data reported indicate that doublet formation may be a result of detection and stimulation of immature megakaryocytes rather than the diminished mpl ligand responsiveness of a proportion of megakaryocyte progenitors. By combining the number of mpl ligand-responsive cells forming single megakaryocytes with those forming megakaryocyte doublets, it is estimated that the size of the immature megakaryocyte pool greatly exceeds previous calculations. Thus we conclude that the immature megakaryocyte population is significantly larger than previously estimated and that these cells are the most sensitive to mpl ligand. Accordingly, these cells are potentially crucial in bone marrow responsiveness to mpl ligand that results from acute thrombocytopenia, being capable not only of endomitosis and maturation but perhaps of cell division as well.