THROMBOPOIETIN, THE LIGAND FOR THE MPL RECEPTOR, SYNERGIZES WITH STEEL FACTOR AND OTHER EARLY ACTING CYTOKINES IN SUPPORTING PROLIFERATION OF PRIMITIVE HEMATOPOIETIC PROGENITORS OF MICE

Recently, the ligand for the MpI receptor (ML) was identified to be th rombopoietin, the principal regulator of megakaryocytopoiesis and thro mbopoiesis. We examined the effects of ML, as a single factor or in co mbinations with early acting factors such as steel factor (SF), interl eukin (IL)-3, IL-ll, IL-6, and granulocyte colony-stimulating factor ( G-CSF), on colony formation from primitive progenitors of mice. Cells enriched for cell cycle dormant primitive progenitors were isolated fr om bone marrow cells of li-fluorouracil (5-FU)-treated mice by a combi nation of Nycodenz density gradient separation, immunomagnetic selecti on for lineage-negative cells, and fluorescence-activated cell sorter (FAGS) sorting for Ly-6A/E(+)Kit(+) cells. ML, in the presence of eryt hropoietin, could support the formation of only a few megakaryocyte co lonies. However, ML acted synergistically with SF or IL-3 to support t he formation of multiple types of hematopoietic colonies including mul tilineage colonies. Effects of the combination of ML and SF on multipo tential progenitors were not mediated through other cells, as demonstr ated by micromanipulation of individual progenitors. In suspension cul ture, the combination of ML and SF increased the number of multipotent ial progenitors. ML also acted synergistically with IL-ll, IL-6, or G- CSF to support colony formation in serum-containing, but not in serum- free, cultures. However, the multilineage colony formation seen in ser um-containing culture was completely abrogated by addition of ACK2, a neutralizing antibody to Kit protein. Serial observation (mapping stud ies) of colony development from multipotential progenitors suggested t hat ML triggers the cell division of dormant progenitors. Based on the se observations, we propose that ML can function as an early acting cy tokine and stimulate the proliferation of cell cycle dormant progenito rs by shortening their G(0) period. (C) 1996 by The American Society o f Hematology.