THROMBOPOIETIN STIMULATES COLONY-FORMING UNIT MEGAKARYOCYTE PROLIFERATION AND MEGAKARYOCYTE MATURATION INDEPENDENTLY OF CYTOKINES THAT SIGNAL THROUGH THE GP130 RECEPTOR SUBUNIT

Thrombopoietin (Tpo), the ligand for the c-Mpl receptor, is a major re gulator of megakaryopoiesis. Treatment of mice with Tpo raises the pla telet count fourfold within a few days. Conversely, c-mpl knock-out mi ce have platelet counts that are 15% that of normal. The subunit struc ture of the c-Mpl receptor is not fully understood. Some cytokines tha t stimulate megakaryopoiesis (IL-6, IL-11, leukemia inhibitory factor, and oncostatin M) bind to receptors that use gp130 as a signal transd uction subunit. For these reasons, we determined whether gp130 functio n was required for Tpo-induced signal transduction, Murine marrow cell s were cultured in semi-solid media in the presence of Tpo or IL-3, wi th or without a neutralizing anti-gp130 monoclonal antibody (RX187) or a soluble form of c-Mpl receptor (soluble Mpl) that blocks Tpo bioact ivity, and the numbers of colony-forming unit-megakaryocyte (CFU-Meg) colonies were counted on day 5. Murine marrow cells were also cultured in suspension under serum-free conditions for 5 days, and megakaryocy te DNA content was measured by flow cytometry, as an index of nuclear maturation, The addition of RX187 did not block Tpo-induced CFU-Meg co lony growth nor CFU-Meg nuclear maturation in suspension culture, Howe ver, IL-3-induced CFU-Meg colony growth and megakaryocyte nuclear matu ration decreased in the presence of RX187, Soluble Mpl completely abla ted Tpo-induced CFU-Meg growth, and partially blocked IL-3-stimulated CFU-Meg growth. Thus the effects of Tpo on megakaryopoiesis in vitro d o not depend on cytokines that signal through gp130. Furthermore, it i s unlikely that gp130 serves as a beta chain for the c-Mpl receptor, a s Tpo signalling is unimpaired in the presence of RX187. In contrast, the effects of IL-3 on CFU-Meg growth are mediated in part through Tpo and through gp130-signalling cytokines. (C) 1996 by The American Soci ety of Hematology.