In order to investigate the biologic processes underlying and resulting from the megakaryocytic hyperplasia that characterizes idiopathic myelofibrosis (IMF), peripheral blood CD34⁺ cells isolated from patients with IMF, polycythemia vera (PV), and G-CSF–mobilized healthy volunteers were cultured in the presence of stem cell factor and thrombopoietin. IMF CD34⁺ cells generated 24-fold greater numbers of megakaryocytes (MKs) than normal CD34⁺ cells. IMF MKs were also shown to have a delayed pattern of apoptosis and to overexpress the antiapoptotic protein bcl-xL. MK hyperplasia in IMF is, therefore, likely a consequence of both the increased ability of IMF progenitor cells to generate MKs and a decreased rate of MK apoptosis. Media conditioned (CM) by CD61⁺ cells generated in vitro from CD34⁺ cells were then assayed for the levels of growth factors and proteases. Higher levels of transforming growth factor-β (TGF-β) and active matrix metalloproteinase-9 (MMP9) were observed in media conditioned with IMF CD61⁺ cells than normal or PV CD61⁺ cells. Both normal and IMF CD61⁺ cells produced similar levels of VEGF. MK-derived TGF-B and MMP-9, therefore, likely contribute to the development of many pathological epiphenomena associated with IMF.