PLATELETS AND THROMBOPOIESIS Notch/Delta4 signaling inhibits human megakaryocytic terminal differentiation

The effects of Notch signaling on human megakaryocytic and erythroid differentiation were investigated by exposing human CD34 progenitor cells to an immobilized chimeric form of the Notch ligand, Delta-like4 (Dll4Fc). Exposure of human cord blood CD34 cells to Dll4Fc induced a modest enhancement of erythroid cell production. Conversely, under megakaryocytic culture conditions, Dll4Fc strongly impaired platelet production by reducing the generation of mature CD41a CD42b megakaryocytes (MKs) and platelet-forming cells. The inhibitory activity of Dll4 on terminal MK differentiation was confirmed by culturing CD34 cells onto Dll-4–expressing stroma cells (engineered to express the membraneanchored form of Dll4). The reduced production of mature CD41a CD42 cells was rescued by inhibiting Notch signaling either with the N-N-(3,5-difluorophenacetylL-alanyl)-S-phenylglycine t-butyl ester -secretase inhibitor or the dominantnegative version of Mastermind. Dll4 impaired the generation of mature CD41a CD42b cells and proplatelet formation without affecting earlier steps of MK differentiation, such as production of megakaryocytic/erythroid progenitors and colony-forming units–MKs. This blockade was accompanied by a modulation of the transcriptional program of megakaryocytic differentiation. All these results indicate that Dll4/Notch signaling inhibits human terminal MK differentiation. (Blood. 2010;116(25):5670-5678)