| 主权项 |
1. A method for producing platelets for clinical transfusion in vitro comprising:
(1) culturing stem cells in a first growth medium to produce a megakaryocyte progenitor cell population, wherein the first growth medium comprises aryl-hydrocarbon inhibitor/stem regenin-1 and notch-ligand delta-1; (2) maturing the expanded megakaryocyte progenitor cells in an artificial bone marrow niche environment comprising a second growth medium in the presence of an oxygen concentration between about 10% and about 30% PO2 to differentiate the megakaryocyte progenitor cells into megakaryocytes, wherein the second growth medium comprises thrombopoietin (TPO), stem cell factor (SCF), nicotinamide, and a Rho/Rock inhibitor; (3) isolating the mature megakaryocytes; (4) culturing the mature megakaryocytes in a three-dimensional matrix and a third growth medium and in the presence of an oxygen concentration between about 10% and about 30% PO2 and a shear stress between about 100 and 400 μl/min to produce platelets, wherein the third growth medium comprises fibrinogen, fibronectin, von Willebrand factor (vWF), an MLCK inhibitor, a Rho/Rock inhibitor, and nicotinamide; and (5) collecting the platelets in a number suitable for transfusion. |
