Charaterization of Human Primitive Megakaryocytes and “Platelets”.

Primitive megakaryocytes (PMs) have been first observed in murine embroynal yolk sac [

CD34+/CD31+/VE-Cadherin+/Flk-1+ hemangioblasts on day 12 were isolated and further cultured for 10–12 days in media containing thrombopoietin (TPO), interleukin 3 (IL3), stem cell factor (SCF). 30% of the non-adherent cells were strongly positive for CD31/CD41/CD61/CD42b, and weakly positive for CD42a/CD45/beta-2-microglobulin (B2M) but negative for CD117 or HLA-DR. The DNA ploidies are exclusively 2N–8N confirming the primitive status of PMs. Morphologically when compared to bone marrow derived megakaryocytes, PMs readily produce long processes with chains of “proplatelet” like structures. By live videomicroscopy, these “proplatelet” structures can actively recruit mitochondria. Interestingly, “platelet-like” fragment shedding from megakaryocytes is significantly stimulated in the presence of nitric oxide and human plasma. The “platelet-like” fragments and megakaryocytes respond to strong agonists, including thrombin and TRAP, with aggregation, alpha granule release and increased affinity for fibrinogen. In contrast, weaker agonists such as ADP only generate minimal response. This study provides preliminary evidence that primitive megakaryocytes have acquired some basic machinery of hemostasis even when the blood flow, liver function and coagulation factors are not present at the early phase of embryonal development. [We thank Dr.William L. Nichols at Mayo Clinic for his critical reviews of this abstract.]