Abstract
Apoptosis is a recognized limitation to generating large numbers of megakaryocytes in culture. The genes responsible have been rigorously studied
in mice, but are poorly characterized in human culture systems. As CD34-positive (
) cells isolated from human umbilical vein cord blood were differentiated into megakaryocytes in culture, two distinct cell populations were identified by flow cytometric forward and side scatter: larger size, lower granularity (LLG), and smaller size, higher granularity (SHG). The LLG cells were CD41a
CD42a
phosphatidylserine
, had an electron microscopic morphology similar to mature bone marrow megakaryocytes, developed proplatelets, and displayed a signaling response to platelet agonists. The SHG cells were CD41a
CD42a
phosphatidylserine
, had a distinctly apoptotic morphology, were unable to develop proplatelets, and showed no signaling response. Screens of differentiating megakaryocytes for expression of 24 apoptosis genes identified
as a novel candidate megakaryocyte apoptosis regulator. Lentiviral
overexpression decreased megakaryocyte apoptosis, increased CD41a
LLG cells, and increased proplatelet formation by 58%. An association study in 154 healthy donors identified a significant positive correlation between platelet number and platelet
mRNA levels. This finding was consistent with the observed increase in platelet-like particles derived from cultured megakaryocytes over-expressing
also induced small, but significant increases in thrombin-induced platelet-like particle αIIbβ3 activation and P-selectin expression. Thus,
restrains apoptosis in cultured megakaryocytes, promotes proplatelet formation, and is associated with platelet number.
is a novel target for improving megakaryocyte and platelet yields in
culture systems.