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C H A P T E R 28

THROMBOCYTOPOIESIS

Alan B. Cantor

Platelets, once regarded simply as “blood dust,” are now recognized tion to total thrombocytopoiesis is estimated to account for at most
to play essential roles in hemostasis. Not only do they form a 7% to 15%.
hemostatic plug and initiate thrombus formation in the event of
vascular injury, but they also repair minute vascular damage that
occurs on a daily basis. Platelets also participate in wound healing Megakaryocyte Progenitors
and angiogenesis via delivery of key growth factors to sites of vas-
cular injury and interact with the innate immune system. Disorders Like other hematopoietic progenitor cells, MkPs can be cultured in
associated with platelet production carry significant morbidity and vitro using semisolid media. Animal studies using these colony assays
mortality in humans because of hemorrhage, thrombosis, bone have allowed delineation of hierarchal developmental pathways of
marrow (BM) fibrosis, BM failure, and/or hematologic malignancy. MkP maturation based on proliferative potential, DNA content,
Platelets are generated from their precursor cells, megakaryocytes, morphologic criteria, and gene expression pattern (Fig. 28.1). This
via a complex process. For a long time, the extreme rarity of pathway can be conceptually divided into three broad stages: prolifer-
megakaryocytes significantly hampered studies aimed at under- ating MkPs, which contain normal DNA content (2N/4N), nonpro-
standing the molecular mechanisms underlying platelet biogenesis. liferating immature megakaryocytes (4N to 8N DNA content), and
However, the purification and cloning (in 1994) of thrombopoietin nonproliferating mature megakaryocytes (DNA content 8N to128N).
(TPO), the major megakaryocyte cytokine, has enabled consider- Within the proliferating MkP compartment, the earliest detectable
able progress to be made. Recent application of whole exome and cell is the megakaryocyte high-proliferative-potential colony-forming
genome DNA sequencing has further stimulated discovery of new cell (Mk-HPP-CFC), which is capable of generating macroscopically
disease-associated genes involved in human thrombocytopoiesis. visible colonies containing a few thousand megakaryocytes. This
These new insights provide an important foundation for improved corresponds to a proliferative capacity of ≈8 to 10 replicative cycles.
diagnosis and treatment of disorders of thrombocytopoiesis. This These cells require IL-3 and simultaneous activation of the protein
chapter reviews the current understanding of megakaryocyte biology kinase C and cyclic adenosine monophosphate signaling pathway.
and platelet production, highlighting connections with human The burst-forming unit-megakaryocyte (BFU-Mk), which is
disease. thought to be a direct progeny of Mk-HPP-CFC, is more mature than
the Mk-HPP-CFC, but retains a high degree of proliferative potential,
developing “bursts” of individual colony-forming cells. These colonies
MEGAKARYOCYTE BIOLOGY contain approximately 100 to 500 megakaryocytes, representing ≈5
to 7 replicative cycles. In humans, BFU-Mk cells require mitogenic
Megakaryocyte Development stimulation with IL-3 or granulocyte-macrophage colony-stimulating
factor (GM-CSF) and synergistic signaling with stem cell factor (SCF;
Although platelets were described as early as the 1840s, it was not also called kit-ligand), interleukin (IL)-11, IL-1α, and TPO. They are
until 1906, in a seminal study by James Homer Wright, that their also resistant to treatment in vitro with 5-fluorouracil.
1
origin from megakaryocytes was first recognized. Megakaryocytes The most mature proliferating cell is the colony-forming cell-
are large polyploid cells that reside predominantly within the BM megakaryocyte (CFU-Mk), which has very limited proliferative
during postnatal life. They are rare cells, constituting only about potential, representing only 2 to 5 cell divisions (4 to 32 megakaryo-
0.1% of nucleated cells under normal steady-state conditions. They cytes per colony). This progenitor responds to a variety of single
develop from common bipotential megakaryocyte-erythroid pro- growth factors, such as IL-3 and GM-CSF, and coregulators such as
genitor (MEP) cells, which are themselves derived from common SCF, FMS-like tyrosine kinase-3 ligand, and TPO. They express early
myeloid progenitor (CMP) cells, and ultimately from totipotent markers of differentiation such as glycoprotein IIb (GPIIb) and
hematopoietic stem cells (HSCs). Recent data suggest that mega- platelet factor 4 (PF4) before initiating endomitotic cell cycles.
karyocyte progenitor (MkP) cells can also develop in a more direct
hierarchical fashion from HSCs. Once committed to the mega-
karyocytic lineage, MkPs undergo a series of dramatic maturational Immature Megakaryocytes: Promegakaryoblasts
steps ultimately tailored to their final task of platelet production
and release. These include changes in proliferative capacity, cell size, Promegakaryoblasts are transitional cells intermediate between pro-
nuclear content, organelle biogenesis, membrane development, and liferating progenitor cells and postmitotic, mature megakaryocytes.
cytoskeletal rearrangement. The large increase in cell size is linked These cells are not readily observed morphologically in vitro or in
to an unusual process termed endomitosis, in which cells replicate BM specimens but may be identified by their expression of
their DNA but fail to undergo cytokinesis. Mature megakaryocytes megakaryocyte-specific or platelet-specific markers, such as platelet
reach diameters of approximately 100 µm and contain DNA peroxidase, platelet GPIIb/IIIa, and von Willebrand factor (vWF).
content as high as 128N. They contain a multilobulated nucleus They have DNA content levels intermediate between proliferating
enclosed by a single nuclear membrane. Their abundant cytoplasm progenitors and mature megakaryocytes. Promegakaryoblasts respond
is filled with ribosomes, platelet-specific granules, mitochondria, to a variety of cytokines in vitro, including IL-3, SCF, IL-6, and TPO,
and complex intracellular membrane systems. Although megakaryo- to produce large polyploid megakaryocytes. At least three distinct
cytes reside predominantly within the BM, they are also found in subpopulations of promegakaryoblasts have been identified based on
peripheral blood, spleen, and lung under normal conditions. These different physiochemical characteristics, morphology, antigen expres-
extramedullary megakaryocytes release platelets, but their contribu- sion, and enzyme content.

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