Page 1843 - Williams Hematology ( PDFDrive )

1818 Part XII: Hemostasis and Thrombosis Chapter 111: Megakaryopoiesis and Thrombopoiesis 1819

chromosomes on a metaphase plate, then the chromosomes begin to (1) a central, electron-dense nucleoid, containing fibrinogen, platelet
separate during early anaphase. However, rather than the dividing chro- factor-4, β-thromboglobulin, transforming growth factor (TGF)-β ,
1
mosomes migrating to opposite poles of the cell to allow the forma- vitronectin, and tissue plasminogen activator–like plasminogen activa-
tion of a cleavage furrow, the chromosomes quickly decondense, the tor; (2) a peripheral zone, containing tubules and von Willebrand factor
nuclear membrane reforms around the entire chromosomal comple- (arranged much like that seen in endothelial cell Weibel-Palade bodies);
ment, and the endomitotic cells reenter G phase followed by S phase. and (3) the granule membrane, containing many of the critical plate-
1
A number of attempts to understand this process at the biochemical let receptors for cell rolling (P-selectin), firm adhesion (GPIb-V-IX),
level have involved leukemic cell lines. Alterations in cyclin B, cdc2, and aggregation (integrin α β ). Proteins present in α granules arise
IIb 3
cell-cycle kinase inhibitors, and aurora kinases have been claimed to be from de novo megakaryocyte synthesis (e.g., GPIb-V-IX, GPIV, integ-
responsible for endomitosis. 43,44 Unfortunately, although these hypoth- rin α β , von Willebrand factor, P-selectin, β-thromboglobulin, plate-
IIb 3
eses possibly explain the polyploidy in various leukemic cell lines, the let-derived growth factor), nonspecific pinocytosis of environmental
hypotheses have not been substantiated in studies of normal endomi- proteins (albumin and immunoglobulin G), or cell surface membrane
totic megakaryocytes. 19,45 Endomitosis departs from a normal mitotic receptor-mediated uptake from the environment (e.g., fibrinogen,
cell cycle at the late anaphase stage, when furrow invagination aborts fibronectin, factor V). Insights into platelet granule formation have
46
short of cell abscission. Additional studies indicate that disordered come from a molecular understanding of Hermansky-Pudlak syndrome
localization of the small G-protein RhoA may be responsible for this (HPS). In this disorder, characterized by oculocutaneous albinism and a
46
property. Confirmation that a decrease in proper RhoA function is qualitative platelet bleeding disorder, a complex of at least eight proteins
critical for endomitosis comes from the, genetic elimination of RhoA form in various granule-associated complexes such as the biogenesis of
from the megakaryocytic lineage; RhoA null megakaryocytes display lysosome-related organelles complexes, which affect δ granule forma-
52
enhanced polyploidy, although the released platelets are characterized tion. These complexes are thought to be involved in cargo transport of
by abnormal membrane rheology, resulting in their rapid clearance a number of subcellular granules, such as lysosomes, melanosomes, and
from the circulation. Proper RhoA localization is controlled by its platelet δ granules.
47
activation by the RhoA guanosine triphosphate (GTP) exchange factor
(GEF) ECT2; ECT2 is down-modulated during the switch from mitosis STAGE III/IV MEGAKARYOCYTES
to endomitosis in megakaryocytes, providing a mechanistic explanation Continued cytoplasmic maturation characterizes stage III/IV megakary-
for the onset of endomitosis. 48
ocyte development (Fig. 111–4). Cells are extremely large (40 to 60 μm
in diameter) and display a low nuclear-to-cytoplasmic ratio. Cytoplas-
Cytoplasmic Development mic basophilia disappears as cells progress from stage III to stage IV.
Early in megakaryocyte development, the cytoplasm acquires a rich The demarcation membrane system gradually replaces the endoplasmic
network of microfilaments and microtubules. Toward stages III and reticulum and Golgi apparatus during the final stages of maturation.
IV, the proteins accumulate in the cell periphery, creating an organelle The nucleus usually is eccentrically placed. Although the nucleus some-
poor peripheral zone. Biochemically, the megakaryocyte cytoskeleton is times appears as several distinct nuclei in biopsy sections, it remains
composed of actin, α-actinin, filamin, nonmuscle myosin (including the highly lobulated but single at all stages of megakaryocyte development.
product of the MYH9 gene), mutated in several giant platelet thrombo- In occasional marrow sections (Fig. 111–4C), neutrophils or other
49
cytopenic syndromes (Chap. 117), β -tubulin, talin, and several other marrow cells are seen transiting through the cytoplasm of the mature
1
actin-binding proteins. Like platelets, megakaryocytes can respond to megakaryocyte, a process termed emperipolesis, and is of no pathologic
external stimuli by changing shape, transporting organelles around the significance.
cytoplasm, and secreting granules. These functions are dependent on the
microfilament and microtubule systems of the cell. In addition, micro- Proplatelet Formation
tubules play a vital role during the later stages of platelet formation. 50 Careful microscopic studies have localized marrow megakaryocytes to
the abluminal surface of sinusoidal endothelial cells. In specially pre-
Regulation of Gene Expression pared specimens, the megakaryocytes can be seen issuing long, slender
As discussed earlier, GATA-1 is vital for committing primitive multi- cytoplasmic processes between endothelial cells and into the sinusoidal
potent progenitors to the erythroid–megakaryocyte pathway. However, lumen, structures termed proplatelet processes (Fig. 111–5). The pro-
53
the transcription factor also is critical later in megakaryopoiesis, for cesses have been reproduced in vitro and in vivo. The processes consist
6
cytoplasmic development. The first convincing evidence that GATA of a β-tubulin cytoskeleton and highway, transporting organelles and
proteins affect megakaryocyte development came from overexpression platelet constituents from the megakaryocyte to the terminal projec-
studies of GATA-1 in a leukemic cell line, in which the transcription tion, the nascent platelet. 17
factor led to partial megakaryocytic differentiation. Reduction in
51
GATA-1 expression also impairs cytoplasmic development in murine Membrane Composition
megakaryocytes, reducing demarcation membranes and platelet-spe- Most of the specific characteristics of platelet membranes are present
29
cific granules. Additional transcription factors expressed during stage at stages III and IV of megakaryocyte development. Megakaryocyte
II megakaryocyte development include RUNX-1, Tal1, and Fli1, but membrane lipid composition progressively changes through develop-
these transcription factors appear to play far greater a role in megakary- ment, achieving approximately four times the content of phospholipids
ocyte maturation and platelet formation, and are discussed in “Stage III/ and cholesterol as found in immature cells. Megakaryocytes contain
IV Megakaryocytes” below. approximately the same amounts of membrane neutral and phospho-
lipid as platelets, but contain relatively more phosphatidylinositol and
Platelet Granule Formation less phosphatidylserine and arachidonic acid.
Although more prominent in later stages of differentiation (Fig. 111–3),
platelet-specific α granules first begin to form adjacent to the Golgi appa- Regulation of Gene Expression
ratus as 300- to 500-nm round or oval organelles in stage II megakary- One transcription factor that plays an important role in the final stages
ocytes. Three distinct compartments are recognized in α granules: of megakaryocyte maturation is nuclear factor-E2. Initially described as

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