Page 2094 - Hematology_ Basic Principles and Practice ( PDFDrive )

C H A P T E R 124

MEGAKARYOCYTE AND PLATELET STRUCTURE

Joseph E. Italiano, Jr. and John H. Hartwig

Platelets are small anucleate fragments that are formed from the expression of cyclin B. 12–15 Cyclins appear to play a critical role in
cytoplasm of megakaryocytes and have a characteristic discoid shape. directing endomitosis. Cyclin D3 is overexpressed in the G 1 phase of
16
To assemble and release platelets, megakaryocytes become polyploid maturing megakaryocytes, but a triple knockout of cyclins D1, D2,
by endomitosis and follow a maturation program that results in the and D3 in mice does not appear to affect megakaryocyte develop-
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conversion of the bulk of their cytoplasm into multiple long processes ment. In contrast, cyclin E-deficient mice exhibit a profound defect
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called proplatelets. To produce its quota of 1000–2000 platelets, a in megakaryocyte development. The molecular programming
megakaryocyte may protrude as many as 10–20 proplatelets, each of involved in endomitosis is characterized by the mislocalization or
which begins as a blunt protrusion that over time thins and branches absence of at least two critical regulators of mitosis: the chromosomal
repeatedly. Platelets form selectively at the ends of proplatelets. As passenger proteins Aurora-B/AIM-1 and survivin. AIM-1, a serine/
platelets develop, their content of granules and organelles is delivered threonine kinase in the Aurora family that is implicated in mitosis,
to them in a stream of individual particles moving from the mega- is downregulated as megakaryocyte polyploidization occurs, suggest-
karyocyte cell body to the nascent platelet buds at the proplatelet tips. ing its loss may lead to the abortive mitosis and polyploidization. 19,20
Platelet formation can be arbitrarily divided into two phases. The first However, while Aurora kinase A is required for hematopoiesis, it is
phase takes days to complete and requires megakaryocyte-specific dispensable for mouse megakarocyte endomitosis and differentia-
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growth factors. Massive nuclear proliferation to 16–32N and enlarge- tion. Whereas deletion of the APC/C cofactor Cdc20 causes mitotic
ment of the megakaryocyte cytoplasm occur as the platelet is filled arrest and severe thrombocytopenia, lack of the kinases Aurora-B,
with cytoskeletal proteins, platelet-specific granules, and sufficient Cdk1, or Cdk2 does not affect endomitosis of megakaryocytes or
membrane to complete the platelet assembly process. The second platelet counts. Deletion of Cdk1 forces a change to endocycles
phase is relatively rapid and can be completed in hours. During this without mitosis, whereas polyploidization in the absence of cdk1 and
phase, megakaryocytes generate platelets by remodeling their cyto- cdk2 occurs in the presence of aberrant replication events. Notably,
plasm first into proplatelets, then preplatelets, which undergo fission ablation of these kinases rescues defects in Cdc20 null megakaryo-
to generate discoid platelets. cytes. The observations suggest that endomitosis can be functionally
replaced by alternative polyploidization mechanisms in vivo. One
explanation for endomitosis could be inhibition of microtubule-based
MEGAKARYOCYTE DEVELOPMENT forces in anaphase B. Spindle pole separation during anaphase B is
believed to be powered by the sliding of antiparallel interdigitating
Endomitosis microtubules past each other mediated by the mitotic kinesin-like
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protein 1. This protein localizes at regions of overlapping microtu-
Hematopoietic stem cells, which are endowed with the genetic capac- bules during anaphase B and studies in vitro indicate that it can slide
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ity to differentiate into multiple lineages, are induced down the microtubules past each other. Therefore lack of spindle pole separa-
pathway to become megakaryocytes by their exposure to certain tion during endomitosis may result from failure of megakaryocytes
1
growth factors. Megakaryocytes become polyploid (i.e., 4N, 16N, to undergo normal spindle orientation and/or the absence of signals
32N, 64N) through repeated cycles of DNA replication without cell that localize or activate a kinesin motor molecule that provides force
2–5
division. Normally ploidy ranges from 4 to 64 times the haploid for sliding.
DNA complement, but the majority of cells fall within three ploidy
classes (8N, 16N, and 32N), with 16N being dominant (Fig.
6,7
124.1). Ploidy number appears to be a predetermined event, pos- Cytoplasmic Maturation
sibly signifying genetic diversity among megakaryocyte populations.
Megakaryocyte polyploidization results in a functional gene amplifi- Megakaryocytes, the largest of the hematopoietic cells, undergo a
8
cation whose likely function is an increase in protein synthesis. This pronounced cytoplasmic maturation to attain their large volumes
process, called endomitosis, is a shortened mitosis caused by a block (15,000 fL). Cytoplasmic maturation begins during endomitosis and
in late anaphase. 9,10 increases considerably after all DNA amplification has ended (Fig.
Whereas cells undergoing diploid mitoses proceed through cyto- 124.2). Megakaryocytes enlarge dramatically as they mature, reaching
kinesis and complete abscission division, megakaryocytes exhibit sizes of 100–150 µm in diameter in culture and in bone marrow.
1
regression of the cleavage furrow and reenter G 1 as polyploid cells. During this process, the megakaryocyte cytoplasm rapidly fills with
During polyploidization of megakaryocytes, the nuclear envelope platelet-specific proteins, organelles, and membrane systems that
breaks down, and an abnormal spherical mitotic spindle forms. The ultimately are subdivided and packaged into platelets (Fig. 124.3).
spindle has attached chromosomes that align from a position equi- Their cytoplasmic space expands and, except for the most cortical
distant from the spindle poles. Sister chromatids segregate and move regions, becomes densely filled with internal membranes that subse-
toward their respective poles (anaphase A). However, the spindle quently serve as the repository for the plasma membrane to be
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poles fail to move apart and do not undergo the microtubule-driven regurgitated for coating proplatelets as they extend. This internal
separation typically observed during anaphase B. Individual chroma- membrane system, which is one of the most striking features of
tids are not moved to the poles, and subsequently a single nuclear mature megakaryocytes, has been referred to as the demarcation
envelope encapsulates the entire set of sister chromatids. 9,10 In most membrane system (DMS). The DMS, first described by Yamada in
cell types, checkpoints and feedback controls ensure that DNA rep- 1957, consists of an extensive, tortuous, branching network of
lication and cell division are tightly coupled. Megakaryocytes appear membrane channels composed of flattened cisternae and tubules.
to be an exception to this rule, indicating they have managed to Initially, the DMS was proposed to play an essential role in platelet
deregulate this process. Proposed mechanisms for regulating endomi- formation by defining preformed “platelet territories” or “platelet
tosis include a reduction in mitosis-promoting factor 11,12 or decreased fields” within the megakaryocyte cytoplasm. 26,27 Release of individual

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