Platelets are formed and released into the bloodstream by precursor cells called megakaryocytes that reside within the bone marrow. The production of platelets by megakaryocytes requires an intricate series of remodeling events that result in the release of thousands of platelets from a single megakaryocyte. Abnormalities in this process can result in clinically significant disorders. Thrombocytopenia (platelet counts less than 150,000/µl) can lead to inadequate clot formation and increased risk of bleeding, while thrombocythemia (platelet counts greater than 600,000/µl) can heighten the risk for thrombotic events, including stroke, peripheral ischemia, and myocardial infarction. This Review will describe the process of platelet assembly in detail and discuss several disorders that affect platelet production. Platelet formationMegakaryocyte development. Megakaryocytes are rare myeloid cells (constituting less than 1% of these cells) that reside primarily in the bone marrow (1) but are also found in the lung and peripheral blood. In early development, before the marrow cavities have enlarged sufficiently to support blood cell development, megakaryopoiesis occurs within the fetal liver and yolk sac. Megakaryocytes arise from pluripotent HSCs that develop into 2 types of precursors, burst-forming cells and colony-forming cells, both of which express the CD34 antigen (2). Development of both cell types continues along an increasingly restricted lineage culminating in the formation of megakaryocyte precursors that develop into megakaryocytes (1). Thrombopoietin (TPO), the primary regulator of thrombopoiesis, is currently the only known cytokine required for megakaryocytes to maintain a constant platelet mass (3). TPO is thought to act in conjunction with other factors, including IL-3, IL-6, and IL-11, although these cytokines are not essential for megakaryocyte maturation (4).Megakaryocytes tailor their cytoplasm and membrane systems for platelet biogenesis. Before a megakaryocyte has the capacity to release platelets, it enlarges considerably to an approximate diameter of 100 µm and fills with high concentrations of ribosomes that facilitate the production of platelet-specific proteins (5). Cellular enlargement is mediated by multiple rounds of endomitosis, a process that amplifies the DNA by as much as 64-fold (6-9). TPO, which binds to the c-Mpl receptor, promotes megakaryocyte endomitosis. During endomitosis, chromosomes replicate and the nuclear envelope breaks down. Although interconnected mitotic spindles assemble, the normal mitotic cycle is arrested during anaphase B. The spindles fail to separate, and both telophase and cytokinesis are bypassed. Nuclear envelope reformation (10, 11) results in a polyploid, multilobed nucleus with DNA contents ranging from 4N up to 128N within each megakaryocyte (12).In addition to expansion of DNA, megakaryocytes experience significant maturation as internal membrane systems, granules, and organelles are assembled in bulk during their development. In particular, there is th...

Megakaryocytes are terminally differentiated cells that, in their final hours, convert their cytoplasm into long, branched proplatelets, which remodel into blood platelets. Proplatelets elongate at an average rate of 0.85 m/min in a microtubule-dependent process. Addition of rhodamine-tubulin to permeabilized proplatelets, immunofluorescence microscopy of the microtubule plusend marker end-binding protein 3 (EB3), and fluorescence time-lapse microscopy of EB3-green fluorescent protein (GFP)-expressing megakaryocytes reveal that microtubules, organized as bipolar arrays, continuously polymerize throughout the proplatelet. In immature megakaryocytes lacking proplatelets, microtubule plus-ends initiate and grow by centrosomal nucleation at rates of 8.9 to 12.3 m/min. In contrast, plus-end growth rates of microtubules within proplatelets are highly variable (1.5-23.5 m/ min) and are both slower and faster than those seen in immature cells. Despite the continuous assembly of microtubules, proplatelets continue to elongate when net microtubule assembly is arrested. One alternative mechanism for force generation is microtubule sliding. Triton X-100-permeabilized proplatelets containing dynein and its regulatory complex, dynactin, but not kinesin, elongate with the addition of adenosine triphosphate (ATP) at a rate of 0.65 m/min. Retroviral expression in megakaryocytes of dynamitin (p50), which disrupts dynactindynein function, inhibits proplatelet elongation. We conclude that while continuous polymerization of microtubules is necessary to support the enlarging proplatelet mass, the sliding of overlapping microtubules is a IntroductionBlood platelets, tiny cells shed by megakaryocytes, circulate throughout blood vessels and survey the integrity of the vascular system. In response to traumatic injuries in which blood vessel continuity is interrupted, platelets bind to exposed collagen, change shape, secrete granule contents, and aggregate with neutrophils to form a hemostatic plug to seal off the damaged blood vessel. The mechanisms by which blood platelets are formed and released from giant precursor cells, called megakaryocytes, in situ remain to be defined. However, the development of megakaryocyte culture systems that produce platelets has provided a means to study the intermediate structures called "proplatelets," long (up to several millimeters), thin extensions of the megakaryocyte cytoplasm that contain multiple platelet-sized beads along their length. [1][2][3][4][5][6][7] Based on multiple lines of evidence, we have speculated that platelets are not preassembled in the megakaryocyte cytoplasm but instead are constructed de novo, predominantly at the ends of the proplatelets. 8 As predicted, megakaryocytes go to great lengths to amplify the number of proplatelet ends, taking the shaft of each proplatelet and bending it multiple times. Each bend yields a bifurcation in the shaft, generating a new end.The formation of proplatelets is highly dependent upon a complex network of protein filaments that extends throug...