Platelet production

• Platelets


  Nonnucleated blood cells that circulate at a concentration of 150 to 400 × 10%/L, with average platelet counts slightly higher in women than in men and slightly lower in members of both sexes who are older than 65 years
• Platelets


  Trigger primary hemostasis on exposure to subendothelial collagen or endothelial cell inflammatory proteins at the time of blood vessel injury
• Megakaryocytes


  Unique bone marrow cells that are the largest cells in the bone marrow and possess multiple chromosome copies (polyploid)
• Megakaryocytes


  • On a Wright-stained bone marrow aspirate smear, each megakaryocyte is 30 to 50 μm in diameter with a multilobulated nucleus and abundant granular cytoplasm
  • Megakaryocytes account for less than 0.5% of all bone marrow cells, and on a normal Wright-stained bone marrow aspirate smear two to four megakaryocytes per 10x low-power field may be identified
• Megakaryocyte differentiation


  1. Megakaryocyte progenitors are recruited from common myeloid progenitors and subsequently differentiate through several maturation stages

  2. They extend proplatelet processes, projections that resemble strings of beads, through or between the endothelial cells and into the venous sinuses, releasing platelets from the tips of the processes into the circulation
• Megakaryocyte progenitors


  Arise from the common myeloid progenitor under the influence of the transcription gene product, GATA-1, regulated by cofactor FOG1
• Megakaryocyte differentiation


  Is suppressed by the transcription gene product, MYB, so GATA-1 and MYB act in opposition to balance megakaryocytopoiesis with erythropoiesis
• Megakaryocyte lineage-committed progenitor stages

  • Burst-forming unit (BFU-Meg)
  • Colony-forming unit (CFU-Meg)
  • Light-density CFU (LD-CFU-Meg)
• BFU-Meg and CFU-Meg


  • Diploid and undergo normal mitosis to maintain a viable pool of megakaryocyte progenitors
  • Their proliferative properties are reflected in their ability to form hundreds (BFU-Megs) or dozens (CFU-Megs) of colonies in culture
• LD-CFU-Meg


  Undergoes endomitosis, a partially characterized form of mitosis unique to megakaryocytes in which DNA replication and cytoplasmic maturation are normal but cells lose their capacity to divide
• Endomitosis


  1. GATA-1 and FOG1-driven transcription slows, and RUNX1 mediates the switch from mitosis to endomitosis by suppressing the Rho/ROCK signaling pathway

  2. In response to the reduced Rho/ROCK signal, inadequate levels of actin and myosin assemble in the cytoplasmic constrictions where separation would otherwise occur, preventing cytokinesis

  3. Subsequently, under the influence of transcription factor NF-E2, DNA replication proceeds to the production of 8N, 16N, or even 32N ploidy with duplicated chromosome sets
• Megakaryocytes


  • Employ their multiple DNA copies to synthesize abundant cytoplasm, which ultimately differentiates into platelets
• Terminal megakaryocyte differentiation


  Megakaryocyte progenitors leave the proliferative phase and enter a series of stages in which they can be recognized by their unique Wright-stained morphology in bone marrow aspirate films or hematoxylin and eosin-stained bone marrow biopsy sections
• Terminal megakaryocyte differentiation stages

  • Megakaryoblast (MK-I)
  • Promegakaryocyte (MK-II)
  • Megakaryocyte (MK-III)
• Megakaryoblast (MK-I)


  • Resembles the myeloblast and pronormoblast (rubriblast), identification by morphology alone is inadvisable
  • Has cytoplasmic "blebs" that resemble platelets
  • Begins to develop most of its cytoplasmic ultrastructure, including procoagulant-laden α-granules, dense granules, and the demarcation system (DMS)
• Promegakaryocyte (MK-II)


  • Cytoplasm is abundant, and the nucleus has minimal lobularity
• Megakaryocyte (MK-III)


  • Nucleus is lobulated with basophilic chromatin
  • Cytoplasm is azurophilic and granular, with evidence of the demarcation system (DMS)
• Thrombocytopoiesis (platelet shedding)


  1. A single megakaryocyte may shed 2000 to 4000 platelets

  2. In an average-size healthy human there are 108 megakaryocytes producing 1011 platelets per day

  3. The total platelet population turns over in 8 to 9 days
• Thrombocytopoiesis


  • The DMS dilates, longitudinal bundles of tubules form, proplatelet processes develop, and transverse constrictions appear throughout the proplatelet processes
  • Proplatelet processes are believed to pierce through or between sinusoid-lining endothelial cells, extend into the venous blood, and shed platelets
• Megakaryocyte membrane markers


  • MPL (TPO receptor)
  • CD34 (stem cell and common myeloid progenitor marker)
  • CD41/CD61 (platelet membrane glycoprotein IIb/IIIa)
  • CD36 (platelet GP 4)
  • CD42 (GP Ib)
  • CD62 (P-selectin)
  • Coagulation factor VIII
  • von Willebrand factor (VWF)
  • Fibrinogen
• Thrombopoietin (TPO)


  A 70,000 Dalton molecule that possesses 23% homology with the red blood cell-producing hormone erythropoietin, and is the ligand that binds the megakaryocyte and platelet MPL receptor
• TPO mRNA has been found in the kidney, liver, stromal cells, and smooth muscle cells, though the liver has the most copies and is considered the primary source
• Thrombopoietin (TPO)


  Hormone that binds to the megakaryocyte and platelet membrane receptor protein MPL, named for the viral oncogene v-mpl associated with murine myeloproliferative leukemia
• TPO


  • Plasma concentration is inversely proportional to platelet and megakaryocyte mass, implying that membrane binding and consequent removal of TPO by platelets is the primary platelet count control mechanism
  • Induces stem cells to differentiate into megakaryocyte progenitors and further induces the differentiation of megakaryocyte progenitors into megakaryoblasts and megakaryocytes
  • Induces the proliferation and maturation of megakaryocytes and induces thrombocytopoiesis
• TPO mimetics (analogs)


  • Romiplostim (NPlate, Amgen)
  • Eltrombopag (Promacta, Novartis)
• Romiplostim


  A nonimmunogenic oligopeptide TPO mimetic that is effective in raising the platelet count in patients with chronic immune thrombocytopenic purpura (ITP)
• Eltrombopag


  A nonpeptide TPO mimetic that binds an MPL site separate from romiplostim, used in the treatment of chronic ITP and in patients with thrombocytopenia resulting from chronic hepatitis C or severe aplastic anemia
• Cytokines that function with TPO to stimulate megakaryocytopoiesis

  • Interleukin-3 (IL-3)
  • IL-6
  • IL-11
• IL-3


  Acts in synergy with TPO to induce the early differentiation of stem cells
• IL-6 and IL-11


  Act in the presence of TPO to enhance endomitosis, megakaryocyte maturation, and thrombocytopoiesis
• IL-11 polypeptide mimetic


  • Oprelvekin (Neumega, Pfizer), stimulates platelet production in patients with chemotherapy-induced thrombocytopenia
• Other cytokines and hormones that participate synergistically with TPO and the interleukins

  • Stem cell factor (kit ligand or mast cell growth factor)
  • Granulocyte-macrophage colony-stimulating factor (GM-CSF)
  • Granulocyte colony-stimulating factor (G-CSF)
  • Acetylcholinesterase-derived megakaryocyte growth stimulating peptide
• Factors that inhibit in vitro megakaryocyte growth, indicating they may have a role in the control of megakaryocytopoiesis in vivo

  • Platelet factor 4 (PF4)
  • B-thromboglobulin
  • Neutrophil-activating peptide 2
  • IL-8
• Transcription factors that diminish megakaryocytopoiesis at different stages

  • FOG1 (progenitor stage)
  • GATA-1 (endomitotic stage)
  • NF-E2 (terminal maturation stage)
• Reticulated platelets


  Also known as "stress platelets", appear in compensation for thrombocytopenia, are markedly larger than ordinary mature circulating platelets, carry free ribosomes and fragments of rough endoplasmic reticulum, and are potentially prothrombotic and may be associated with increased risk of cardiovascular disease
• Reticulated platelets can be quantitatively evaluated using nucleic acid dyes like thiazole orange, but platelet dense granule nucleotides may interfere with this measurement, falsely raising the reticulated platelet count
• Platelet plasma membrane


  • Composed of a phospholipid bilayer with polar head groups oriented toward the aqueous blood plasma (neutral phospholipids) and platelet cytoplasm (charged phospholipids), and nonpolar fatty acid tails that orient toward the center
  • Contains esterified cholesterol that maintains membrane integrity and function
  • Anchors glycoproteins and proteoglycans that support surface glycosaminoglycans, oligosaccharides, glycolipids, and essential plasma surface-oriented glycosylated receptors
• Surface-connected canalicular system (SCCS)

  Plasma membrane invades the platelet interior, producing a sponge-like system that enables the platelet to store additional quantities of the same hemostatic proteins found on the glycocalyx, and allows for enhanced interaction of the platelet with its environment
• Platelet


  • Thick adhesive surface layer that responds readily to hemostatic demands
  • Maintains a negative surface charge that repels other platelets, other blood cells, and the endothelial cells that line the blood vessels
• Surface-Connected Canalicular System (SCCS)

  Plasma membrane invades the platelet interior, producing a unique sponge-like system that enables the platelet to store additional quantities of the same hemostatic proteins found on the glycocalyx and allows for enhanced interaction of the platelet with its environment