Platelets in Hemostasis

• Megakaryocytes, the precursors of platelets, arise from the common myeloid progenitor, CCFU-GEMM, under the influence of the transcription gene product, GATA-1, as regulated by cofactor Fog1.
• Megakaryocyte differentiation is suppressed by another transcription gene product, MYB, so GATA-1 & MYB act in opposition to balance megakaryopoiesis in one arm with differentiation to the RBC line in another arm.
• From the myeloid progenitor, there arise three megakaryocyte lineage-committed progenitor stages: Burst-Forming Unit (BFU-Meg), Colony-Forming Unit (CFU-Meg), and Light Density Colony-Forming Unit (LD-CFU-Meg).
• The BFU-Meg, CFU-Meg, and LD-CFU-Meg are diploid and participate in normal mitosis, maintaining a viable pool of megakaryocyte progenitors.
• The proliferative properties of these stages are reflected in their ability to form hundreds (BFU-Megs) or dozens (CFU-Megs) of colonies in culture.
• Endomitosis is a form of mitosis that lacks telophase and cytokinesis, resulting in no daughter cells.
• Some megakaryocyte nuclei replicate 5x, reaching 128 N, which is unusual and may signal hematologic disease.
• As GATA-1 and FOG1 transcription slows, another transcription factor, RUNX1, mediates the switch from mitosis to endomitosis by suppressing the Rho/ROCK signaling pathway, which suppresses the assembly of the actin cytoskeleton.
• Endoreduction is a step under endomitosis.
• Polyploidization is the process of increasing DNA content without cytoplasmic division, resulting in no identical daughters being produced.
• In response to the suppression of the actin cytoskeleton, inadequate levels of actin and myosin assemble in the cytoplasmic constrictions where separation would otherwise occur, preventing cytokinesis.
• Subsequently, DNA replication proceeds under the influence of another transcription factor, NF-E2, to the production of 8N, 16N or even 32N ploidy with duplicated chromosome sets.
• The LD-CFU-Meg does not have the capacity to divide, but retains its DNA replication and cytoplasmic maturation, a partially characterized form of mitosis unique to megakaryocytes known as Endomitosis.
• CD34 is a stem cell and common myeloid progenitor marker that disappears as differentiation proceeds.
• Megakaryocyte membrane markers that can be measured by flow cytometry include MPL and CD34.
• In megakaryocyte cultures examined by TEM, dilation of DMS leads to the formation of longitudinal bundles of tubules, providing a structural framework for cells to take shape.
• Development of proplatelet processes in the BM environment is believed to involve proplatelet processes piercing through or between sinusoid-lining endothelial cells, extending into the venous blood, and shedding platelets.
• Thrombopoiesis leaves behind naked megakaryocyte nuclei to be consumed by marrow macrophages.
• Platelet budding/shedding cannot be reliably observed in megakaryocytes in situ, even in well-structured BM biopsy preparations.
• The TPO receptor site is present at all maturation stages of megakaryocytes.
• Megakaryopoiesis and thrombopoiesis are controlled by hormones and cytokines.
• Platelet membrane glycoprotein (GP) IIb/IIIa (CD41/CD61) first appears on megakaryocyte progenitors and remains present throughout maturation, along with the following immunologic markers: CD36 (PLATELET GP 4), CD42 (GP Ib), and CD62 (P-selectin).
• Factor VIII, VWF, and fibrinogen are detected in the fully developed megakaryocyte by immunostaining.
• TPO induces stem cells to differentiate into megakaryocyte progenitors, the differentiation of megakaryocyte progenitors into megakaryoblasts and megakaryocytes, the proliferation and maturation of megakaryocytes, and thrombopoiesis.
• Acting in the presence of TPO, IL-6 enhances endomitosis, megakaryocyte maturation, and thrombopoiesis.
• mRNA for TPO has been found in the kidney, liver, stromal cells, and smooth muscle cells.
• Early acting multilineage cytokine IL-3 functions with TPO to stimulate megakaryopoiesis.
• The liver has the most copies of mRNA for TPO and is considered the primary source.
• Megakaryocyte-specific cytokine TPO (Thrombopoietin) is a growth factor with a molecular weight of 70,000 D and possesses 23% morphology with the RBC-producing hormone EPO.
• The ligand that binds the megakaryocyte and platelet membrane receptor protein, MPL (named for v-mpl), is a viral oncogene associated with murine myeloproliferative leukemia.
• Stem cell factor (aka Kit-ligand or Mast cell growth factor) and granulocyte-macrophage colony stimulating factor (GM-CSF) participate synergistically with TPO and interleukins.
• Acting in the presence of TPO, IL-11 enhances endomitosis, megakaryocyte maturation, and thrombopoiesis.
• TPO circulates as a hormone in plasma.
• The plasma concentration of TPO 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.
• Thrombokinetics: Platelet life span and turnover rate indicate that under healthy steady-state conditions, the rate of platelet release from the megakaryocytes is equivalent to the rate of platelet removal from the circulation.
• Normal platelets carry FREE RIBOSOMES and FRAGMENTS OF RER, analogous to RBC reticulocytes.
• Nucleic acid dyes such as THIAZOLE ORANGE bind the RNA of the ER and are exploited by hema instruments to provide a quantitative evaluation of reticulated platelet production.
• The presence of these free ribosomes and fragments of reticulocytes triggers speculation that normal platelets arise from early and rapid proplatelet extension and release.
• The net rate of production is expressed as PLATELET TURNOVER.
• Normal platelets exhibit EDTA-induced changes that cause them to round up and CITRATED WB become cylindrical and beaded, resembling fragments of megakaryocyte proplatelet processes.