Platelets-intro.pdf

lOMoARcPSD|51543588 Hematology - Week 2 BS Medical Technology (University of Negros Occidental-Recoletos) Scan to open on Studocu Studocu is not sponsored or endorsed by any college or university Downloaded by...

lOMoARcPSD|51543588 Hematology - Week 2 BS Medical Technology (University of Negros Occidental-Recoletos) Scan to open on Studocu Studocu is not sponsored or endorsed by any college or university Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588 HEMATOLOGY II CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION TABLE OF CONTENTS → LD-CFU-Meg – the more mature progenitor, light density I. MEGAKARYOCYTOPOIESIS colony forming unit A. Megakaryocyte IV. PLATELET ACTIVATION  All three progenitor stages resemble lymphocytes and Differentiation and A. Adhesion: Platelets cannot be distinguished by Wright stain light Progenitors Bind Elements of the microscopy. B. Terminal Vascular Matrix B. Aggregation:  BFU and CFU-Meg are diploid and undergo normal Megakaryocyte Differentiation Platelets Irreversible mitosis to maintain a viable pool of megakaryocyte C. Megakaryocyte Cohere progenitors. BFU form hundreds while CFU form dozens Membrane Receptors C. Secretion: Activated of colonies in culture. and Markers Platelets Release  The third stage LD-CFU-Meg undergoes endomitosis D. Hormones and Granular Contents D. Generation of which is a partially characterized form of mitosis unique Cytokines of Megakaryocytopoiesis Platelet to megakaryocytes in which DNA replication and II. PLATELETS Microparticles cytoplasmic maturation are normal, but cells lose their III. PLATELET V. PLATELET ACTIVATION capacity to divide. ULTRASTRUCTURE PATHWAYS A. Resting Plasma A. G-Proteins Membrane B. Eicosanoid Synthesis B. TERMINAL MEGAKARYOCYTE DIFFERENTIATION B. Surface-Connected C. Inositol As endomitosis proceeds, megakaryocyte progenitors leave Canalicular System Triphosphate- the proliferative phase and enter terminal differentiation. C. Dense Tubular Diacylglycerol  A series of stages in which microscopists become able System Activation Pathway to recognize their unique Wright-stained morphology. D. Cytoskeleton: VI. PLATELET PROTEOME VII. R → Megakaryoblast (MK-I stage) Microfilaments and Microtubules VIII. Citation  the least differentiated megakaryocyte precursor. E. Platelet Granules: α- IX. References  Cannot be reliably distinguished from myeloblast or Granules, Dense pronormoblast using light microscopy. Granules, and  Morphologists may occasionally see a vague clue which Lysosomes F. Plasma Membrane are plasma membrane blebs or blunt projections that Receptors That resemble platelets. Provide for Adhesion  The megakaryoblast begins to develop its ultrastructure G. The Seven- including α-Granules, dense granules, and the Transmembrane demarcation system. Repeat Receptors o Is a series of membrane-lined channels that I. MEGAKARYOCYTOPOIESIS invade from the plasma membrane and grow inward to subdivide the entire cytoplasm. Platelets are nonnucleated blood cells. Platelets circulate at a concentration of 150 to 400 x 109/L Platelet counts are slightly higher in women than in men Platelets are lower in both sexes who are older than 65 years old. Platelets arise from unique bone marrow cells called Megakaryocytes.  the largest cells in the bone marrow  30 to 50 μm in diameter, multilobulated nucleus and abundant granular cytoplasm  In a normal wright stained bone marrow aspirate smear, 2-4 megakaryocytes per 10 magnification low power field may identify A. MEGAKARYOCYTE DIFFERENTIATION AND PROGENITORS Megakaryocyte progenitors arise from the common myeloid progenitor under the influence of the transcription → Promegakaryocyte (MK-II stage) gene product, GATA-1.  Nuclear lobularity becomes apparent as an indentation Megakaryocyte differentiation is suppressed by another rendering the cell identifiable as an MK-II stage or transcription product, MYB, so GATA-1 and MYB act in promegakaryocyte. opposition to balance megakaryocytopoiesis. → BFU-Meg – least mature, burst forming unit → CFU-Meg – the intermediate colony forming unit Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 1 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588 → Megakaryocyte (MK-III stage) C. MEGAKARYOCYTE MEMBRANE RECEPTORS AND  At the most abundant MK-III stage, megakaryocyte is MARKERS easily recognized at 10 magnification on the basis of its Immunostaining of fixed tissue, flow cytometry with 30 to 50 μm diameter. immunologic probes, and FISH with genetic probes are used  The nucleus is intensely indented or lobulated. The to identify visually indistinguishable megakaryocyte chromatin is variably condensed with light and dark progenitors. patches. The cytoplasm is azurophilic, granular, and There are several megakaryocyte membrane markers that platelet-like because of the spread of the demarcation can be measured including MPL and CD34. system in alpha granules. o The CD34 marker disappears as differentiation proceeds. The platelet membrane glycoprotein IIb/IIIa first appears on megakaryocyte progenitors and remains present throughout maturation along with CD36, CD42, and CD62. Cytoplasmic coagulation factor VIII, Von Willebrand factor, and fibrinogen may be detected in the fully developed megakaryocyte by immunostaining. D. HORMONES AND CYTOKINES OF MEGAKARYOCYTOPOIESIS → Thrombopoietin (TPO)  70,000 Dalton molecule that possesses 23% homology with the RBC producing hormone, erythropoietin.  mRNA for TPO has been found in the kidneys, liver, stromal cells, and smooth muscle cells. → Thrombocytopoiesis (Platelet Shedding)  The liver is considered the primary source of  At full maturation, platelet shedding, or Thrombopoietin. Thrombocytopoiesis, proceeds.  The plasma concentration of TPO is inversely  The total platelet population turns over in 8 to 9 days proportional to platelet and megakaryocyte mass; (the so-called platelet lifespan). membrane binding and consequent removal of TPO by  A single megakaryocyte may shed 2,000 to 4,000 platelets is the primary platelet count control platelets. The total platelet population turns over in 8- mechanism. 9 days, the platelet lifespan.  Thrombopoietin in synergy with other cytokines  In the bone marrow environment proplatelet processes induces stem cells to differentiate into megakaryocyte are believed to pierce through or between sinusoid progenitors and that it further induces the lining endothelial cells extend into the venous blood differentiation of megakaryocyte progenitors into and shed platelets. Megakaryocyte is adjacent to megakaryoblasts and megakaryocytes. endothelial cell of the bone marrow and extends a  TPO also induces the proliferation and maturation of proplatelet process through or between the endothelial megakaryocytes and induces Thrombocytopoiesis. cells into the vascular sinus. Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 2 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588  Other cytokines that function with TPO to stimulate  Fatty acid chains esterified to carbons 1 and 2 of the megakaryocytopoiesis includes IL-3, IL-6, and IL-11. phospholipid triglyceride backbone orient toward each  IL-3 seems to act in synergy with TPO to induce the other perpendicular to the plane of the membrane to early differentiation of stem cells whereas IL-6 and IL- form a hydrophobic barrier sandwiched within the 11 act in the presence of TPO to enhance endomitosis, hydrophilic layers. megakaryocyte maturation, and Thrombocytopoiesis.  The neutral phospholipids phosphatidylcholine and sphingomyelin predominate in the outer blood plasma  Other cytokines and hormones that participate layer. synergistically with TPO and the ILs are kit ligand or  The anionic or polar phospholipids mast cell growth factor, GM-CSF, G-CSF and phosphatidylinositol, phosphatidylethanolamine, and acetylcholinesterase-derived megakaryocyte grown phosphatidylserine predominate in the inner stimulating peptide. cytoplasmic layer. Platelet factor 4, β-thromboglobulin, neutrophil activating  Cholesterol stabilizes the membrane, maintains fluidity, peptide 2, IL-8, and other factors inhibit in vitro and helps control the transmembranous passage of megakaryocyte growth which indicates that they may have a materials through the selectively permeable plasma role in the control of megakaryocytopoiesis in vivo. membrane.  Anchored within the membrane are glycoproteins and proteoglycans that support surface II. PLATELETS glycosaminoglycans, oligosaccharides, glycolipids, and essential plasma surface-oriented glycosylated cells consisting of granular cytoplasm with a membrane but receptors that respond to cellular and humoral stimuli, no nucleus, into the venous sinus of the bone marrow called ligands or agonists, transmitting their stimulus On a Wright-stained wedge- preparation blood film, platelets through the membrane to activation organelles internal are distributed throughout the red blood cell monolayer at 7 to the platelet. to 21 cells per 100 magnification field, and they average 2.5  The platelet membrane surface, called the glycocalyx μm in diameter absorbs albumin, fibrinogen, and other plasma MPV ranges from 8 to 10 fL. Heterogeneity in the MPV of proteins, in many instances transporting them to normal healthy humans reflects random variation in platelet internal storage organelles using a process called release volume and is not a function of platelet age or vitality. endocytosis. At 20 to 30 nm the platelet glycocalyx is Resting platelets are biconvex. The platelets in blood thicker than the analogous surface layer of leukocytes collected using EDTA tend to “round up”. or erythrocytes. The normal peripheral blood platelet count is 150 to 400 x SURFACE-CONNECTED CANALICULAR SYSTEM 109/L. The platelet count decreases with increasing age, such  The plasma membrane invades the platelet interior, that after 65 years of age, platelet counts of 122 to 350 x producing a unique surface-connected canalicular 109/L and 140 to 379 x 109/L are seen in men and women, system respectively. This count represents only two thirds of total  twists sponge-like throughout the platelet, enabling the body platelets; the remaining one third is sequestered within platelet to store additional quantities of the same the spleen. hemostatic proteins found on the glycocalyx Reticulated platelets sometimes known as stress platelets  allows for enhanced interaction of the platelet with its appear in compensation for thrombocytopenia. environment, increasing access to the platelet interior Are markedly larger than ordinary mature circulating as well as increasing egress of platelet release products. platelets; diameter in blood films exceeds 6 μm and their MPV reaches 12 to 14 fL. DENSE TUBULAR SYSTEM They also round up in EDTA but are cylindrical and beaded in  Is parallel and closely aligned to the SCCS citrated whole blood (light blue tubes).  A condensed remnant of the rough endoplasmic reticulum.  Sequesters Ca2+ and bears a number of enzymes that III. PLATELET ULTRASTRUCTURE support platelet activation including phospholipase A2, RESTING PLASMA MEMBRANE cyclooxygenase, thromboxane synthetase &  The platelet plasma membrane resembles any biologic phospholipase C. membrane: a bilayer composed of proteins and lipids.  The predominant lipids are phospholipids which form CYTOSKELETON: MICROFILAMENTS AND MICROTUBULES the basic structure and cholesterol which distributes  a thick circumferential bundle of microtubules asymmetrically throughout the phospholipids. maintains the platelet’s discoid shape.  Phospholipids form a bilayer: polar heads oriented  Circumferential microtubules parallel the plane of the toward aqueous environments toward blood plasma outer surface of the platelet and reside just within, externally and the cytoplasm internally. although not touching, the plasma membrane. Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 3 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588 CYTOSKELETON: MICROFILAMENTS AND MICROTUBULES  8 to 20 tubules composed of multiple subunits of  CAMs integrate their ligands which they bind on the tubulin that disassemble at refrigerator temperature outside of the cell with the internal cytoskeleton or when platelets are treated with colchicine. When triggering activation. microtubules disassemble in the cold platelets  GP Ia/IIa or α2β1 is an integrin that binds the become round, but on warming to 37° C they recover subendothelial collagen that becomes exposed in the their original disc shape. On cross section, damage blood vessel wall, promoting adhesion of the microtubules are cylindrical with a diameter of 25 nm. platelet to the vessel wall. The circumferential microtubules could be a single  α5β1 and α6β1 bind laminin and fibronectin, spiral tubule. adhesive endothelial cell proteins which further  Between the microtubules and the membrane lies a promotes platelet adhesion. thick meshwork of microfilaments composed of actin.  GP VI is another collagen-binding receptor from the Actin is contractile in platelets (as in muscle) and immunoglobulin gene family. GP IV is a key collagen anchors the plasma membrane glycoproteins and receptor that also binds the adhesive protein proteoglycans. It is also present throughout the thrombospondin. platelet cytoplasm, constituting 20% to 30% of platelet  GP Ib/IX/V is a leucine-rich-repeat family CAM that protein. In the resting platelet, actin is globular and arises from the genes GP1BA, GP1BB, GP5 and GP9. It amorphous. As cytoplasmic calcium concentration is composed of two molecules each of GP Ibα, GP Ibβ, rises actin becomes filamentous and contractile. and GP IX and one molecule of GP V which are bound  The cytoplasm also contains intermediate filaments, noncovalently. rope-like polymers 8 to 12 nm in diameter, of desmin and vimentin. THE SEVEN-TRANSMEMBRANE REPEAT RECEPTORS  Thrombin, thrombin receptor activation peptide PLATELET GRANULES: α-Granules, Dense Granules, and (TRAP), adenosine diphosphate, epinephrine, Lysosomes serotonin, thromboxane A2 and other prostaglandins  There are 50 to 80 α-granules in each platelet which can function individually or in combination to activate stain medium gray in osmium-dye transmission platelets. electron microscopy preparations. o These platelets antagonists are ligands for  α-granules are filled with proteins. Several α-granules seven-transmembrane repeat receptors and are membrane bound. are named for their unique membrane-  As the platelet becomes activated, α-granule anchoring structure. membranes fuse with the SCCS where their contents  The STRS have seven hydrophobic anchoring domains flow to the nearby microenvironment. They supporting an external binding site and an internal participate in platelet adhesion and aggregation and terminus that interacts with G proteins to mediate support plasma coagulation. outside-in platelet signaling.  There are 2 to 7 dense granules per platelet. These  Thrombin cleaves two STRs, protease-activated appear later than α-granules in megakaryocyte receptor 1 (PAR1) and PAR4, that together have a total differentiation and stain black (opaque) when treated of 1800 membrane copies on an average platelet. with osmium in transmission electron microscopy. o Thrombin cleavage of either of these two  Dense granules migrate to the plasma membrane and receptors activates the platelet through G- release their contents directly into the plasma on proteins that in turn activate at least two platelet activation. internal physiologic pathways.  Membranes of dense granules support the same  Thrombin also interacts with platelets by binding or integral proteins as the α-granules. digesting two CAMs in the leucine-rich repeat family,  Platelets contain a few lysosomes similar to those in GP Iba and GP V, both of which are parts of the GP neutrophils. Ib/IX/V VWF adhesion receptor  There are about 600 copies of the high-affinity ADP PLATELET MEMBRANE RECEPTORS THAT PROVIDE FOR receptors P2Y1 and P2Y12 per platelet. ADHESION  P2Y1 signaling leads to an increase in intracellular  The platelet membrane contains more than 50 distinct calcium levels and contributes to initial platelet receptors including members of the cell adhesion activation, shape change, and the formation of small molecule (CAM) integrin family the seven- reversible aggregates transmembrane receptor family and some  P2Y12 signaling leads to a decrease in cyclic adenosine miscellaneous receptors. monophosphate (cAMP) levels and supports the  Integrins bind collagen, enabling the platelet to formation of irreversible platelet aggregates. adhere to the injured blood. Integrins are o These receptors are linked to different G- heterodimeric or composed of two dissimilar proteins. proteins and produce distinct intracellular signals that have complementary effects on platelet aggregation Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 4 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588  TPα and TPβ bind TXA2. This interaction produces  VWF, circulates as a globular protein. Under shear more TXA2 from the platelet, a G-protein-based stress, VWF becomes thread-like as it unrolls and autocrine (self-perpetuating) system that activates exposes sites that weakly bind the GPIbα portion of neighboring platelets. the platelet membrane GP Ib/ IX/V leucine-rich  Epinephrine binds a2-adrenergic sites that couple to receptor. G-proteins and open membrane calcium channels.  The interaction between platelet and VWF remains o The a2-adrenergic sites function similarly to localized by a liver-secreted plasma enzyme, those located on heart muscle. ADAMTS13, also called VWF-cleaving protease, which  The receptor site IP binds prostacyclin (prostaglandin digests larger VWF multimers into smaller, less I2, PGI2), a prostaglandin produced by endothelial biologically active forms. cells.  Type I fibrillar collagen binding to platelet GP VI,  Prostacyclin binding results in an increase in the which is anchored in the platelet membrane by an Fc internal cAMP concentration of the platelet and an receptor-like molecule, triggers internal platelet inhibition of platelet activation. activation pathways, releasing TXA2 and ADP, an  The platelet membrane also contains STRs for “outside-in” reaction serotonin, platelet-activating factor, prostaglandin E2,  Agonists attach to their respective receptors: TPα and PF4, and β-thromboglobulin. TPβ for TXA2, and P2Y1 and P2Y12 for ADP, triggering an “inside-out” reaction that raises the affinity of ADDITIONAL MEMBRANE RECEPTORS integrin α2β1 for collagen.  The CAM immunoglobulin family includes the  The combined effect of GP Ib/IX/V, GP VI, and α2β1 intercellular adhesion molecules, or ICAMs (CD50, causes the platelet to become firmly affixed to the CD54, CD102), which play a role in inflammation and damaged surface, where it subsequently loses its the immune reaction discoid shape and spreads.  platelet–endothelial cell adhesion molecule, or PECAM (CD31), which mediates platelet-to-white AGGREGATION: PLATELETS IRREVERSIBLE COHERE blood cell and platelet-to-endothelial cell adhesion  Blood vessel injury exposes tissue factor expressed on  FcgIIA (CD32), a low-affinity receptor for the subendothelial smooth muscle cells and fibroblasts. immunoglobulin Fc portion that plays a role in a Tissue factor triggers the production of thrombin, dangerous condition called heparin-induced which cleaves platelet PAR1 and PAR4. thrombocytopenia  The activation generates the “collagen and thrombin  P-selectin (CD62) is an integrin that facilitates platelet activated” or COAT platelet, integral to the cell-based binding to endothelial cells, leukocytes, and one coagulation model another; found on the a-granule membranes of the  The platelet activators TXA2 and ADP are secreted resting platelet but migrates via the SCCS to the from the platelet granules to the microenvironment, surface of activated platelets they activate neighboring platelets through their o P-selectin quantification by flow cytometry respective receptors and trigger inside-out activation is a common means for measuring in vivo of integrin αIIbβ3 (GP IIb/IIIa receptor), enabling it to platelet activation. bind RGD sequences of fibrinogen and VWF and support platelet-to-platelet binding referred to as platelet aggregation. IV. PLATELET ACTIVATION  P-selectin from the a-granule membranes moves to the surface membrane to promote binding of ADHESION: PLATELETS BIND ELEMENTS OF THE VASCULAR platelets with leukocytes. MATRIX  In conjunction with aggregation, platelets change in  shape from discoid to round and extend pseudopods. Vessel walls create stress or shear force, measured in This allows platelets to cover more surface area and it units labeled as s-1 enhances platelet binding to other platelets and  Shear forces range from 500 s-1 in venules and veins to foreign surfaces. 5000 s-1 in arterioles and capillaries and up to 40,000  Membrane phospholipid asymmetry is lost, with the s-1 in stenosed (hardened) arteries. more polar molecules, especially phosphatidylserine,  when the shear rate is more than 1000 s-1, platelet flipping to the outer layer. As platelet aggregation adhesion and aggregation require the process of primary hemostasis: the process of platelet response continues, membrane integrity is lost, and a syncytium to blood vessel injury (refer to figure 10.9, p.147). or massive clump of platelets forms as the platelets  Injury to the blood vessel wall disrupts the collagen of exhaust internal energy sources the extracellular matrix (ECM)  Platelet aggregation is a key part of primary  Damaged endothelial cells release VWF from hemostasis, which in arteries may end with the cytoplasmic storage organelles, which then adheres to formation of a “white clot,” a clot composed primarily sites of injury of platelets and VWF. Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 5 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588  Aggregation is a normal part of vessel repair. The increases the platelet intracellular concentration of presence of white clots often implies inappropriate calcium. platelet activation in seemingly uninjured arterioles  Elevated levels of intracellular calcium result in an and arteries and is the pathologic basis for arterial inhibition of the enzymes responsible for maintaining thrombotic events the asymmetric distribution of phospholipids in the o Such as acute myocardial infarction, plasma membrane and an activation of intracellular peripheral artery disease, and ischemic calpain, which cleaves the platelet cytoskeleton. stroke. The risk of these cardiovascular o lead to the outward blebbing of the plasma events rises in proportion to the number and membrane and the formation of platelet avidity of platelet membrane α2β1 and GP VI microparticles. receptors  The combination of polar phospholipid exposure on → Platelet microparticles: activated platelets, platelet fragmentation with  most abundant microparticles in the circulation cellular microparticle release, and secretion of the  are formed after exposure of platelets to strong platelet’s a-granule and dense granule contents agonists or shear stress. triggers secondary hemostasis, called coagulation.  made up of the plasma membrane and cytosolic  Fibrin and red blood cells deposit around and within material of the parent cell from which they are derived the platelet syncytium to form a bulky “red clot.” and retain the cell surface proteins found on the parent  Red clot is essential to wound repair. It may also be cell characteristic of inappropriate coagulation in venules  microparticles have been found to modulate and veins, resulting in deep vein thrombosis and inflammation, oxidative stress, angiogenesis, and pulmonary embolism. thrombosis. V. PLATELET ACTIVATION PATHWAYS SECRETION: ACTIVATED PLATELETS RELEASE GRANULAR CONTENTS G-PROTEINS  Outside-in activation of the platelet through STRs  G-proteins control cellular activation for all cells at the (such as ADP binding to P2Y12) and the inner membrane surface. immunoglobulin gene product GP VI triggers actin  G-proteins are αβy heterotrimers (proteins composed microfilament contraction. of three dissimilar peptides) that bind guanosine o Intermediate filaments also contract, diphosphate (GDP) when inactive. moving the circumferential microtubules  Membrane receptor–ligand (agonist) binding promotes inward compressing the granules. GDP release and its replacement with guanosine  Contents of a-granules and lysosomes flow through triphosphate (GTP) the SCCS, while dense granules migrate to the plasma  The Gα portion of the three-part G molecule briefly membrane where their contents are secreted. disassociates, exerts enzymatic guanosine  Dense granule contents are small molecule triphosphatase activity, and hydrolyzes the bound GTP vasoconstrictors and platelet agonists that amplify to GDP, releasing a phosphate radical primary hemostasis; most of the a-granule contents  The G-protein resumes its resting state, but the are large molecule coagulation proteins that hydrolysis step provides the necessary phosphorylation participate in secondary hemostasis to trigger eicosanoid synthesis or the IP3-DAG pathway  Phosphatidylserine is the polar phospholipid on which the factor IX/VIII (tenase) and factor X/V EICOSANOID SYNTHESIS (prothrombinase) complexes assemble.  The eicosanoid synthesis pathway, alternatively called o formation of both complexes is supported by the prostaglandin, cyclooxygenase, or thromboxane ionic calcium secreted by the dense pathway, is one of two essential platelet activation granules. pathways triggered by G-proteins found in platelets o The α-granule contents fibrinogen, factors V  The platelet membrane’s inner leaflet is rich in and VIII, and VWF (which binds and stabilizes phosphatidylinositol, a phospholipid whose number 2 factor VIII) are secreted and increase the carbon binds numerous types of unsaturated fatty localized concentrations of these essential acids, but especially 5,8,11,14-eicosatetraenoic acid, coagulation proteins, further supporting the commonly called arachidonic acid. action of tenase and prothrombinase  Membrane receptor-ligand binding and the consequent G-protein activation triggers phospholipase A2, a GENERATION OF PLATELET MICROPARTICLES membrane enzyme that cleaves the ester bond  Microparticles are membrane-derived vesicles that connecting the number 2 carbon of the triglyceride form in response to an activating stimulus that backbone with arachidonic acid. Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 6 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])  lOMoARcPSD|51543588  Cleavage releases arachidonic acid to the cytoplasm, V. PLATELET PROTEOME where it becomes the substrate for cyclooxygenase,  Proteomic analysis has indicated that platelets anchored in the DTS. Cyclooxygenase converts contain thousands of unique transcripts that can be arachidonic acid to prostaglandin G2 and prostaglandin translated in response to platelet activation and H2, and then thromboxane synthetase acts on ligand binding to the GPIIb/IIIa receptor. Such prostaglandin H2 to produce TXA2. mechanisms allow platelets to alter their phenotype  TXA2 binds membrane receptors TPα or TPβ, inhibiting in response to the level of activation. it is suggested adenylate cyclase activity and reducing cAMP that the protein Italics concentrations, which mobilizes ionic calcium from the DTS.  Rising cytoplasmic calcium level causes contraction of actin microfilaments producing platelet shape change and further platelet activation.  When reagent arachidonic acid is used as an agonist in the laboratory assay, it bypasses the membrane and directly enters the eicosanoid synthesis pathway.  The cyclooxygenase pathway in endothelial cells incorporates the enzyme prostacyclin synthetase in place of the thromboxane synthetase found in platelets  The eicosanoid pathway end point for the endothelial cell is prostaglandin I2, or prostacyclin, which binds the IP receptor activating the IP3-DAG pathway, leading to an acceleration of adenylate cyclase, an increase in cAMP, and a sequestration of ionic calcium to the DTS. The unavailability of ionic calcium shuts down platelet function.  Endothelial cell eicosanoid pathway suppresses platelet activation in the intact blood vessel, creating a dynamic equilibrium with the eicosanoid pathway within the platelet where platelet activation occurs.  Thromboxane B2 is acted on by a variety of liver enzymes to produce an array of soluble urine metabolites, including 11-dehydrothromboxane B2, which is stable and measurable. INOSITOL TRIPHOSPHATE-DIACYLGLYCEROL ACTIVATION PATHWAY  The IP3-DAG pathway is the second G-protein- dependent platelet activation pathway.  G-protein activation triggers the enzyme phospholipase C  Phospholipase C cleaves membrane phosphatidylinositol 4,5-bisphosphate (PIP2) to form IP3 and DAG, both second messengers for intracellular activation.  IP3 promotes release of ionic calcium from the DTS, which triggers actin microfilament contraction. IP3 may also activate phospholipase A2.  DAG triggers a multistep process: activation of phosphokinase C, which triggers phosphorylation of the protein pleckstrin, which regulates actin microfilament contraction. Trans # 1 MTY1215_WEEK2_LEC_ CHAPTER 10: PLATELET PRODUCTION, STRUCTURE AND FUNCTION (766-771) 7 of 7 Rodak's Hematology Clinical Principles and Applications 6E – Page 137-150 | Module: LABORATORY EVALUATION OF HEMOSTASIS Downloaded by Bxbbx Xjchx ([email protected])