Platelets and the Vascular System Flashcards
Describe the platelet maturation process, from CFU-Meg (of CFU-MK) to mature platelet.
The CFU-Meg (or CFU-MK) is a progenitor cell found in the bone marrow. Under the influence of humoral factors GM-CSF, IL-3 and TPO, it can differentiate into an immature megakaryocyte. Still in the BM, with further regulation from TPO, the megakaryocyte undergoes endomitosis and matures. This process takes about 4-5 days, after which the megakaryocyte enters the bone marrow sinus. Here, based on the regulation from TPO, the megakaryocyte’s cytoplasm breaks down along its demarcation membrane, producing many platelets which enter into circulation or into the spleen.
What is endomitosis? Is this process common to all cells, or unique to platelets?
Round of nuclear DNA synthesis without nuclear or cytoplasmic division
Unique to platelets
The four platelet zones
Peripheral zone
Structural zone
Organelle zone
membrane zone
Peripheral platelet zone
Function
Components
Membrane
Function: Adhesion and aggregation, contributes to platelets, overall neg charge and contain receptors
Components:
factor 5, von Willebrand factor and fibrinogen
Glycocalyx: proteins, glycoproteins and glycopolysaccharides
Membrane:
GP Ib/IX: receptor for von Willebrand factor and important for platelet adhesion
GP IIb/IIIa: receptor for Fibrinogen
Structural platelet zone
Function
Components
Function: Structure and Support
Components • Microtubules (tubulin) • Protein Network o Actin 15-20% of platelet proteins Globular (G) and Filamentus (F)—active form o Other contractile proteins
Organelle platelet zone
Function
Components
Function: Secretion and Storage
Components:
- Alpha and Delta Granules
- ADP —Agonist for platelets
- ATP —Agonist for cells other than platelets, Activates Ca+2 influx
- Calcium—Platelet activation
- Serotonin—Vasoconstriction; platelet agonist
Membrane Systems (platelet zone)
Function
Components
Functions: Secretion and Storage; Regulate Intracellular (Calcium)
Components:
• Open Canalicular system—connect the inside of platelet with membrane
• Dense Tubular system canals running within the cell, stores and regulates, prostaglanding and thromboxane synthesis
Steps of primary hemostasis
adhesion
aggregation
secretion
activation
Primary Hemostasis is measured by what?
Bleeding time
Platelet function test
Platelet aggregation test
Platelet role in adhesion
Platelet adheres to collagen from injured tissue with vWF as a bridge. Platelets have GPIb/IX receptors for vWF
Platelet role in aggregation
- Changes occurs in shape (from disks to spiny spheres)
- Metabolic biochem occurs with movement of intracellular calcium from dense bodes through the membrane to the cytoplasm. Phospholipase C and A2 and adenyl cyclase activate
- GP IIb/IIIa receptor for fibrinogen appears
- Membrane surface changes occur and allow coag factors to bind. Calcium, collagen and ADP are main agonists
Platelet role in secretion
The contents of the granules are secreted to the outside of the platelet through the open canalicular system after aggregation. Agonists attract more platelets
Platelet role in secondary hemostasis
Platelet contractile proteins causes the fibrin clot to shrink, which strengthens the clot. Platelets provide a surface for the reactions.
Describe the main functions of blood vessels in hemostasis.
Damaged blood vessels initiate hemostasis through vasoconstriction by interaction with thromboxane A2 and serotonin which are released from activated platelets. Heparin sulfate and thrombomodulin are released from the endothelial cells and inhibit the fibrin formation while vWF and tissue thromboplastin result in fibrin formation and platelet adhesion
Thrombocytopenia
deficiency of platelets in the blood <150 x 109/L
Thrombocytosis
disorder in which the body produces too many platelets. >400 x 109/L
Thrombophilia
tendency to form blood clots abnormally
Marfan Syndrome
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Vascular disorder
Pathophysiology: Problem with Fibrillin
Abnormal structure/elasticity of vessels
Gene for fibrillin-1
Clinical Symptoms: Aortic aneurism, defects in mitral valve, death is common, long extremities hyper extendable joints
Diagnostic Lab Tests and results: Diagnosis by exclusion; bleeding time and platelet aggregation may be abnormal
Osteogenesis Imperfecta
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Vascular disorder
Pathophysiology:
Patchy defective BM leading to brittle bones that fracture easily.
Gene for type 1 procollagens
Clinical Symptoms: Intercranial hemorrhage due to collapsed cranial bones, epistaxis, coughing up blood and spontaneous bruising
Diagnostic Lab Tests and results: Diagnosis by exclusion, platelet aggregation may be abnormal
Ehlers-Danlos Syndrome
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Vascular disorder
Pathophysiology: Lack of collagen creating fragile vessels
Clinical Symptoms: Easy bruising, stretchable fragile skin, hyper extendable joints
Diagnostic Lab Tests and results: Diagnosis by exclusion; bleeding time may be abnormal
Hereditary Hemorrhagic Telangiectasia
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Vascular disorder
Pathophysiology: No capillaries, arterioles connect directly to the venioles
Over dilation causes vessels to be thin and fragile and prone to rupture
Gene for endolin-1
Clinical Symptoms: Bleeding symptoms progress with age
Starts with Epistaxis and leads to more dangerous bleeding
Diagnostic Lab Tests and results: Diagnosis by exclusion, gene testing
Pseudoxanthoma Elasticum
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Vascular disorder
Pathophysiology: Calcification of elastic tissue in skin and arteries
Clinical Symptoms: Appears 20-30 year old
Hemorrhage in GI, eyes, kidneys, nose, skin, easy bruising and petechia
Diagnostic Lab Tests and results: Diagnosis by exclusion
Purpura due to decreased connective tissue
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Acquired Vascular disorder
Pathophysiology:
- -Senile purpura: caused by ↓collagen with age, results in vessels prone to break
- -Cushing syndrome: from supportive tissue that is weakened because of ↑ glucopolysaccharides that are present in condition or result from cortical steroid therapy
- -Scurvy: deficiency in vit C, causes ↓collagen production resulting in vessels that are weakened
Clinical Symptoms: Bleeding, bruising and petechiae
Diagnostic Lab Tests and results: specific to cause
Purpura associated with paraprotein disorders
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Acquired Vascular disorder
Pathophysiology: Paraproteins bind (deplete) calcium; paraprotein complexes deposited in vessel wall
Clinical Symptoms: Nonspecific bruising and petechia
Diagnostic Lab Tests and results: Specific to cause
Purpura due to vasculitis
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Acquired Vascular disorder
Pathophysiology: Immune complexes bind to vessel wall, activating complement
Clinical Symptoms: Nonspecific bruising and petechia
Diagnostic Lab Tests and results: Specific to cause
Heparin induced Thrombocytopenia (HIT)
Defective Body System
Pathophysiology
Clinical Symptoms
Lab Tests
Defective Body System: Platelet disorder
Pathophysiology: Heparin/antibody/PF4 platelet complex initiates thrombosis, rapidly depleting platelet count
Clinical Symptoms: Thrombosis petechiae and/or bleeding
Diagnostic Lab Tests and results: Platelet count decreased PTT increased (heparin present)
Disseminated Intravascular Coagulation (DIC)
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Acquired disorder of hemostasis
Pathophysiology: Consumption of coagulopathy
Systemic: circulating thrombin and unregulated platelets
Unregulated thrombin generation: depletion of I, II, V, VIII, XIII, platelet activation
Clinical Symptoms: Life threatening (50%)
Bleeding and clotting simultaneously
Acute: most common (80-90%) bleeding symptoms predominate with bleeds @ 3 or more sites
Chronic: thrombotic systems predominate leading to vessel occlusion and organ failure
Diagnostic Lab Tests and results: Platelet count decreased
PT, PTT, DD, TT elevated
Aspirin ingestion
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Platelet disorder
Pathophysiology: Aspirin inactivates cyclooxygenase, preventing the formation and release of ThromboxaneA2, and thus platelet secretion
Clinical Symptoms: Bleeding patients with underlying bleeding conditions or in cases of overdose
Diagnostic Lab Tests and results: Abnormal platelet aggregation test no secondary wave for ADP or epinephrine
Acute Immune Thrombocytopenia Purpura
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Platelet disorder
Pathophysiology: Immune mediated destruction of platelets
Clinical Symptoms: Petechiae
Diagnostic Lab Tests and results: Decreased platelet count
Bernard Soulier Disease
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Qualitative platelet disorder
Pathophysiology: Defective or deficient GPIb/IX receptor complex; results in platelet adhesion problems
Clinical Symptoms: Mild to severe bleeding in homozygotes
Diagnostic Lab Tests and results: abnormal platelet aggregation with ristocetin not corrected with vWF
Glansmann’s Thrombasthenia
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Qualitative platelet disorder
Pathophysiology: Defective or deficient GPIIb/IIIa receptor complex; results in platelet aggregation problems
Clinical Symptoms: Mild to severe bleeding in homozygotes
Diagnostic Lab Tests and results: Normal platelet count; abnormal platelet aggregation test (abnormal ADP, collage and epinephrine) increased bleeding time
Von Willebrand disease
Defective Body System
Pathophysiology
Clinical Symptoms
Diagnostic Lab Tests and Results
Defective Body System: Inherited Secondary Hemostasis disorder
Pathophysiology: Genetic mutation that results in defective vWF processing or disruption of binding sites
Clinical Symptoms: mucosal and gum bleeding, epistaxis
Diagnostic Lab Tests and results:
Platelet aggregation test: Ristocetin is abnormal and Ristocetin + vWF is normal
What is an agonist?
A substance that causes platelet activation
Agonists
Strength, source, and platelet receptors
Collagen Thrombin Epinephrin ADP Serotonin Thromboxane PAF
Collagen - STRONG, Nonplatelet, GPIa/IIa, GPVI
Thrombin - STRONG nonplatelet, PAR1, PAR4
Epinephrin - weak, nonplatelet, A2 adrenergic receptor
ADP - weak, platelet, P2Y1, P2Y12
Serotonin - weak, platelet, 5HT24
Thromboxane - weak, platelet, TP
Platelet activating factor (PAF) - weak, platelet, PAFR
List the common agonist reagents used in platelet aggregation testing, and which component of platelet function (adhesion, aggregation, secretion) it is evaluating.
ADP Collagen Epinephrine Ristocetin Ristocetin + vWF
ADP: primary wave (aggregation) and secondary wave (secretion)
Collagen: secretion
Epinephrine: aggregation, second wave (secretion)
Ristocetin: adhesion
Ristocetin + vWF: adhesion
Template Bleeding Time
Purpose Principle Procedure Normal Reference Intervals Significance of Abnormal Results
Purpose: Evaluate the forming of the primary hemostatic plug
Principle: Measure the amount of time it takes to form the primary hemostatic plug
Procedure:
Dab filter paper on bleed every 30 seconds and count how many spots on paper
Normal Reference Intervals:
1-10 min
Significance of Abnormal Results:
↑ possible vascular or platelet problem
↓ not clinically significant
Manual Platelet Studies
Purpose Principle Procedure Normal Reference Intervals Significance of Abnormal Results
Purpose: Quantitatively evaluate platelets
Principle: Count the number of platelets per a designated unit
Procedure (include calculation):
Manual (Hemocytometer) and Automated
Normal Reference Intervals:
150,000-400,000/uL
Significance of Abnormal Results:
↑ thrombocytosis
↓ Thrombocytopenia
Platelet Aggregation Studies
Purpose
Principle
Procedure
Normal Reference Intervals
Purpose: Qualitatively evaluate platelets
Principle: Measures % transmittance in patient’s plasma after being mixed with agonist
Procedure:
Varying reagent agonists are added to whole blood sample; platelet aggregation monitored by aggregometry, producing a pattern or curve that is compared to a normal curve to determine the type of faulty platelet function
Normal Reference Curves:
ADP, Epinephrine — has two waves
Collagen, Ristocetin, Ristocetin + vWF – only one wave
Platelet Estimate (from a peripheral blood smear)
Purpose
Procedure
Normal Reference Intervals
Significance of Abnormal Results
Purpose: screen peripheral blood for platelet quantity; verify automated platelet count
Procedure: Determine the average # of platelets per 100x field; multiply average by 20,000 to get /uL
Normal Reference Intervals: Should correlate with automated value
Significance of Abnormal Results: indicates problem with the automated analyzer
Platelets
Production Distribution Lifespan Normal Reference Intervals Morphology
Production: Demarcation membranes form throughout megakaryocyte cytoplasm forming proplatelets that further breakdown into platelets
Distribution: 2/3 in active circulation, 1/3 sequestered in spleen
Lifespan: 9-12 days in circulation
Normal Peripheral Blood Reference Intervals: 150-400 x 109/L (150,000-450,000/uL)
Morphology: 2-3um; anuclear; granular; irregular (heterogeneous) shape
Megakaryocytes
Distribution
Lifespan
Normal Reference Intervals
Morphology
Distribution: Bone Marrow
Lifespan: ~7 days in BM and eventually breaks down into platelets
Normal Bone Marrow Reference Intervals: ~1-3/lpf (10x) feather edge of smear
Morphology: (Stage 1&2) High N:C ratio; cytoplasm more blue; agranular, single lobed nucleus that may have nuclei; fine chromatin in nucleus
(Stage 3&4) Low N:C ratio; abundant, granular cytoplasm with a lavender color; multi-lobed nucleus with condensed chromatin
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
Presence of EDTA-induced clumping
False decrease; analyzer does not count platelet in the clump
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
A clotted EDTA sample
Falsely decreased (they are all in clot and not counted)
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
A sample showing platelet satellitism
Platelets surround WBCs and don’t get counted causing a falsely low count
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
A patient with many larger than normal platelets
Counted as different cell, falsely decrease count
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
A patient with many RBC schistocytes present
False increase; RBC fragments are counted as platelets
Describe how (ex. falsely increase, falsely decrease, no impact) and why each of the following might impact an automated platelet count result.
A patient that has taken aspirin
No impact; aspirin impacts function not quantity
What is aspirin’s “mode of action”?
Aspirin interferes with prostaglandins. It irreversibly inhibits cyclo-oxygenase which is an enzyme needed to make Thromboxane A2 (TXA2). During platelet secretion, TXA2 is released to further activate the platelet.
The normal pathway of TXA2 synthesis:
Increased Phosopholipase A2 causes the release of increased amounts of Arachidonic Acid. Cyclo-oxidase converts Arachidonic Acid to PGG2; PGG2 is converted to TXA2 by Thromboxane synthase.
What impact does aspirin have on platelet function?
Stops the production of Thromboxane A2 (TXA2) which is a platelet activation enhancer
Which of the following laboratory tests would be impacted by aspirin ingestion and what specific effect would it have on the results?
Template bleeding time:
Would be prolonged because the activation is affected, therefore the platelet would struggle to form the platelet plug.
Which of the following laboratory tests would be impacted by aspirin ingestion and what specific effect would it have on the results?
Platelet aggregation studies:
The second wave of aggregation would not occur (where platelets respond to their own agonists) because the activation is inhibited
Which of the following laboratory tests would be impacted by aspirin ingestion and what specific effect would it have on the results?
Platelet function test:
It would be affected because TXA2 is not produced
Which of the following laboratory tests would be impacted by aspirin ingestion and what specific effect would it have on the results?
Platelet count:
Not affected
Which of the following laboratory tests would be impacted by aspirin ingestion and what specific effect would it have on the results?
Clotting times (ex. PT, aPTT):
No impact because they evaluate clotting factors not platelets
Hemolytic Uremic Syndrome
Pathophysiology
Symptoms
Pathophysiology: verotoxin producing E coli (VTEC) or inherited, Likely due to toxin induced endothelial damage (primarily in renal glomeruli) that results in microvascular thrombi
Symptoms: Hemolytic anemia, acute renal failure that is more severe than TTP, thrombocytopenia, CNS involvement, microangiopathic hemolytic anemia, fever
Neonatal Alloimmune Thrombocytopenia
HPA-1a is most common platelet antigen (97% population has this antigen)
Maternal IgG anti-HPA-1a—can cross placenta and attack baby’s platelets
Posttransfusion Purpura
Rare, results in abrupt bleeding of mucous membranes and lasts 2-6 weeks
7-10 days post transfusion
The donor contains Anti-HPA-1a, when given, attacks the platelets of receiver
Thrombotic Thrombocytopenic Purpura
Large vWF multimers attach to platelet surface—induce agglutination
Acquired or inherited deficiency of ADAMTS-13 (responsible for cleaving large vWF multimers)
Blood smear: acanthocyte, schistocytes, micro RBCs, and platelets
Increased Splenic Sequestration of Platelets
Normally—spleen sequesters 1/3 of platelets from BM
Usually the platelet count doesn’t drop below 20 x 109/L therefore not life threatening
Dilutional Thrombocytopenia
Caused by massive hemorrhage, occurs when patients recieve disproportionate amount of RBCs than platelets in transfusion
>10 units packed RBCs in 24 hours
Platelets and plasma proteins become depleted in proportion to amount of RBCs
Primary Thrombocytosis
Platelet count: >1,000,000/uL
Pathophysiology: Essential Thrombocytosis, CML, Myeloproliferative diseases
Labs: Platelet count increased, Abnormal platelet aggregation test
Secondary Thrombocytosis
Platelet count <1,000,000/uL
Pathophysiology: surger, IDA, recovery from thrombocytopenia
Lab: Platelet count, Diagnosis of underlying condition
Discuss the function of thromboxane A2 and prostacyclin in the regulation of the kinetics of platelets
Arachidonic acid transforms into thromboxane A2 or prostacyclin depending on physiologic need
Thromboxane A2 is an aggregating agent and constricts the blood vessels
Prostacyclin is an anti-aggregating agent a strong platelet inhibitor and vasodilator
