Week 5: Primary Hemostasis

Hemostasis Vascular problems

Platelet function Allergy
Platelet anatomy Viral infection
Megakaryocyte Collagen disorders
Platelet kinetics Vitamin C deficiency
Platelet count Ehlers-Danlos
Aggregation studies Render-Osler-Weber
Von Willebrands telangiectasia
Bernard-Soulier Petechiae, ecchymoses
Aspirin (salicylate) Bleeding time
Vascular Injury
Serotonin and thrombaxane A2 (TxA2) for
vasoconstriction
Prostacyclin PGI-2 for arteriole relaxation
to increase blood flow
Exposure of basement membrane and
collagen (negatively charged surface)
Inside of a vessel SEM x 2,500
Process of Hemostasis
Vascular injury
Platelet adhesion and activation
Platelet aggregation (1o hemostatic plug)
Fibrin formation via cascade (2o
hemostasis)
Clot retraction (thrombasthenin)
Fibrinolysis and healing
Role of Platelets
Surveillance for
vascular integrity
Formation of 1o
hemostatic plug
Activation of 2o
hemostasis
Healing
Platelet Formation
Megakaryoblast undergoes endomitosis
Intermiediate stage promegakaryocyte
without granules
Megakaryocyte (2N to 64N) with over
100 diameter
IL3, GM-CSF, thrombopoietin
20% of platelet stored in spleen
Platelet
2 - 4 diameter
Round or oval
Hyalomere - clear peripheral zone
Granulomere - highly stained area with
granules
Platelet Anatomy
Peripheral zone with glycoprotein
receptors
Structural zone with contractile
microtubules (thrombasthenin)
Organelle zone with granules
Membrane with open cananicular and
tubule systems for increased surface area
and rapid release
 Electron micrograph of a platelet x 25,000

Longitudinal peripheral microtubule (brown), endoplasmic reticulum

(blue), mitochondria (green), glycogen (black)
Platelet Function
Adhesion to basement membrane or collagen with
vWF and GP-Ib
Activation
Shape change from discoid to distorted

Exposure of GP-IIb/IIIa and other receptors

TxA2 synthesis (cyclo-oxygenase dependent)

Membrane phospholipid Arachidonic acid

Arachidonic acid TxA2 and prostaglandins
Platelet Function:
Glycoprotein Group or Gene Families
Integrins
Leucine rich glycoprotein family
Selectin family
Quadraspanin family
Immunglobulin supergene family
Integrins
Integral to membrane
Ca++ dependent GPIIb/IIIa most abundant
Cell-cell or cell-substrata interaction
Receptor to Fib, vWF, vitronectin,
fibronectin
Leucine Rich GP Family
Adhesion to subendothelial collagen (COL)
mediated by vWF by transmembrane
complex GPIb/IX
Stabilizes PLT membrane by interaction
with cytoskeleton
Selectin Family
GMP-140 mediates adhesion of neutrophils
and monocytes to PLT
Quadraspanin Family
Plasma membrane protein p24/CD9
interacts with GPIIb/IIIa, modulating
adhesion molecules
Leads to Ca++ increase and subsequent PLT
activation and aggregation
Immunoglobulin Supergene Family
Functional role unclear
Has role in cellular interactions
Platelet Activation
1o aggregation with agonists: ADP, epinephrin,
serotonin, PF4 (anti-platelet)
Release or secretion facilitated by TxA2
o
Dense body for 2 aggregation and
vasoconstriction (ADP, Ca++, serotonin)
-granule for heparin neutralization and clot

Platelet derived growth factor (PDGF) for

healing
Retraction
Signs and Symptoms of 1o
Hemostasis Problems
Ecchymoses
Petechiae
Mucus membrane bleeding
Hematoma
Prolonged bleeding after minor surgery
Hereditary Vascular Problems
Hereditary (spider) telangiectasis (Osler-
Rendu-Weber): dilated superficial
capillaries
Ehlers-Danlos: collagen disorder
Marfan syndrome: connective tissue
Osteogenesis imperfecta
Acquired Vascular Problems
Senile purpura (Batemans): altered
connective tissue support
Cushing syndrome: metabolic
Scurvy: abnormal collagen
Allergy: vascular inflammation
Viral infection
Bleeding Time
For vascular and platelet functions
Duke (1910) on earlobes
Ivy (1941) on arm with 1mm x 3mm
incision
Mielke (1969) with 1mm x 10mm template
1980s: disposable devices (e.g., Simplate,
Surgicutt)
Bleeding Time
 Quantitative Platelet Disorders
Thrombocytopenia
<100,000/ml BT prolonged
10,000 Bleeding in trauma or OR
<10,000 Spontaneous, CNS bleeding
Thrombocytopenia due to destruction
ITP (acute in children, chronic in young women)
with anti-glycoprotein
Drug reaction

Heparin induced thrombocytopenia

DIC and TTP

About Thrombotic Thrombocytopeneic
Purpura (TTP)
Disorder of systemic platelet aggregation in
microvasculature
Stimulus: unusually large vWf
In children: likely to be deficiency in vWf
metalloproteinase to break down vWf
In adults: vWf metalloproteinase inhibited by
autoantibodies
Low PLT count, intravascular hemolysis, RBC
fragmentation, high LDH
Quantitative Platelet Disorders
Thrombocytopenia due to decreased production
Aplastic anemia (e.g., Fanconis)

Fibrosis

Acute leukemia

Megaloblastic anemia

Hereditary (e.g., May-Hegglin, Wiscott-Aldrich,

Bernard-Soulier)
Splenic sequestration
HELLP syndrome (hemolysis, elevated liver enzyme,
low PLT) in pre-eclampsia
Dilution (massive transfusion)
Platelet Satellitosis in EDTA
Quantitative Platelet Disorders
Thrombocytosis
Primary with dysfunctions (e.g., CML,
ET)
Post splenectomy: also see HJ, etc.

Hemolytic anemia

Acute hemorrhage and surgery

Pseudo Thrombocytosis
Red cell abnormalities
HJ bodies

Clumped Pappenheimer bodies

nRBC

Malaria

Microspherocytes and schistocytes

White cell abnormalities

Unlysed WBC

WBC fragments and necrobiotic cells

Platelet Count
Rees-Ecker with brilliant cresyl blue
Brecker-Cronkite with ammonium oxalate
Unopette: similar to BC
Electronic counters
Qualitative Platelet Disorders
Berhard-Soulier: GP-Ib deficiency, adhesion
problem
Von Willebrands: vWF deficiency, adhesion
problem
Glanzmanns thrombasthenia: GP-IIb/IIIa
deficiency, aggregation problem -- cannot bind
vWF and Fib
Storage pool disease: dense body defect, secretion
problem
Qualitative Platelet Problems
Aspirin: inhibits cyclo-oxygenase (COX),
secretion problem, no TxA2
Plavix (Clopidogrel) inhibits ADP receptor
Other medications affect GPIIa/IIIb
interaction with Fib
Uremia, secretion problem
Gray platelet syndrome: -granule defect
Hypofibrinogenemia
Aggregation Studies
ADP
reversible 1o wave

if ADP is released, then 2o wave

abnormal with aggregation and release problems

Epinephrin
similar to ADP

Collagen
direct release so only one wave of aggregation

Ristocetin
antibiotic

aggregation only with vWF and GP-Ib

Platelet Aggregometry
 Platelet

Aggregation