Transcript Platelet Function in Cardiothoracic Surgery

Platelet Function in
Cardiothoracic Surgery
Mike Poullis
Overview
• Basic science
– What are they?
– How do they work?
• Methods of assessing platelet function
– Full blood count
– Microaggregation
– Macroaggregation
– Thromboelastography (TEG)
– Platelet function analyser
– Bleeding time
• Clinical scenarios
– Drugs
– Medical conditions
– Congenital disorders
– Cardiopulmonary bypass
– Aprotinin
– HIT
– On the ward
Basic science
• What are they?
• How do they work?
What Is a Platelet?
Blood Film
Ultra Structure
Coagulation System
• Platelets
• Soluble Factors
• Red Cells
• Soluble factors interact with platelets to
form a mesh that RBCs stick to
How Do Platelets Work ?
• Afferent pathway
• Inside platelet pathway
• Efferent pathway
Afferent pathway
Inside
platelet
pathway
Efferent pathway
Platelet Agonists (Afferent)
Agonist
Receptor
Adrenaline
Collagen
ADP
Thrombin
alpha
VLA
ADP
PAR I and IV
The platelet
Thrombin
PAR-1 AP
Adrenaline
ADP Collagen
Aprotinin
PAR-1 receptor
G protein
PMA
PKC
Ca2+ Flux
Platelet
Platelet
aggregation
Microaggregation and
Macroaggregation
Mechanism of G protein receptor
activation by soluble ligand eg ADP
Soluble ligand
(Reversible)
Cell Membrane
G protein
Mechanism of G protein receptor activation
by protease eg Thrombin
Protease eg thrombin
(Irreversible)
Cell Membrane
G protein
(cont)
Tethered ligand
(Irreversible)
Peptide
Cell Membrane
G protein
Activation
PAR activating peptides
PAR activating peptide
(reversible)
Cell Membrane
G protein
Activation
Actions proteolytic inhibitors eg
Aprotinin
Protease eg thrombin
Aprotinin
Cell Membrane
G protein
PAR deactivation
Protease eg thrombin
aa 41/42
Deactivator eg elastase
aa 43/44 & 55/56
(Irreversible)
NH2
Cell Membrane
G protein
Platelet Thrombin desensitisation
1
Neutrophil elastase cleaves PAR-1
Specific cleavage inhibits activation
2
Thrombin fragment deactivation theory
Efferent
Agonist
Receptor
GP IIb/IIIa
GP Ib
Fibrinogen
Von Willebrand Factor
GP IIb/IIIa Cross Linking
Techniques to Understand
• Microaggregation
• Macroaggregation
• Platelet function analysers
• Thrombelastography
• Calcium fluxes
Microaggregation and How to
Count Platelets
No
No
Size
Size
Macroaggregation
Increasing
aggregation
Time
Platelet Function Analyser
Thromboelastography (TEG)
Principles of
Thrombelastography
Readout
What the Numbers/letters Mean
• R: Time from initiation to initial fibrin formation
• k: Time of clot formation until amplitude of 20 mm
• Alpha angle: Acceleration (kinetics) of fibrin build up
and cross-linking
• MA - Maximum amplitude strength of clot (number
function platelets, fibrin)
• MA60: The rate of amplitude reduction 60 min. after
MA (stability) of the clot
Tips and Tricks
• Heparinase
• Adding c7E3 Fab (ReoPro) to the TEG sample
will eliminate platelet function from the
thromboelastogram.
• Antifibrinolytic agents such as EpsilonAminocaproic Acid, Tranexamic acid and
Aprotinin
Normal Coagulation Profile
Heparinase
No Heparinase
Heparin Effect
Heparinase
No Heparinase
General Shapes Of TEGS
Calcium Flux Measurements
Calcium Flux Measurements
1 Thrombin
2 Adrenaline
A control
B Aprotinin
Tests of Platelet Function
• Full blood count
• Whole Blood tests
– Microaggregation
– Thrombelastography
• Purified Platelet tests
– Microaggregation
– Macroaggregation
– Platelet function analysers
– Calcium flux
• Skin bleeding time
Advantages of Techniques
• Full blood count
– Quick, easy, reproducible, understandable
• Whole Blood tests
– Microaggregation, Thrombelastography
Easy
MAJOR ADVANTAGE IS NO SAMPLE PREPERATION
• Purified Platelet tests
– Microaggregation
– Macroaggregation
– Platelet function analysers
– Calcium flux
• Skin bleeding time
– Whole body answer
Easy
PRECISE DEFECT
PRECISE DEFECT
PRECISE DEFECT
Limitations of Techniques
• Full blood count
– Number not function
• Whole Blood tests
– Microaggregation
– Thrombelastography
No commercial kit
?sensitivity
• Purified Platelet tests
YOU HAVE TO PREPARE THE PLATELETS
– Microaggregation
No commercial kit
– Macroaggregation
Experienced technician
– Platelet function analysers
No enzymes available
– Calcium flux
Expensive and experience needed
• Skin bleeding time
– Invasive, not specific
Clinical Scenarios
•
•
•
•
•
•
•
Drugs
Medical conditions
Congenital disorders
Cardiopulmonary bypass
Aprotinin
HIT
On the ward
Effect of Drugs on Platelet
Function
•
•
•
•
•
•
Aspirin (Non steroidal anti inflammatory drugs)
Clopidogrel, Ticlopidine
ReoPro, Tirofiban
Prostacyclin
Hirudin
Ancrod
Aspirin (Non steroidal anti inflammatory
drugs)
TxA2
Platelet
PgI2
Endothelium
Clopidogrel / Ticlopidene
Thrombin
PAR-1 AP
Adrenaline
ADP Collagen
Aprotinin
PAR-1 receptor
G protein
PMA
PKC
Ca2+ Flux
Platelet
Platelet aggregation
ReoPro / Tirofiban / Integrilin
Platelet
GP 11b/111a
Platelet
GP 11b/111a
Fibrin
Effect of Drugs on Platelet
Function - Cont
• Prostacyclin
• Hirudin
• Ancrod
Medical Conditions
• Hyperglobulinaemia
– Multiple myeloma and
– Waldestrons macroglobulinaemia
• Renal failure Uraemia
• Liver disease
• Myeloproliferative disorders
– Essential thrombocythaemia
– Polycytheamia
– Myelodysplasias
Congenital Platelet disorders
• All rare
• VonWillibrands (commonest)
• Glanzmann Thrombasthenia (Acquired common)
• Bernard-Soulier
• Platelet storage disorders
Von Willebrands
Platelet
Factor V111
vWF
Endothelium
Stabilisation and adhesion
Glanzmann Thrombasthenia
• Genetic platelet disorder
• Gp IIb/IIIa deficient or dysfunctional
• FACS analysis
Bernard-Soulier
• Large platelets
• Phospholipid not made available
• GP 1b deficiency
Platelet Storage Disorders
• ADP or 5-HT release deficiency
• Defect in dense or alpha granules
How are platelets protected
during CPB?
• Heparin only works on soluble factors!
Thrombin
Intrinsic
Extrinsic VII
HEPARIN
CLOT
Platelet Dysfunction Post CPB
•
Activation decreased numbers and causes
clumping
•
Thrombin receptor desensitisation (Neutrophil
elastase)
•
Thrombin fragment deactivation theory
•
GP Ib depletion
•
Other
Platelets and Aprotinin
• Aprotinin inhibits thrombin induced platelet
aggregation but not adrenaline, collagen,
and ADP induced aggregation
• Aprotinin protects GP1b levels
• Neutrophil enzymes can cleave PAR
receptor and make it functionless
Aprotinin Inhibiting the Effects
of Thrombin
Increasing
aggregation
Time
HIT, and
HITT
HIT, HITT, CPB and Aprotinin
• PF-4 and heparin antibody
– Clinical suspicion
– Immunoassay
– Bioassay
• How do you develop HIT?
–
–
–
–
Tissues and blood cell activation
Heparin exposure
Antibody formation
Antibody has to have a functional Fc
PAF
Drugs to treat HIT
•
•
•
•
•
LMW
Prostacyclin
Ancrod
Aprotinin
Thrombin antagonists Hirudin
• Ancrod - aprotinin interaction is important
Thrombocytopenia in the well
patient on day 5 ready for home
• Blood film for platelet clumping
• Repeat in Citrate not EDTA
• Look at coulter trace
• ? Need extra dose of aspirin or clopidogrel
Blood Film
Microaggregation and Platelet
Clumping
No
No
Size
Size
Interpreting Cardiac Surgery
Papers on Platelet Function
•
•
•
•
Agonist
Technique
What is the likely defect?
Is it relevant?
• Methodology quirk “Correcting count for
tests”
Any Questions?