7 - Haemostasis Flashcards
State the steps involved in haemostatic plug formation from the time of injury.
- Vessel constriction
- Formation of an unstable platelet plug (platelet adhesion and aggregation)
- Stabilisation of plug with fibrin (blood coagulation)
- Dissolution of clot and vessel repair (fibrinolysis)
What is the very first initial response to vessel wall injury?
vessels constriction - this is a local contractile response (important for smaller vessels in particular)
What are the 2 important steps of formation of an unstable platelet plug?
platelet adhesion to the damaged blood vessel
platelet aggregation
What must happen after the formation of an unstable platelet plug to snore that it doesn’t get broken down?
blood coagulation and formation of a fibrin network to stabilise it
What are the two main functions of the endothelium (in reference to platelet plug formation)?
- maintains the barrier between the blood and procoagulant sub endothelial structures
- synthesis of prostaglandins, thrombomodulin, vWF and plasminogen activators
What component found underneath the endothelium is involved in triggering the coagulation cascade?
- Procoagulant subendothelial structures e.g. collagen
- Tissue factor
Where is tissue factor found in the body?
expressed on the surface of cells underlying blood vessels
but it is NOT normally expressed within the circulation itself
State some important factors produced by endothelial cells.
Prostacyclin
Thrombomodulin
Von Willebrand Factor
Plasminogen Activator
Briefly, why is von Willebrand factor important?
important in the initial platelet interaction with the damaged area
Briefly, why is plasminogen factor important?
involved in the breakdown of clots
What process during maturation of the megakaryocytes is important for the formation of platelets?
Granulation
What do the dense granules in platelets contain that is important for platelet function?
ADP
What do alpha granules in the platelets contain?
vWF
Factor V
State the two ways in which platelets can bind to collagen. Name the receptors involved.
- It can bind via vWF to collagen (via the GlpIb receptor)
- It can bind directly to the collagen (via the GlpIa receptor)
What happens following the passive adhesion of platelets to the (sub-endothelial) collagen and engagement of receptors?
- The receptors signal inside the cell to release ADP from the storage granules and to synthesise thromboxane
- These bind to receptors on the surface of the platelets and activate them
- Once activated, GlpIIb/IIIa receptors become available, which can bind to fibrinogen and allows the platelets to aggregate
Which receptors on the platelets become available following activation of the platelets and what do they bind to?
GlpIIb/IIIa
These bind to fibrinogen
What else can activate platelets?
Thrombin
What does platelet activation lead to?
prostaglandin metabolism
Which enzyme converts phospholipids to arachidonic acid?
Where does this take place
Phospholipase
in the activated endothelial cells
What does COX convert arachidonic acid to?
Endoperoxides
Describe what happens to endoperoxides in platelets and in the endothelial cells.
Platelets – thromboxane synthetase converts endoperoxides to thromboxane (potent inducer of platelet aggregation)
Endothelial Cells – prostacyclin synthetase converts endoperoxides to prostacyclin (important regulator of haemostasis)
Describe what happens to endoperoxides in platelets and in the endothelial cells.
Platelets – thromboxane synthetase converts endoperoxides to thromboxane (potent inducer of platelet aggregation)
Endothelial Cells – prostacyclin synthetase converts endoperoxides to prostacyclin (important regulator of haemostasis)
What effect does aspirin have on the prostaglandin metabolism pathway?
Aspirin is a COX1 inhibitor
State some important drug targets in platelet aggregation.
COX
GlpIIb/IIIa
ADP Receptor
State two ADP receptor antagonists.
Clopidogrel
Prasugrel
State three GlpIIb/IIIa antagonists.
Abciximab
Tirofiban
Eptifibatide
What is the most important test for monitoring platelets and their function?
Platelet count
What is the normal range for platelet count?
150-400 x 10^9/L
What is a common cause of spontaneous bleeding?
What are the symptoms?
Autoimmune thrombocytopenia (autoimmune antibodies clear platelets from the circulation) This results in purpura, multiple bruises and ecchymoses
Why do you get thrombocytopenia in leukaemia?
Leukaemic cells populate the bone marrow so it crowds out the megakaryocytes so the platelets aren’t produced in sufficient numbers
Describe the platelet aggregation test. What is it used to diagnose?
The platelets are stimulated with ADP/thromboxane/collagen to study their function
This is used to diagnose platelet disease e.g. von Willebrand disease
What does the platelet count test for?
thrombocytopenia
Where is von Willebrand factor produced?
Endothelial cells and a little bit by megakaryocytes
What factors do megakaryocytes produce?
Factor V
Von Willebrand Factor
Where are most of the coagulation factors produced?
in the liver
What factor is prothrombin?
factor 2
What factor does thrombin activate?
factor 13 (to its activate form factor 13a)
What is the Amin factor involved with tissue factor?
factor 7a
How is fibrin stabilised?
by covalent cross linking of fibrin by factor 13a
Tissue factor activates the clotting cascade by converting 9 to 9a and by converting 10 to 10a. What difference does this make?
9 to 9a – slower but produces more thrombin
10 to 10a – faster
State two accelerating factors. What are they activated by?
Factor VIII
Factor V
They are activated by trace amounts of thrombin
Which factors are activated on the surface of the platelet? Describe how this works.
- 10 to 10a
For 9a to activate 10 it needs to come in close proximity with 10. They both bind to the surface of the platelet mediated by calcium ions, and factor VIIIa bring the two close together so that 9a can proteolytically cleave 10 to 10a - 2 to 2a (prothrombin to thrombin)
Factor Va does the same with 10a and 2 (prothrombin)
Which factors are affected by warfarin?
2, 7, 9, 10
What is common to all of the factors that are affected by warfarin and what is the significance of this common feature?
They have a cluster of glutamic acid
The glutamic acid is recognised by a liver enzyme and is modified in the presence of vitamin K to Gamma-carboxyglutamic acid
Once this extra carboxyl group is added, calcium can facilitate the binding of gamma carboxyglutamic acid to the activated platelet membrane phospholipid
Name the enzyme that warfarin inhibits and explain how this leads to inhibition of the post-translational modification of these factors (2,7,9,10)?
Warfarin inhibits vitamin K epoxide reductase thus inhibiting the gamma carboxylation of factors 2, 7, 9 and 10
What effect does heparin have on anti-thrombin?
Heparin potentiates the action of anti-thrombin
In what situation is heparin used?
Heparin is used for immediate anticoagulation in venous thrombosis and pulmonary embolism
Describe how anti-thrombin inhibits the clotting factors.
Dircet inhibition of factor 10a and thrombin
Anti-thrombin has a reactive loop that irreversibly inhibits the active site on the clotting factors
So anti-thrombin acts as a scavenger in stopping inappropriate action of clotting factors
How does heparin potentiate the action of anti-thrombin and why is the chain length important?
Once bound to anti-thrombin, heparin changes the position of the reactive loop and makes the inhibition occur faster
- for factor 10a, a relatively SHORT chain of heparin is sufficient (low molecular weight heparin)
- for thrombin, you require a LARGE chain (standard/unfractionated heparin)
NOTE: standard/unfractionated heparin inhibits either Factor 10a or thrombin
What are the two forms of heparin? Which factor do they inhibit the action of?
- low molecular weight heparin - inhibits the action of factor 10a
- standard/unfractionated heparin - inhibits the action of both factor 10a and thrombin
State three laboratory tests for blood coagulation.
Activated Partial Thromboplastin Time (APTT)
Prothrombin Time (PT)
Thrombin Clotting Time (TCT)
What do each of the laboratory tests represent?
Activated Partial Thromboplastin Time (APTT
Prothrombin Time (PT)
Thrombin Clotting Time (TCT))
APTT – detects abnormalities in the INTRINSIC and COMMON pathways (coagulation is triggered by activation of factor 12)
PT – detects abnormalities in the EXTRINSIC and COMMON pathways (tissue factor is added to trigger the extrinsic pathway)
TCT – shows abnormality in the fibrinogen fibrin conversion (not important any more)
What are the main uses of these laboratory tests?
(Activated Partial Thromboplastin Time (APTT)
Prothrombin Time (PT)
Thrombin Clotting Time (TCT))
APTT and PT are used together for screening causes of bleeding disorders
APTT is used to monitor heparin therapy for thrombosis
PT is used to monitor warfarin treatment
What two proteins assemble on the surface of a clot to allow fibrinolysis to take place? Where are these proteins made?
- Plasminogen
- Tissue Plasminogen Activator (tPA)
Plasminogen is a plasma protein
tPA is produced by endothelial cells
What is produced from the break down of the fibrin clot and how does this level change in DIC?
Fibrin degradation products (FDP)
This is elevated in DIC
What factors are used in therapeutic thrombolysis of myocardial infarction?
tPA and bacterial activator streptokinase
Describe the function of the Protein C anticoagulant pathway.
Thrombin also has a role in anticoagulation
It binds to thrombomodulin on the surface of endothelial cells and by binding to thrombomodulin it activates protein C, which, along with co-factor protein S, INACTIVATES FACTOR Va and FACTOR VIIIa
