Haemostasis Flashcards
1
Q
What are the 3 main important functions of haemostasis?
A
- Trigger coaguation when needed
- Prevent thrombosis (excessive or generalised blood clot)
- Trigger fibrinolysis when needed
2
Q
What 3 things does the body respond with following trauma?
A
- Contraction of blood vessels (vasoconstriction)
- Formation of an unstable platelet plug at the site of the vessel wall damage (primary haemostasis)
- Formation of a stable fibrin clot (secondary haemostasis/coagulation)
3
Q
What are platelets? What is their circulating lifespan?
A
- Platelets are discoid, non-nucleated, granule-containing cells that are derived from myeloid stem cells.
- Platelets are formed in the bone marrow by the fragmentation of megakaryocyte cytoplasm.
- They have a circulating lifespan of around 10 days.
4
Q
Describe the process of platelet adhesion and what happens when this is achieved?
A
- Injury to the vessel wall -> platelets stick to the damaged endothelium directly to collagen using GPIa receptor or indirectly via von Willebrand factor (VWF), with GPIb receptor.
- Adhesion to wall -> platelets activated -> change shape from disc to more rounded form with spicules -> encourages platelet-platelet interaction.
- Activation -> release of the contents of platelet storage granules.
- Important contents = ADP, fibrinogen and von Willebrand factor.
- Platelet activation also causes a conformational change in the GPIIb/IIIa receptor (known as ‘inside-out’ or ‘flip-flopping’) to provide binding sites for fibrinogen.
- Fibrinogen binding to GPIIb/IIIa causes ‘outside-in’ signalling which further activates the platelets.
5
Q
What are the two main types of ultrastructurally-identifiable granules in platelets?
A
α-granules and dense granules
6
Q
How do platelets and endothelial cells use arachidonic acid?
A
- arachidonic acid is derived from the cell wall.
- prostaglandins are effective vasoconstrictors
- prostacyclins are effective vasodilators.
- cyclo-oxygenase is a common enzyme that allows the two types of cells to use arachidonic acid.
7
Q
How is platelet aggregation triggered?
A
- The granular release of ADP and generation of thromboxane A2 result in further platelet recruitment activation and aggregation. They do this by binding respectively to the P2Y12 and thromboxane A2 receptor.
- Platelet activation also causes a conformational change in the GPIIb/IIIa receptor (known as ‘inside-out’ or ‘flip-flopping’) to provide binding sites for fibrinogen.
- Fibrinogen binding to GPIIb/IIIa causes ‘outside-in’ signalling which further activates the platelets. Fibrinogen has a key role in linking platelets together to form the platelet plug.
8
Q
How is inappropriate platelet aggregation prevented?
A
The aggregation triggered by fibronogen binding to activated platelets is normally counterbalanced by the active flow of blood and the release of prostacyclin (PGI2) from endothelial cells; prostacyclin is a powerful vasodilator and suppresses platelet activation, thus preventing inappropriate platelet aggregation.
9
Q
Summary of platelet adhesion, release and aggregation
A
10
Q
What are two antiplatelet drugs and how do they work?
A
- Antiplatelet drugs are widely used for the prevention and treatment of cardiovascular and cerebrovascular disease.
- Aspirin inhibits the production of thromboxane A2 by irreversibly blocking the action of cyclo-oxygenase (COX), resulting in a reduction in platelet aggregation. Although prostacyclin production is also inhibited by cyclo-oxygenase, endothelial cells can synthesise more COX whereas the non-nuclear platelet cannot. The effect of a single dose of aspirin therefore persists for around 7 days, until most of the platelets present at the time of aspirin ingestion have been replaced by new platelets.
- Clopidogrel works by irreversibly blocking the ADP receptor (P2Y12) on the platelet cell membrane. Therefore the effect of clopidogrel ingestion also lasts for 7 days until new platelets have been produced.
11
Q
What is Von Willebrand Factor (VWF)
A
- Von Willebrand factor (VWF) is a glycoprotein that is synthesised by endothelial cells and megakaryocytes and circulates in plasma as multimers of different sizes.
- VWF mediates the adhesion of platelets to sites of injury and promotes platelet-platelet aggregation.
- In addition to its adhesive properties VWF is a specific carrier for factor VIII (FVIII).
12
Q
Where are most clotting factors synthesised? What are the exceptions to this?
A
- Most clotting factors are synthesised in the liver.
- The exceptions to this are factor VIII and VWF, which are made by endothelial cells. VWF is also made in megakaryocytes and incorporated into platelet granules.
13
Q
What clotting factors need vitamin K and why?
A
- Factors II (prothrombin), VII, IX and X are dependent on Vitamin K for carboxylation of their glutamic acid residues.
14
Q
What needs to be done to inactive forms of clotting factors to generate active forms?
A
- Each step of blood coagulation is characterised by the conversion of an inactive zymogen (proenzyme) into an active clotting factor by the splitting of one or more peptide bonds and exposure of the active enzyme site
15
Q
What clotting factors are co-factors?
A
Factors V and VIII
16
Q
What site are the clotting factors believed to work at and what helps them to bind there?
A
- Many clotting factors are believed to work on the exposed phospholipid surface of platelets, which helps to localise and accelerate these reactions. Calcium ions play an important role in the binding of activated clotting factors to the phospholipid surfaces of platelets.
23
Q
Diagram of 3 phases of coagulation
A
26
Q
How do protein C, protein S and antithrombin inhibit coagulation?
A
- Thrombin binds to thrombomodulin on the endothelial cell surface leading to activation of protein C to activated protein C (APC). APC inactivates factors Va and VIIIa in the presence of a co-factor protein S.
- Thrombin and factor Xa are inactivated by the circulating inhibitor antithrombin. The action of antithrombin is markedly potentiated by heparin: this occurs physiologically by the binding of antithrombin to endothelial cell-associated heparins.
35
Q
How does Tranexamic acid work and what is it used for?
A
- Tranexamic acid is a synthetic derivative of the amino acid lysine that works by binding to plasminogen (see figure below).
- In doing so it prevents plasminogen from binding to the lysine residues of fibrin (competitive inhibition).
- This prevents the activation of plasminogen to plasmin, which would otherwise result in fibrinolysis
- Tranexamic acid is used widely to treat bleeding in trauma and surgical patients as well as in patients with inherited bleeding disorders.
37
Q
How did the intrinsic/extrinsic model work? And what discoveries lead to the acceptance of the current cellular based model?
A
- ‘Intrinsic’ refers to a system in which all components are in the plasma (factors XII, XI, IX, X and co-factors VIII and V)
- While the ‘extrinsic’ system comprises TF and factors VII, X, and co-factor V.
- It was previously believed that the extrinsic and intrinsic pathways ran in parallel, with initiation of the intrinsic pathway resulting from contact activation of factor XII.
- Through a greater understanding of factor XI and the recognition that people with inherited deficiencies of factor XII do not have bleeding problems, it became clear that the intrinsic-extrinsic model did not represent the physiological pathway of coagulation.
38
Q
What pathway’s integrity does Prothrombin Time (PT) measure and how does it work?
A
- Measures the integrity of the ‘extrinsic’ pathway
- Blood is collected into a bottle containing sodium citrate (usually blue-topped as in the picture), which chelates calcium thus preventing the blood from clotting in the bottle
- The sample is spun to produce platelet-poor plasma
- A source of TF and phospholipid is added to the citrated plasma sample, together with calcium to start the reaction; the length of time taken for the mixture to clot is recorded.
- The PT may be prolonged if there is a reduction in the activity of factors VII, X, V, II (prothrombin) or fibrinogen i.e. (‘prothrombin’ is a misnomer)
- Nowadays a recombinant thromboplastin is often used as the source of both TF and phospholipid
- When the PT is used to monitor vitamin K antagonist anticoagulant therapy such as warfarin, the results are expressed as the international normalised ratio (INR). This involves a correction for the different thromboplastin reagents used by different laboratories and means that all laboratories would be expected to obtain the same INR result for a given sample irrespective of the source of thromboplastin.
39
Q
What pathway’s integrity does Activated Partial Thromboplastin Time (APTT) measure and how does it work?
A
- Measures the integrity of the ‘intrinsic’ pathway
- Performed by the contact activation of factor XII by a surface such as glass, or using a contact activator such as silica or kaolin.
- Contact activator, together with phospholipid, is added to the citrated plasma sample followed by calcium; the time taken for this mixture to clot is measured
- Prolongation of the APTT is seen in a variety of situations where there is a reduction in a single or multiple clotting factors; in the the latter there may also be an associated prolonged PT
- An isolated prolonged APTT (i.e. normal PT) is seen in patients with haemophilia A (factor VIII deficiency), haemophilia B (factor IX deficiency) and factor XI deficiency. However this may also be caused by factor XII deficiency which does not result in bleeding. (Note that FXII does not appear in the cell-based model described in ‘Coagulation (secondary haemostasis): formation of the stable fibrin clot’ and is not important for clotting in vivo).
40
Q
Under normal conditions, what determines the fluidity of blood within the blood vessels in terms of equilibrium?
A
- Under normal conditions the confinement of circulating blood to the blood vessels and maintenance of blood in a fluid state are dependent on maintaining a state of equilibrium between these processes:
41
Q
What happens when the haemostatic balance is lost resulting in bleeding?
A
- Loss of this balance leading to bleeding may be caused by:
- Reduction in platelet number or function (primary haemostasis –platelet plug)
- Reduction in coagulation factor(s) (secondary haemostasis – fibrin clot)
- Increased fibrinolysis
42
Q
What happens when the haemostatic balance is lost resulting in thrombosis?
A
- Thrombosis is the term used to describe the formation of a blood clot within an intact blood vessel.
- This usually results in obstruction of the blood flow with serious and possibly fatal consequences.
- The German physician, Rudolf Virchow recognised that there were three contributory factors to pathological clotting or thrombosis. This is known as ‘Virchow’s triad’:
- Blood: dominant in venous thrombosis
- Vessel wall: dominant in arterial thrombosis
- Blood flow: complex, contributes to both arterial and venous thrombosis
- Changes in blood constituents become more important when we come to consider venous thrombosis:
- Changes in blood that increase the risk of venous thrombosis include:
- a) Reduced levels of anticoagulant proteins. These usually have a genetic basis, e.g. inherited antithrombin deficiency, an example of an inherited thrombophilia
- b) Reduced fibrinolytic activity. An example of this is pregnancy where there is inhibition of plasminogen activation through the production of a specific inhibitor by the placenta (PAI-2)
- Increased levels of clotting factors or platelets. For example:
- Levels of factor VIII increase during pregnancy
- The activity of factor V is increased by a single point mutation in the factor V gene, known as factor V Leiden. Factor V Leiden makes factor V more resistant to inactivation by protein C. Around 7% of the population are carriers (heterozygotes) for Factor V Leiden, making it the most common of the inherited thrombophilias.
- Platelets are increased in number in some myeloproliferative disorders, where the bone marrow output is increased.
