haemostasis Flashcards
What does blood do under normal conditions?
flows within the vascular system, transporting oxygen, nutrients and hormonal info around the body and removing metabolic waste
What are the confinement of circulating blood and maintenance of blood in fluid state dependent on the balance of?
- fibrinolytic factors, anticoagulant proteins
- coagulation factors, platelets
Why is balance important for normal haemostasis/what does the balance allow?
- allow the stimulation of blood clotting processes following injury, in which blood changes from its liquid state = coagulation
- limit the extent of the response to the area of injury to prevent excessive or generalised blood clotting = thrombosis
- start the process that eventually leads to the breakdown of the clot as part of the process of healing = fibrinolysis
What does haemostasis describe?
the âhalting of bloodâ following trauma to blood vessels and results from three intertwined proceses
Which three intertwined processes does haemostasis result from?
- 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)
Why is it important to understand these haemostatic mechanisms/in order to what? (5)
- diagnose and treat bleeding disorders
- identify risk factors for thrombosis
- treat thrombotic disorders
- monitor the drugs that ar used to treat bleeding and thrombotic disorders
- control bleeding in individuals who do not have an underlying bleeding disorder
Describe an overview of haemostasis.
(response to injury) vessel constriction -> formation of an unstable platelet plug (platelet adhesion&aggregation) -> stabilisation of the plug with fibrin (blood coagulation) -> dissolution of clot and vessel repair (fibrinolysis)
What is described as primary haemostasis?
formation of an unstable platelet plug (platelet adhesion & platelet aggregation)
What is described as secondary haemostasis?
stabilisation of the plug with fibrin (blood coagulation)
What are platelets?
discoid, non-nucleated, granule-containing cells that are derived from myeloid stem cekls
Where are platelets formed?
in the bone marrow by the fragmentation of megakaryocyte cytoplasm
What is the circulating lifespan of platelets?
around 10 days
What is important for plateletsâ interactions?
glycoproteins (plasma membrane contains glycoproteins (GPs))
What do the platelets do following injury to the vessel wall?
platelets stick to the damaged endothelium, either directly to collagen via the platelet GPIa receptor or indirectly via von Willebrand factor (VWF), which binds to the platelet GPIb receptor
What are the two ways platelets can stick to the damaged endothelium?
- directly to collagen via the platelet GPIa receptor
- indirectly via von Willebrand factor (VWF), which binds to the platelet GPIb receptor
What does the adhesion of platelets lead to for the platelets?
- causes them to become activated
- changes their shape from a disc to a more rounded form with spicules to encourage platelet-platelet interaction
- release of the contents of their storage granules
What are the two main types of ultrastructurally-identifiable granules in platelets?
α-granules and dense granules
How does the platelet release granules?
platelet membrane is invaginated to form a surface-connected cannalicular system through which the platelet granules are released
What are important components of the contents of platelet granules?
ADP, fibrinogen, von Willebrand factor
What are platelets stimulated to produce?
prostaglandin thromboxane A2 from arachidonic acid that is derived from the cell membrane
What are the roles of thromboxane A2?
- platelet aggregation
- vasoconstrictor
- especially important during tissue injury and inflammation
What effects do platelet granular release of ADP and generation of thromboxane A2 have?
positive feedback effects resulting in further platelet recruitment activation and aggregation
How does the granular release of ADP and generation of thromboxane A2 result in further platelet recruitment activation and aggregation?
by binding respectively to the P2Y12 and thromboxane A2 receptor
What does platelet activation cause?
a conformational change in the GPIIb/IIIa receptor (known as âinside-outâ or âflip-floppingâ) to provide binding sites for fibrinogen
What does fibrinogen binding to GPIIb/IIIa cause?
âoutside-inâ signalling which further activates the platelets
What does fibrinogen have a key role in?
linking platelets together to form a platelet plug
What are the effects of fibrinogen counterbalanced by?
- active flow of blood
- release of prostacyclin (PGI2) from endothelial cells
What is prostacyclin (PGI2)?
- powerful vasodilator
- suppresses platelet activation
thus preventing inappropriate platelet aggregation
What are antiplatelet drugs widely used for?
the prevention and treatment of cardiovascular and cerebrovascular disease
What are the two most commonly used antiplatelet drugs?
aspirin and clopidogrel
What does aspirin do?
inhibits the production of thromboxane A2 by irreversibly blocking the action of cyclo-oxygenase (COX)
How does aspirin inhibit the production of thromboxane A2?
by irreversibly blocking the action of cyclo-oxygenase (COX), resulting in a reduction in platelet aggregation
What is also inhibited by cyclo-oxygenase?
prostacyclin production but, endothelial cells can synthesis more COX whereas the non-nuclear platelet cannot
How long do the effects of a single dose of aspirin last for?
around 7 days, until most of the platelets present at the time of aspirin ingestion have been replaced by new platelets
How does clopidogrel work?
by irreversibly blocking the ADP receptor (P2Y12) on the platelet cell membrane
How long do the effects of clopidogrel ingestion last?
7 days until new platelets have been produced
What is Von Willebrand factor (VWF)?
a glycoprotein that is synthesised by endothelial cells and megakaryocytes and circulates in plasma as multimers of different sizes
What do van Willebrand factors do?
mediates the adhesion of platelets to sites of injury and promotes platelet-platelet aggregation
What is another property of VWF in addition to its adhesive properties?
VWF is a specific carrier for factor VIII (FVIII)
What is coagulation?
- secondary haemostasis
- formation of the stable fibrin clot
When is primary platelet plug sufficient?
for small vessel injuries
Primary platelet plugs would fall apart for large vessels. What stabilises the platelet plug?
fibrin formation
What do blood coagulation pathways centre on?
on the generation of thrombin, which cleaves fibrinogen to generate a fibrin clot that stabilises the platelet plug at sites of vascular injury
Where are most clotting factors synthesised?
in the liver
What are the exceptions for clotting factors synthesised in the liver?
factor VIII and VWF
What are factor VIII and VWF made by?
endothelial cells
Where else are VWF made in?
megakaryocytes and incorporated into platelet granules
Which clotting factors are dependent on Vitamin K?
factors II (prothrombin), VII, IX, X
What are factors II (prothrombin), VII, IX and X dependent on Vitamin K for?
carboxylation of their glutamic acid residues, which is essential for the function of these clotting factors
What does the process of blood coagulation involve?
a number of steps that are activated in sequence
What is each step of blood coagulation characterised by?
by the conversion of an inactive zymogen (proenzyme) into an active clotting factor
How is inactive zymogen (proenzyme) converted into an active clotting factor?
by the splitting of one or more peptide bonds and exposure of the active enzyme site
What are factors V and VIII?
co-factors
Where are many clotting factors believed to work on?
on the exposed phospholipid surface of platelets, which helps to localise and accelerate these reactions
What is the role of calcium ions in clotting factors?
calcium ions play an important role in the binding of activated clotting factors to the phospholipid surfaces of platelets
What is the trigger to initiate coagulation at the site of injury?
the tissue factor (TF) exposed on the surface of endothelial cells and leukocytes and on most extravascular cells in an area of tissue damage
Where is TF mainly located?
at sites that are not usually exposed to the blood under normal physiological conditions
What is a result of TF being mainly located at sites that are not usually exposed to the blood under normal physiological conditions?
blood only encounters TF at sites of vascular injury
What does the binding of TF to factor VIIa lead to?
the activation of factors IX to IXa and X to Xa
What does the activation of factors IX to IXa and X to Xa lead to?
the activation of prothrombin (factor II) to generate a small initial amount of thrombin (factor IIa)
What is the phase of activation of prothrombin (factor II) to generate a small initial amount of thrombin (factor IIa) known as?
Initiation phase
What does the small initial amount of thrombin generated in the Initiation phase do?
mediates the activation of the co-factors V and VIII, the zymogen factor XI and platelets (Amplification phase)
What is the Amplification phase?
When the small amount of thrombin (generated in the Initiation phase) mediates the activation of the co-factors V and VIII, the zymogen factor XI and platelets
What does factor XI convert?
factor XI converts more factor IX to IXa
What does IXa do in concert with factor VIIIa?
amplifies the conversion of factor X and Xa
What happens after the amplification of the conversion of factor X to Xa?
a rapid burst in thrombin generation
What phase is the rapid burst in thrombin generation known as?
Propagation phase
What does the propagation phase (rapid burst in thrombin generation) do?
cleaves the circulating fibrinogen (soluble) to form the insoluble fibrin clot
What do the action of inhibitory mechanisms ensure?
that coagulation is confined to the site of injury and prevent spontaneous activation of coagulation
What are the names of the most important inhibitory mechanisms?
protein C, protein S, antithrombin
What is the role of thrombin?
binds to thrombomodulin on the endothelial cell surface leading to activation of protein C to activated protein C (APC).
What does APC do?
inactivates factors Va and VIIIa in the presence of a co-factor protein S
What are thrombin and factor Xa inactivated by?
by the circulating inhibitor antithrombin
What is the action of antithrombin markedly potentiated by?
heparin: this occurs physiologically by the binding of antithrombin to endothelial cell-associated heparins
What are the main anticoagulant drugs?
heparin, warfarin and the direct oral anticoagulants (DOACs)
What are the main anticoagulant drugs widely used in?
the prevention and treatment of thrombosis
What is heparin
- an anticoagulant drug
- a mixture of glycosaminylglycan chains extracted from porcine mucosa
How does heparin work?
it works indirectly by potentiating the action of antithrombin leasing to the inactivation of factors Xa and IIa (thrombin)
What does inactivation of thrombin require?
requires longer chains of heparin chains, which are able to wrap around both the antithrombin and thrombin
How is heparin administered?
intravenously or by subcutaneous injection
What is warfarin?
- an anticoagulant drug
- derived from coumarin
- vitamin K antagonist
How does warfarin work?
by interfering with protein carboxylation
- it therefore reduces synthesis of functional factors I, VII, IX and X by the liver
How is warfarin given and monitored?
- given as an oral tablet
- its anticoagulant effect needs to be monitored by regular blood testing
How many days does it take for warfarin to take effect and why?
- several days to take effect
- because it reduces synthesis of coagulation factors rather than inhibiting existing factor molecules
What are direct oral anticoagulants (DOACs)?
orally available drugs that directly inhibit either thrombin for factor Xa (i.e., without the involvement of antithrombin)
Do direct oral anticoagulants (DOACs) need monitoring?
they do not usually require monitoring
What mechanism does the body have after haemostasis has been achieved?
a mechanism to break down (lyse) clots
What is the bodyâs mechanism to break down (lyse) clots called?
fibrinolytic system
What is the principal fibrinolytic enzyme?
plasmin
In what form does the principal fibrinolytic enzyme plasmin circulate?
in its inactive zymogen form plasminigen
What is the activation of plasmin mediated by?
tissue plasminogen activator (t-PA)
When does t-PA not activate plasminogen?
t-PA does nit activate plasminogen until these (????) are both brought together by binding to lysine residues on fibrin
What does the breakdown of fibrin lead to?
the generation of fibrin-degradation produces (FDPs)
Is plasmin specific?
plasmin is not specific for fibrin and can also break down other protein components of plasma, including fibrinogen and the clotting factors Va and VIIIa
What is plasmin inhibited by?
antiplasmin, which circulates in the blood
What else is plasmin inhibited by?
plasmin is one of the many proteases inhibited by alpha 2 macroglobulin, the broadly active alpha-2-globulin (from Peaseâs plasma lecture)
What is an example of a thrombolytic agent?
recombinant t-PA
How does recombinant t-PA (a thrombolytic agent) work?
by generating plasmin to lyse clots
How and when is recombinant t-PA (a thrombolytic agent) administered?
intravenously to selected patients presenting with ischaemic stroke
Describe the administration of t-PA and the reason for this.
the benefit is time-dependent and so t-PA needs to be given to eligible patients as quickly as possible, preferably within one hour of the onset of symptoms
What is a risk associated with t-PA use?
high risk of bleeding associated
When can thrombolytic therapy be given, other than for ischaemic stroke?
can also be given to patients with life threatening pulmonary emboli
- was previously used in patients with myocardial infarction, although this has largely been replaced with angioplasty and the insertion of stents to open the diseased coronary vessels
What is tranexamic acid?
a synthetic derivative of the amino acid lysine
How does tranexamic acid work?
by binding to plasminogen
What does tranexamic acid do by binding to plasminogen?
prevents plasminogen from binding to the lysine residues of fibrin- known as competitive inhibition
What does the competitive inhibition of tranexamic acid prevent?
the activation of plasminogen to plasmin, which would otherwise result in fibrinolysis
What is tranexamic acid widely used for?
to treat bleeding in trauma and surgical patients, as well as in patients with inherited bleeding disorders
tests of coagulation
What does Prothrombin time (PT) measure?
the integrity of the âextrinsicâ pathway
What are two tests of coagulation?
- prothrombin time (PT)
- activated partial thromboplastin time (APTT)
What is the first step of prothrombin time (PT)?
blood is collected into a bottle containing sodium citrate (usually blue-topped), which chelates calcium thus preventing the blood from clotting in the bottle
What is the second step of prothrombin time (PT)?
the sample is spun to produce platelet-poor plasma
What is the third step of prothrombin time (PT)?
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 misture to clot is recorded
Why might the PT be prolonged?
if there is a reduction in the activity of factors VII, X, V, II (prothrombin) or fibrinogen i.e., (âprothrombinâ is a misnomer)
What is often used nowadays as the source of both TF and phospholipid?
a recombinant thromboplastin
What is PT used to monitor?
vitamin K antagonist anticoagulant therapy such as warfarin
How are the results of monitoring vitamin K antagonist anticoagulant therapy using PT expressed?
as the international normalised ratio (INR)
What does the international normalised ratio involve?
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
What does the activated partial thromboplastin time (APTT) measure?
the integrity of the âintrinsicâ pathway
How is activated partial thromboplastin time (APTT) performed?
performed by the contact activation of factor XII by a surface such as glass, or using a contact activator such as silica or kaolin
How do you find out the activated partial thromboplastin time (APTT)?
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
When is prolongation of the APTT seen?
it is seen in a variety of situations where there is a reduction in a single or multiple clotting factors; in the latter there may also be an associated prolonged PT
When is an isolated prolonged APTT seen.
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
