Lecture 10: Physiological Coagulation Flashcards
What can excess bleeding be due to?
Excessive bleeding due to:
- Lacking one or more coagulation factors
- Lacking or impaired platelets
- Too many inhibitors (i.e. medicines) probably unlikely
Describe the difference between haemostasis and thrombosis
Spectrum between haemostasis and thrombosis
Haemostasis is a normal process where we’re using our clotting proteins and platelets to heal the defects in the blood vessel wall (Small and platelet rich)
Thrombosis is the formation of an abnormal clot and the vessel wall is generally in tact.
- Venous: red cells and fibrin
- Arterial: platelet rich; occlusive
Describe
Step 1: platelet plug (primary haemostasis)
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Primary haemostasis starts following vessel injury. It responds by vasoconstriction, leading to reduced blood flow.
Damage to vessel wall (disrupted endothelium) leads to exposure of collagen in subendothelial tissue. This triggers platelet activation which leads to platelet plug formation. This process occurs within a few seconds of injury and leads to development of an unstable clot.
- Platelets are localised to injury site where they stick to exposed collagen (left figure below). This process is platelet adhesion via circulating von Willebrand factor (VIII-vWF), which fixes platelet receptor Gplb (glycoprotein) to collagen.
- Following adhesion, platelet aggregation occurs.
- It requires a shape change where platelet morphology alters from a disc to a _spiney spher_e.
- Shape change is associated with release action, which involves release of vasoconstricting amines (e.g. serotonin) and adenine nucleotides (e.g. ADP), which cause further platelet activation and initiate platelet aggregation.
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Aggregation is process by which one platelet sticks to another to form platelet plug.
- Aggregation is mediated by fibrinogen (a plasma protein) binding platelets together via integrin αIIbβ3 (also known as glycoprotein IIb IIIa). This receptor is activated during shape change process.
- It requires a shape change where platelet morphology alters from a disc to a _spiney spher_e.
Prostaglandin pathways, particularly in the platelet, are also important in these processes as they convert arachidonic acid in platelet membrane to thromboxane A2 (TXA2), which is a potent vasoconstrictor and aggregator of platelets.
Describe Secondary Haemostasis
(Coagulation pathway)
At the _same time as platelet activatio_n, coagulation pathway is activated.
- When platelet aggregate first occurs, it is unstable. A sequence of enzymatic reactions is initiated that culminates in formation of fibrin strands.
- A fibrin mesh (also called a clot) is then formed, entraps around platelet plug and _stabilizes clot (_right figure above; RBC become trapped giving a venous clot a red appearance).
Primary role of coagulation pathway is to g_enerate thromb_in, which converts circulating fibrinogen to fibrin to make a stable clot.
- It is possible to activate coagulation pathway in two different ways, which are (1) tissue factor pathway, (2) contact activation on a charged surface.
- Coagulation occurs through tissue factor pathway (predominant pathway), first at vessel wall and then on activated platelet surface. Contact pathway (XII) is not required for repair of damaged vessels.
Draw the BASIC flow diagram showing the principles of coagulation
1) Vessel injury and platelet plug formation
2) This and the tissue factors activates the clotting factors
3) Thrombin (IIa)
4) this does lots of things, but the thing it does for the fibrin net, is to convert the fibrinogen to fibrin.
Primary role of coagulation pathway is to_______________, which __________________ (function)
In what state are coagulation factors found in in the plasma vs site of injury
Coagulation factor is in an inactive form in the plasma
Protein cleavage occurs by another protease (either a part is lobbed off, or cut causes a shape change)
Coagulation factor is in an active form at the site of injury
Describe the concept of enzyme catalytics (in the coagulation cascade)
We have a protein that has been activated (protease)
If there were nothing to line the target protein up with the protease, the target protein may not bind accurately (process is inefficient and slow)
However, if we have a cofactor which can line these up, ther reaction is much more efficient.
The coagulation process requires lots of cofactors to make this pathway effecient.
What are the 2 steps (PHYSIOLOGICAL) of the coagulation pathway?
1) Coagulation occurs through tissue factor pathway fir_st at the vessel wall_ and then on the a_ctivated platelet surface_
2) Contact pathway (XII is not required for repair of damaged vessels
Describe the i_nitial phase_ (at site of injury) of the Tissue Factor pathway (extrinsic pathway)
Initial Phase (At Site of Injury)
Tissue factor (e.g. TFVII) is necessary to initiate coagulation.
- It is a trans-membrane protein that is expressed on subendothelial tissues (e.g. fibroblasts, smooth muscle).
- It is not expressed on cellular blood components or endothelium of blood vessels in the resting state.
Following trauma to a vessel, t_issue factor is exposed_ to circulation after endothelial submatrix is disrupted.
Factor VII binds to tissue factor and undergoes a shape change that allows it to be a_ctivated by trace amounts of other clotting factors_. Factor VIIa (activated) is then able to activate factor X to factor Xa, and factor IX to factor IXa.
Factor Xa can then slowly convert a small amount of prothrombin (II) to thrombin (IIa) (requires calcium and phospholipid).
Thrombin amplifies coagulation pathway to produce more thrombin. It does this by activating factor V, VIII and XI (cofactors).
- Factor Va and VIIIa are important cofactors that dramatically speed up coagulation to physiological rate. They align protease on the lipid membrane so it can cleave efficiently.
- Factor XIa amplifies coagulation by converting factor IX to IXa (feedback loop).
—–above occurs at the site of vessel wall injury—–
Describe the Second phase (on activated platelet surface) of the Tissue Factor pathway (extrinsic pathway)
Second Phase (On Activated Platelet Surface)
Factor IXa (protease) and _VIIIa (cofactor) f_orms intrinsic tenase complex (requires calcium and phospholipid). This complex rapidly converts factor X to Xa.
Factor Xa (protease) and Va (cofactor) forms prothrombinase complex (also requires calcium and phospholipid). This complex rapidly converts prothrombin (II) to thrombin (IIa). Therefore, huge burst of thrombin is formed (more than initial phase).
The fibrinogen then becomes fibrin
The coagulation cascade starts with….
Tissue factor
IX a is the ______
VIIIa is the __________
IX a is the protease
VIIIa is the cofactor
For the reactions that occur on the platelet surface to occur, you need……
Calcium/Phosopholipid
So the proteins can sit on the phospholipids, so their tails are bound (lined up properly)
Calcium helps with the configuration of the protein
Describe the formation of the Fibrin Clot
Formation Of Fibrin Clot (after initial and second phase)
Thrombin then cross links fibrinogen molecules to form a fibrin mesh.
- Fibrinogen is composed of three strands (alpha, beta and gamma chains) in plasma. Thrombin cleaves small fragments of both alpha (fibrinopeptide a) and beta chains (fibrinopeptide b). This allows the chains to polymerise into long fibrin strands.
- These chains are stabilised by factor XIIIa. Factor XIII is activated by thrombin.
GIve a summary of the coagulation pathway (extrinsic)
This pathway using tissue factor and factor VII is measured in prothrombin time. This test involves addition of thromboplastin (form of tissue factor) to make blood clot. Because an extrinsic factor had to be added, this was referred to as extrinsic pathway. This pathway takes place in two phases.
- Initial burst of factor VIIa activity with the production of a small amount of factor Xa and thrombin.
- Second phase of thrombin feedback, which activates factors V and VIII to rapidly accelerate thrombin generation.
Phospholipid in these reactions is provided by platelet endothelium of activated platelets. It helps to keep process of coagulation localised to the site of injury.
What is involved in the Initiation complex
Tissue factor, VIIa, IIa (?)
What is involved in complex 2?
Xa, VIIIa, IXa
What is involved in complex 3?
Xa binds with Va to convert Prothrombin II to thrombin IIa
Xa is a protease
Va speeds up reaction
Huge burst of thrombin formed
Much ore than extrinsic Xase (tissue factor/VIIa)
Calcium and phospholibid is required.
Describe 4 Inhibitors of the coagulation pathway.
Inhibitor proteins prevent u_ncontrolled coagulation_ occurring within blood vessels, which is achieved by inhibiting activated proteins.
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Antithrombin (previously called antithrombin III) is the most important inhibitor (requires natural heparin like compounds).
- This primarily inhibits thrombin by forming a 1:1 complex.
- It also i_nhibits factor Xa_ and to lesser extent it inhibits factors IXa and XIa.
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Proteins C and protein S form the other main inhibitory system.
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Protein C is activated by thrombin via cofactor thrombomodulin.
- Thrombomodulin is an endothelial bound protein formed in the endothelial cells. Activated protein C (APC) functions by inactivating clotting factors Va and VIIIa.
- _Protein S i_s a cofactor that enhances the action of activated protein C.
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Protein C is activated by thrombin via cofactor thrombomodulin.
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Tissue factor pathway inhibitor (TFPI) controls the i_nitial phase of coagulation._
- This inhibitor is present in plasma and platelets and accumulates at _coagulation sit_e due to local platelet activation.
- It functions by binding to factor Xa and extrinsic tenase complex (VII+TF), which stops further factor Xa activation.
- This prevents widespread activation of coagulation away from site of injury.
“think money”
think about this illustration of the coagulation pathway
What factors are the proteases?
What factors are the cofactors?
Factor IXa, Factor Xa are proteases
Factor VIIIa and Factor Va are the cofactors
What is needed to convert X to Xa?
1) Ca
2) IXa
3) Phosphoplipid
4) VIIIa
what is needed to convert Prothrombin to Thrombin?
1) Ca
2) Xa
3) Phospholipid
4) Va
What are the vitamin K dependent proteins?
1) VII
2) X
3) IX
4) prothrombin (II)
5) Protein C
6) Protein S
Describe the Contact activation of the Coagulation pathway
NON-physiological
Factor XIIa can activate Factor XI which can trigger this pathway in the lab
Factor XII deficiency is not associated with bleeding
Factor XI deficiency is mild
Have roles in thrombosis when platelet medaiated
What is the role of antithrombins?
Requires natural hepairn like compoudns
1) Inhibits thrombin
2) Inhibits factor Xa
3) Less effecient inhibitor of IXa and XIa
What is Fibrinolysis?
There is a close interrelationship between inflammation, thrombosis and fibrinolysis.
Fibrinolytic system becomes activated when any clot forms.
- Fibrinolysis is the body’s way of keeping coagulation from becoming excessive and breaking down clots.
- It functions to digest the clot so that the normal structure of the vessel can be re-established.
Fibrinolytic cascade involves conversion of plasminogen (inert plasma protein) to digestive enzyme plasmin.
- Conversion of plasminogen involves two major plasminogen activators, which are tissue plasminogen activator (tPA) and urokinase like palsminogen activator (uPA).
How is Fibrnolysis inhibited?
Fibrinolysis is inhibited by at least two different inhibitors.
- Antiplasmin binds to plasmin and forms 1:1 inactive complex, therefore inhibit plasmin activity.
- Thrombin activated fibrinolysis inhibitor (TAFI) is activated by thrombin in presence of thrombomodulin (see protein C above). Activated TAFI removes lysine residues from fibrin and prevents plasminogen binding to clot and inhibits fibrinolysis.
The action of plasmin on cross-linked fibrin generates a heterogeneous mixture of fragments (fibrin degradation products).
How does Fibrinolysis work?
Fibrinolytic cascade involves c_onversion of plasminogen_ (inert plasma protein) to digestive enzyme plasmin.
- Conversion of plasminogen involves two major plasminogen activators, which are tissue plasminogen activator (tPA) and urokinase like palsminogen activator (uPA).
What is the action of plasmin?
The action of plasmin on cross-linked fibrin generates a heterogenous mixture of fragments
