Haem 7 - Haemostasis Flashcards
What is haemostasis
Cellular and biochemical processes that enable both specific and regulated cessation of bleeding in response to vascular insult
What is haemostasis for
Prevent blood loss from intact and injured vessels, enable tissue repair
What may tip the haemostatic balance towards bleeding
Increased fibrinolytic factors or anticoagulant proteins
What may tips the balance towards the thrombotic side
Increased coagulant factors, platelets
Haemostatic plug formation occurs in response to injury to endothelial cell lining
- Vessel constriction - VSMC contract locally —> limits blood flow to injured vessel
- Formation of unstable platelet plug - platelet adhesion + platelet aggregation —> limits blood loss + provides surface for coagulation
- Stabilisation of plug with fibrin - blood coagulation –> stops blood loss
- Vessel repair and clot dissolution - cell migration/proliferation and fibrinolysis –> restores vessel integrity
On vessel wall, the endothelial cell is (by definition) anticoagulant - e.g. EPCR. The Layers under the endothelial cell are procoagulant.
Y
Subendothelium = procoagulant - basement membrane contains elastin, collagen.
VSMC and fibroblasts also present, alongside Tissue factor (TF)
Describe platelets
- Small (2-4 um)
- Anuclear
- Lifespan = 10 daysish
- Platelet count = 150-350 x10^9/l
All platelets in our blood are derived from?
Megakaryocytes (4000 can be derived from just one MK)
Haematopoietic stem cell –> promegakaryocyte –> megakaryocytic —> platelet
What are the various roles that platelets have
- Haemostasis and thrombosis
- Cancer
- Atherosclerosis
- Infection
- Inflammation
Explain how VWF is involved in haemostasis (platelet adhesion)
- Multimeric VWF circulates in plasma in globular conformation. Platelet binding site hidden from GP1B on platelet
- Vascular injury - damages endothelium and exposes sub-endothelial collagen
- Various collagen binding sites that are exposed bind globular VWF
- VWF recruited and undergoes structural transformation - unravelled in response to shear forces of flowing blood
- VWF binds to GP1B of platelet - recruits platelets to site of vessel damage
- IF LOW SHEAR CONDITIONS (i.e. not arteries/capillaries) - platelets can also bind directly to collagen via GPVI and A2B1.
- Platelets activated - further recruit platelets
Explain platelet activation.
- Collagen and thrombin also activate platelets
- Platelets bound to collagen/VWF release ADP and thromboxane - activates/primes platelets
- Activated platelets (a2bB3) recruit additional platelets. a2bB3 also binds fibrinogen. Platelet plug develops. This helps slow bleeding and provides surface for coagulation
4.
Upon adhesion, activation and aggregation, how does the platelet change?
It starts to spread (once it has become a spreading platelet then irreversible adhesion - but the stage before (hemisphere shaped platelet) is reversible adhesion)
What is the most common inherited bleeding disorder
VWD
Causes ineffective platelet recruitment as lack/dysfunctional VWF
What platelet disorders may contribute to abnormal haemostasis
Low platelet count (thrombocytopenia) or improperly behaving platelets
Tethered VWF may also get unravelled by shear forces of flowing blood
What is the key role of thrombin
To cleave fibrinogen into fibrin
Where do most of our clotting factors come from?
Liver
the factors are all serine proteases domain containing
What are the cofactors involved in coagulation
TF
F5a
F8a
Serine proteases contain a catalytic triad - which is?
His/Asp/Ser
The serine proteases cleave substrates after specific Arg and Lys residues
How is coagulation initiated?
extrinsic pathway - TF
Tissue damage causes Tissue factor (TF - integral membrane protein) to come into contact with F7a (plasma protein) - they both bind and switches on F7a.
Hence, TF is a cellular receptor for F7/7a - TF is the only coagulation factor that doesn’t require an activation step
Where is TF located?
At extravascular sites - expressed higher in certain organs
Describe the domain structure that F7, F9, F10 and Protein C & S all share
Serine protease, EGF2, EGF1, Gla domain
(going closer to membrane)
Gla domain binds to phospholipid surfaces
EGF domain involved in protein-protein interactions
All the relevant factors circulate in zymogen form
What is the formation of a Gla domain dependent on?
Vit K
Glutamic acid —- (Vit K carboxylase) —> adds on a carboxylic acid - so there are now 2 negatively charged carboxylic acid domains –> amino acids can now bind Ca very tightly
Binding of Ca causes the structural transition so it can bind phospholipids
THIS IS HOW WARFARIN works
What 2 things does the TF-F7a complex do?
Proteolytically activated F10 and F9 by removing activation peptide - yields active enzyme
What does F10a do?
Activates prothrombin - generates thrombin
However, this step is inefficient AF, as only little quantities of thrombin are generated.
But thrombin activates F8a, and F8a is a cofactor for F9a. (Thrombin also activates f5a)
This F8a/F9a complex can then activate F10a. Activation of f10a this way is way more efficient than activation of f10a by TF.
Also, F5a (also activated by thrombin) forms a complex with f10a and can convert prothrombin into thrombin much more efficiently)
Haemophilia A is a deficiency in?
wb Haemophilia b?
A = F8
B = F9
They are both X-linked
What are the 3 means of regulating coagulation
- TFPI - tissue factor pathway inhibitor
- APC and protein S
- Anti-thrombin
How does TFPI work
usually v low conc in blood
TFPI-F10a complex can bind/inactivate TF-F7a active site via Kunitz domain.
Essentially docks the 4 mofos.
TFPI regulates the initiation of coagulation
Describe the protein C pathway
Thrombin aka F2A
Protein C - plasma protein.
Activated by thrombin-thrombomodulin complex on endothelial cells (Protein C –> APC)
APC inhibits thrombin generation - proteolytically inactivates procoagulant cofactors F5A and F8A
Protein C pathway regulates propagation phase of coagulation - down regulates thrombin generation (doesn’t inhibit thrombin)
What are the 2 cofactors
F5A and f8a
What happens when thrombin binds to thrombomodulin
It turns from a procoagulant protease to an anticoagulant protease —> activates PC (by removing activation peptide)
Activation of protein C by definition occurs at the edge of a clot
What is antithrombin (AT)
SERPIN (SERine Protease INhibitor).
AT inactivates many activated serine proteases (F10A, thrombin, F9A, F11A) - it mops up free serine proteases that escape the site of vessel damage
How does heparin work?
Binds to thrombin and makes it much better at inhibiting thrombin or F10A
Deficiencies of AT, protein C and protein S are important RFs for?
Thrombosis
How is the clot dissolved?
tpa (tissue plasminogen activator) converts plasminogen into plasmin
Plasmin then helps to degraded the clot
tPA can be used therapeutically for thrombosis - promotes dissolution of the clot
Name common anticoagulants
Heparin, warfarin, DOACs
Name common anti platelet agents
Aspirin, P2Y12 blockers
D-dimer can be used as a test to determine?
How much haemostasis has been going on
D-dimer is a fragment of fibrin that is produced when a clot gets degraded - gives snapshot of how much haemostasis has been occurring beforehand
