5.2. Hemostasis and the role of thrombocytes. Blood coagulation. Fibrinolysis. Physiological anticoagulant mechanisms. Flashcards
I. Hemostasis
1. Definition of hemostasis
prevention of bleeding from a damaged vessel (prevention of hemorrhage)
I. Hemostasis
2. What are the stages of hemostasis?
PRIMARY HEMOSTASIS
1. Vasoconstriction
2. Increased tissue pressure
3. Formation of a platelet plug
SECONDARY HEMOSTASIS
Coagulation (formation of a blood clot)
I. Hemostasis
3A. What are the mechanism and role of vasoconstriction (primary hemostasis)?
1/ Mechanism:
- Upon injury of the blood vessel wall, there is an immediate response called vasoconstriction
- Vasoconstriction will decrease blood flow towards injured area, leading to decrease in blood loss
2/ Role: to give more time for more effective hemostatic processes to take place
I. Hemostasis
3B. What are the causes of vasoconstriction (primary hemostasis)?
1/ Neurogenic reflex
2/ Endothelin release
3/ Release of vasoconstrictor substances
I. Hemostasis
3B1. How does neurogenic reflex participate in vasoconstriction (primary hemostasis)?
produces vasoconstriction
I. Hemostasis
3B2. How does Endothelin release participate in vasoconstriction (primary hemostasis)?
- Endothelin Binds to to ET-receptor on SMC (smooth muscle cell), leading to vasoconstriction
I. Hemostasis
3B3. What are the vasoconstrictor substances that are released by thrombocytes? Consequences?
Chemical released by thrombocytes
- Thromboxane A2 (TXA2, synthesized by platelets)
=> Ca2+ signal
=> Vasoconstriction - Serotonin (synthesized by platelets)
- Epinephrine (not synthesized, taken up by platelets) => ⍺1 adrenergic receptor acting on smooth muscle contraction
I. Hemostasis
3C. Mechanism and role of Increased tissue pressure?
Increased tissue pressure contributes to hemostasis because:
- It decreases radius, which causes significant decrease in blood flow; decrease r by a factor of 2 would diminish flow by a factor of 16 according to - Poiseuille law.
E.g, We do this naturally by pressing a finger against a small cut to stop the bleeding.
I. Hemostasis
3D. What is the role of Platelet plug formation (step 3 in primary hemostasis)?
platelets plug small ruptures in the endothelium
I. Hemostasis
3E. What are the stages of Platelet plug formation (step 3 in primary hemostasis)?
1/ Adhesion
2/ Activation
3/ Aggregation
I. Hemostasis
3F. What are platelets?
platelets are fragments of megakaryocytes
I. Hemostasis
3G. What are characteristics (structure) with corresponding functions of platelets
1/ Have a dense tubular system
-> Acts as a Ca2+ storage
2/ Have dense granules containing ADP, serotonin and polyphosphates to activate platelets
3/ Have alpha granules which contain clotting factors
4/ Have actin + myosin for later contraction to reduce the size of the clot
5/ Have adhesion glycoproteins found on their surface
ex: GPIa/IIa, GPVI
I. Hemostasis - Stages of platelet formation
3H1. Why is platelet adhesion necessary? When will platelet adhesion occur?
1/ Platelets do not adhere to endothelium directly because platelets have negative surface charge and so do the endothelium (proteoglycans – heparan sulfate).
2/ Platelet adhesion occurs in response to ↑ shear stress at platelet surface or endothelial cells, and in response to vessel injury or humoral signals.
I. Hemostasis - Stages of platelet formation
3H2. What are the steps af platelet adhesion?
1/ Platelets first adhere to the injured space via glycoprotein-collagen binding with glycoprotein GP VI and GP Ia – IIa
=> This is a Direct platelet interaction
2/ Endothelial cells also release the von Willebrand factor (vWF) which also binds to exposed collagen and then create an indirect interaction between GP Ib – IX – B
I. Hemostasis - Stages of platelet formation
3I1. How are platelets activated?
Binding of GP VI to collagen initiates tyrosine phosphorylation cascade -> Phospholipase C (PLC) activation -> IP3 -> Ca2+-release = platelet is activated
*Note: Inositol trisphosphate or inositol 1,4,5-trisphosphate abbreviated InsP3 or Ins3P or IP3
I. Hemostasis - Stages of platelet formation
3I2. What are the 5 consequences of increased concentration of intracellular Ca2+?
- Exocytosis of granules
→ ADP, 5-HT, epinephrine, Ca2+, procoagulant factors, fibrinogen…
→ Positive feedback - Phospholipase A2 (PL2) activation
→ TXA2-production → positive feedback → platelet activation increased - Cytoskeletal rearrangement
→ platelets undergo shape change, increased surface - Exposure of GPIIb-IIIa integrin complex
→ fibrinogen binding, platelet aggregation - Phospholipid externalization
→ coagulation cascade
I. Hemostasis - Stages of platelet formation
3I3. The 5 consequences of increased contraction of intracellular Ca2+
-> What are the consequences of Exocytosis of granules?
→ ADP, 5-HT receptors (serotonin receptors), epinephrine, Ca2+, procoagulant factors, fibrinogen…
→ Positive feedback
I. Hemostasis - Stages of platelet formation
3I4. The 5 consequences of increased contraction of intracellular Ca2+
-> What are the consequences of PLA2 activation?
→ TXA2-production → positive feedback → platelet activation increased
I. Hemostasis - Stages of platelet formation
3I5. The 5 consequences of increased contraction of intracellular Ca2+
-> What are the consequences of 3. Cytoskeletal rearrangement?
→ platelets undergo shape change, increased surface
I. Hemostasis - Stages of platelet formation
3I6. The 5 consequences of increased contraction of intracellular Ca2+
-> What are the consequences of 4. Exposure of GPIIb-IIIa integrin complex?
→ fibrinogen binding, platelet aggregation
I. Hemostasis - Stages of platelet formation
3I7. The 5 consequences of increased contraction of intracellular Ca2+
-> What are the consequences of 5. Phospholipid externalization?
→ coagulation cascade
I. Hemostasis - Stages platelet formation – 2. Platelet activation
3j. How is platelet activation inhibited?
By synthesis of PGI2 and NO in endothelial cells, it will inhibit the increase of intracellular Ca2+ concentration which subsequently inhibit platelet activation and aggregation
- PGI2 -> PGI2 – R (GS) -> Cause increase in cAMP
- NO -> GC -> Increase in cGMP
*Note:
- Prostaglandin I2 (PGI)
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 3. Platelet plug formation
3K. What are the Steps of platelet aggregation?
1/ Thrombus initially develops through the stabilization of discoid platelet aggregate
2/ Shedding of microparticles from the platelet plasma membrane (caused by a high shear stress) also contributes to aggregation
3/ ADP + TXA2 released from platelets attract more platelets to the affected area
4/ Exposed GP IIb-IIIa binds to RGD sequence and RGD proteins
- RGD sequence is a tripeptide Arg-Gly-Asp motif in a subunit of fibrinogen
- RGD proteins are vWF, fibrinogen, fibronectin and vitronectin
=> Platelets cross-bind to each other
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation a
3L. What are Characteristics of blood coagulation?
1/ Occurs primarily because of the conversion of fibrinogen to fibrin (via thrombin)
2/ Thrombin itself must first be activated and have the necessary cofactors to perform this function
- Thrombin activation occurs via intrinsic and extrinsic pathway
- Intrinsic (contact activation) and extrinsic (tissue factor) converge at the activation of factor X
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation a
3M. How does Intrinsic pathway work?
1/ Intrinsic pathway: activated by factors that are found within blood.
2/ Factors XII gets activated = XIIa -> XIIa converts factor XI to factor XIa -> XIa activates factor IX to factor IXa
-> IXa forms the “intrinsic tenase complex” with VIIIa, phosphotidylserine (PS) and Ca2+ = (Ixa – VIIIa – PS – Ca2+)
-> Intrinsic tenase converts factor X to factor Xa
-> Factor Xa is generated on the platelet surface
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation a
3N. How does Extrinsic pathway work?
1/ Extrinsic pathway: activated by tissue factor found outside of the blood (VSM)
2/ With injury to a blood vessel, the factor VIIa is exposed to tissue factor -> form a complex together with Ca2+ called the ‘’extrinsic tenase complex’’ (TF-VIIa-Ca2+)
3/ Extrinsic tenase complex activates factor X -> factor Xa (common pathway) and factor IX -> factor IXa (intrinsic pathway)
-> Factor Xa is generated by extrinsic tenase complex on the surface of fibroblasts
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation a
3L. How does Final common pathway work?
1/ Factor Xa (gotten from both intrinsic + extrinsic pathway) forms a complex with factor Va, that is called ‘’prothrombinase complex’’
2/ Prothrombinase complex is used to activate prothrombin (II) into thrombin (IIa)
-> Thrombin cleaves fibrinogen to fibrin (fibrin mesh network)
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation
3O. What are the 4 4 pro-coagulative effects by thrombin (IIa)?
1/ Activate platelets directly via PAR-2
2/ Catalyze the proteolysis of fibrinogen (I) into fibrin (Ia).
3/ Activate factor XIII (stabilizing factor) for cross links
4/ Activate factor V, VIII (positive feedback)
I. Hemostasis - Stages of hemostasisPrimary hemostasis – 4. Blood coagulation a
3P. Compare Extrinsic vs Intrinsic pathway
1/ Extrinsic
- Starts early
- Explosive (within seconds)
2/ Intrinsic
- Starts later
- Slower (within minutes)
- Accelerates fibrin formation
- Stabilizes the blood clot
I. Hemostasis - Stages of hemostasis - Secondary hemostasis – 4. Blood coagulation
3Q1. What is the structure of Gla proteins?
A complex consist of
- Procoagulants: prothrombin, factor VII, IX, X
- Anticoagulants: protein C & S
I. Hemostasis - Stages of hemostasis - Secondary hemostasis – 4. Blood coagulation
3Q2. What does the synthesis of Gla proteins require?
1/ Requires vitamin K
2/ Occur in liver
I. Hemostasis - Stages of hemostasis - Secondary hemostasis – 4. Blood coagulation
3R. How does Vitamin K-dependent carboxylation occur?
1/ γ-glutamyl carboxylase uses a vitamin K cofactor to convert glutamic acid into γ-carboxyglutamic acid, Gla (by adding carboxylic acid).
2/ In the process of serving as a cofactor (donate ē), vitamin K is converted to vitamin K epoxide. Vitamin K epoxide gets converted back into vitamin K quinone by epoxide reductase = reusable!
I. Hemostasis - Stages of hemostasis - Secondary hemostasis – 4. Blood coagulation
3S. Importance of vitamin K in blood coagulation?
1/ Gla structure allows Ca++ to bind, which induces factor II, VII, IX, X to activate.
2/ Warfarin inhibits epoxide reductase, blocking vitamin K from getting recycled = coagulation cascade would not work properly.
I. Hemostasis - Stages of hemostasis - Secondary hemostasis – 4. Blood coagulation
3T. What are the Steps of fibrin formation?
1/ The formation of fibrin takes place when thrombin converts fibrinogen dimers to fibrin monomers
2/ To stabilize the fibrin mesh network, factor XIIIa (transglutaminase) (activated by thrombin) converts the fibrin monomers to a network of fibrin polymers
=> The fibrin network is then insoluble and much more difficult to break
=> When plenty of thrombin is available, a dense fibrin network is formed and traps blood cells and plasma, preventing further bleeding
II. Anticoagulation mechanisms
1. What are the factors involved in anticoagulation?
1/ Paracrine factors
2/ Anticoagulant factors – inhibition of the clotting cascade
- Tissue factor pathway inhibitor
- Antithrombin
- Thrombomodulin
- Protein C and S
3/ Clearance of activated clotting factors
II. Anticoagulation mechanisms
1A. How are paracrine factors involved in anticoagulation?
Endothelial cells generate prostacyclin, which promotes vasodilation and inhibits platelet activation. They also generate NO (stimulated by thrombin), which inhibits platelet adhesion and aggregation through cGMP.
II. Anticoagulation mechanisms
1B. How are Tissue factor pathway inhibitor (TFPI) and Antithrombin involved in anticoagulation?
1/ Tissue factor pathway inhibitor (TFPI)
=> Glycosylphosphatidylinositol (GPI) – anchored to endothelial cell membrane inhibits VIIa
2/ Antithrombin III binds and inhibits thrombin & Xa
=> Heparins (mast cells, basophils) & Heparin-sulfate (endothelial surface) promotes this, inhibiting coagulation
II. Anticoagulation mechanisms
1C. How is Thrombomodulin involved in anticoagulation?
Thrombomodulin is the Glycosaminoglycan on endothelial cells
-> Binds and removes thrombin from circulation
-> Binds to protein C
II. Anticoagulation mechanisms
1D. How is protein C involved in anticoagulation?
Protein C binds to thrombin-thrombomodulin complex
-> Activated by thrombin
-> Protein C inactivates Va & VIIa
II. Anticoagulation mechanisms
1E. How is protein S involved in anticoagulation?
Protein S is a cofactor of protein C
III. Procoagulant effects of thrombin
1. What are the 4 Procoagulant effects of thrombin?
1/ Activation of platelets (through PAR-1)
2/ Activation of downstream components in the clotting cascade
3/ Positive feedback at several upstream levels of the cascade
4/ Inhibition of early fibrinolysis
III. Procoagulant effects of thrombin
1A. What are consequences of Activation of downstream components in the clotting cascade?
catalyze the proteolysis of fibrinogen
-> activates XIII factor
III. Procoagulant effects of thrombin
1B. What are consequences of Positive feedback at several upstream levels of the cascade?
- catalyze the proteolysis of prothrombin
-> catalyze the activation of XI factor
-> catalyze the formation of the cofactors Va and VIIIa
III. Procoagulant effects of thrombin
1C. What are consequences of Inhibition of early fibrinolysis?
high local concentration → activation of fibrinolysis-inhibitor
IV. What are Anticoagulant effect(s) of thrombin?
Activation of Protein C (through thrombomodulin)
V. Fibrinolysis
1. What are definition and role of Fibrinolysis?
1/ Definition: Breakdown of blood clot
2/ Role
- Prevents accumulatio of non-aquedate blood clot
- Removes already unnecessary blood clot
V. Fibrinolysis
2. What is the mechanism of Fibrinolysis?
1/ When a clot is formed, a large amount of plasminogen is trapped in the clot along with other plasma protein
2/ The injured tissues and vascular endothelium slowly release t-PA and u-PA , which coverts plasminogen to plasmin a Ser protease)
3/ Plasmin causes degradation of fibrin
V. Fibrinolysis
3. How is fibrinolysis inhibited?
1/ Process: synthesis of plasminogen activator inhibitor such as ⍺2-antiplasmin and ⍺2-macroglobulin
- Function: restriction of fibrinolysis
2/ TAFI (thrombin-activatable fibrinolysis inhibitor)
- Function: inhibit the conversion of plasminogen into plasmin
VI. What are some anticoagulants medications?
1/ vitamin K antagonists (e.g, warfarin) which inhibit FIX, FX, FII, FVII
2/ Direct thrombin inhibitors (e.g, dabigatran) which inhibit FIIa
3/ Anticoagulant outside the body
- Heparin
- Ca2+ chelators
