Lecture 7/8 - Haemostasis

Question 1

Haemostasis: what is it and what are the 5 main components of it

Answer 1

The ability to minimise blood loss after injury

Blood vessels - vasoconstriction
Platelets
Coagulation factors
Coagulation inhibitors
Fibrinolysis

Question 2

What is the process of hemostatic plug formation?

Answer 2

Vessel injury - collagen exposure - platelet adhesion - platelet activation - platelet aggregation - primary hemostatic plug

Vessel injury + platelet acivation - vasoconstriction - reduced blood flow - primary hemostatic plug

Vessel injury (tissue factor) + platelet phsopholipid (platelet activation) - thrombin - fibrin + primary hemostatic plug - hemostatic plug

Question 3

Endothelial cells: what do they do, how do they affect hemostasis, and what do they produce

Answer 3

Maintain vascular integrity

Have potent anti-hemostatic influence (prostacyclin, nitric oxide, vasodilatory properties, platelet adhesion inhibition)

Von Willebrand factor (vWF), tissue plasminogen activator (fibrinolysis)

Question 4

Vasoconstriction

Answer 4

Immediate (few minutes) smooth muscle cell activation before other mechanisms take over

Question 5

Platelets

Answer 5

Produced in bone marrow as a fragment (2-4µm diameter) of a megakaryocyte (1 ->4000)

Lifespan of 9-10 days, normal platelet count 150-400x10^9/L

Destroyed in spleen and liver by Kupffer cells

700-40-1

Question 6

Platelet production

Answer 6

Endomitotic synchronous nuclear replication
Nuclear lobes increase in multiples of two. Cytoplasmic volume increases and becomes more granular and platelets are produced

Negative feedback - number of platelets circulating

Stimulated by thrombopoietin (TBO) and stimulated by IL-3 and GM-CSF

Question 7

Platelet structure: what are the components, what do they do, and what do glycoproteins do?

Answer 7

• Glycocalyx (contain glycoproteins (GPIa, GPIb, GPIIb, etc))
• Glycogen (energy supply), dense tubular system (prostaglandin and thromboxane A2 synthesis)
• Platelet contractile proteins (make aggregation irreversible)
• Electron dense (delta) granules (contain Ca²⁺, ADP, ATP, and serotonin)
• Alpha granules (platelet-derived growth factor, platelet factor 4, fibrinogen, vWF, factors V and VIII)
• Canalicular system - allow granule release

The glycoproteins allow adhesion to surfaces like collagen

Question 8

Platelet function

Answer 8

Adhesion, secretion (granule release reaction), and aggregation

vWF often binds with GPIIb and GPIIIa

Question 9

Hemostatic stable plug

Answer 9

Secretes ADP - and thromboxane A2 - increasing aggregation

Platelets swell due to the ADP presence

Positive feedback occurs

Question 10

Coagulation factor I: name and active form

Answer 10

Fibrinogen

Fibrin

Question 11

Coagulation factor II: name and active form

Answer 11

Prothrombin

Serine protease

Question 12

Coagulation factor III: name and active form

Answer 12

Tissue factor

Co-factor

Question 13

Coagulation factor V: name and active form

Answer 13

Labile factor

Co-factor

Question 14

Coagulation factor VII: name and active form

Answer 14

Proconvertin

Serine protease

Question 15

Coagulation factor VIII: name and active form

Answer 15

Antihaemophilic factor

Co-factor

Question 16

Coagulation factor IX: name and active form

Answer 16

Christmas factor

Serine protease

Question 17

Coagulation factor X: name and active form

Answer 17

Stuart-Prower factor

Serine protease

Question 18

Coagulation factor XI: name and active form

Answer 18

Plasma thromboplastin antecedent

Serine protease

Question 19

Coagulation factor XII: name and active form

Answer 19

Hageman factor

Serine protease

Question 20

Coagulation factor XIII: name and active form

Answer 20

Fibrin stabilising factor

Serine protease

Question 21

Coagulation cascade initiator

Answer 21

TF is the main initiator and binds with coagulation factors (VII-VIIa)

Question 22

The process of fibrin generation

Answer 22

Injury -> Factor VII is activated and binds to tissue factor forming factor VIIa TF

VIIa TF -> activates both factor X and IX

IXa and VIIa -> activate factor X

Xa and Va -> convert prothrombin into thrombin

Thrombotic activation -> Va, XIa, and IXa and VIIIa formation

Thrombin converts fibrinogen and fibrin

XIa -> activates more XI and IX

Tissue factor pathway inhibitor (TFPI)

Question 23

Coagulation cascade initiation: what are the steps?

Answer 23

Initiation:
* Extrinsic tenase complex
* Generation of trace thrombin
* Down-regulation of initiation by TFPI

Amplification

Propagation:
* Formation of intrinsic tenase complex
* Formation of prothrombinase complex
* Burst of thrombin generation

Question 24

Initiation: Extrinsic tenase complex

Answer 24

Involves FVIIa, TF, FX, phospholipids, Ca²⁺

Question 25

Initiation: Generation of trace thrombin

Answer 25

Factor IIa is activated, Ca²⁺ is present, and factor Xa activates a small amount of thrombin

Question 26

Initiation: Down-regulation of initiation by TFPI

Answer 26

Factor Xa is limited by TFPI

Question 27

Amplification

Answer 27

The produced factor IXa and thrombin, although not enough to convert fibrinogen into fibrin, can amplify the process by feedback and activate factor VIII, a co-factor for factor IX, and factor V, a co-factor for factor X This results in an amplified cascade

Question 28

Propagation: Formation of intrinsic tenase complex

Answer 28

FXa and FVIIa activate FIX and the intrinsic tenase complex is formed, consisting of FIXa, FX, FVIIIa, Ca²⁺, and Phospholipids (obviously meaning this is occurring on the membrane, near the site of injury(?))

Question 29

Propagation: Formation of prothrombinase complex

Answer 29

The prothrombinase complex is composed of FXa, FVa, FII, Ca²⁺, and phospholipids Since the process has been amplified, the inhibitor has less of a role in inhibiting FX This process ends with prothrombin being activated

Question 30

Propagation: Burst of thrombin generation

Answer 30

FXa, FVa, FIIa, Ca²⁺ FVa and FXa - convert prothrombin into thrombin Thrombomodulin, a thrombin co-factor, has procoagulant and anticoagulant properties The produced thrombin increases the process to be stimulated by positive feedback The end result of this process is a stable fibrin clot, only degradable by fibrinolysis

Question 31

The core components of the extrinsic tenase, intrinsic tenase, and prothrombinase pathways?

Answer 31

Extrinsic tenase: FVIIa, TF, FX, Ca²⁺, and K⁺ Intrinsic tenase: FIXa, FVIIIa, FX, Ca²⁺ and K⁺ Prothrombinase: FXa, FVa and FII, Ca²⁺ and K⁺

Question 32

Fibrin clot: when is it added, what happens to the platelet plug, what is the final form of the hemostatic plug, and what mediates clot retraction?

Answer 32

Fibrin is added to the primary hemostatic plug, causing platelet mass and platelet-induced clot retraction/compaction Fibrin increases as the platelet plug degranulates/autolyses After a few hours, the entire haemostatic plug is composed of cross-linked fibrin Clot retraction is medicated by glycoprotein receptors (GpIIb/IIIa)

Question 33

What are the inhibitors present in the activating of the tissue factors?

Answer 33

Tissue Factor Pathway Inhibitor (TFPI) – VIIa and Xa Antithrombin III – Xa and thrombin Protein C and Protein S – PS inactivate Va and VIIIa, the action of PS is enhanced by PC and PC also enhances fibrinolysis

Question 34

Tissue factor pathway inhibitor: how quickly does it act, where is it found, what causes its accumulation, and what is inhibited?

Answer 34

1st inhibitor to act Found in plasma and platelets Accumulates due to platelet activation Inhibits FVIIa and FXa

Question 35

Antithrombin: where is it synthesised and what does it inhibit/inactivate?

Answer 35

Synthesized in the liver and endothelium. Inactivates serine proteases, principally factor Xa and thrombin

Question 36

Protein C: what is it, how is it activated, how is its action enhanced, and what does it do?

Answer 36

Vitamin K-dependent protein made in the liver Activated via a complex which inactivates factors Va and VIIIa The action of protein C, enhanced by the action of protein S, binds protein C to the platelet surface Activated Protein C enhances fibrinolysis by inactivating the tissue plasminogen activator (tPA) inhibitor

Question 37

Protein S: what is it, how is it activated, and what does it do?

Answer 37

Vitamin K-dependent protein made in the liver Activated via a complex which inactivates factors Va and VIIIa Enhances the action of protein C

Question 38

Fibrinolysis: what is it, how does it occur, and what is generated?

Answer 38

The degradation of fibrin (clot) tPA activates plasminogen to plasmin which degrades fibrin Generates soluble fragments called fibrin degradation products (FDPs)

Question 39

Fibrinolytic systems

Answer 39

Plasminogen is activated into plasmin by tissue plasminogen activator (tPA) as well a factor XIIa, and then plasmin breaks fibrinogen down into fibrin

Question 40

Streptokinase: what is it?

Answer 40

A drug used to break down clots extremely effectively

Question 41

What inhibits the fibrinolytic system?

Answer 41

Plasminogen activator inhibitor (PAI-1) inhibits tPA and factor XII α2-antiplasmin inhibits the action of plasmin

Question 42

What methods are used to detect haemolytic issues?

Question 43

Bleeding time: how is it monitored, how long should bleeding normally last for, and in what conditions is the bleeding time normal/abnormal?

Answer 43

Application of pressure to the upper arm (blood pressure cuff) with small incisions made in the forearm surface skin Bleeding normally stops in 3-8 mins Prolonged in thrombocytopenia (reduced platelet count) but normal in cases of abnormal vascular bleeding

Question 44

Platelet count and function: how are platelet counts done and how is platelet function monitored?

Answer 44

* Full blood count – platelet count * Bone marrow examination - when platelet issues are suspected * Platelet aggregometry – most valuable * Measure the change in light absorbance as the platelets aggregate * Adhesion studies * Von Willebrand assay

Question 45

Prothrombin time: what does it do, what is the normal PT, and how is it standardised?

Answer 45

Measure factors VII, X, V, prothrombin, and fibrinogen and their integrity Normal PT is 10-14 seconds Prolonged in liver disease, oral anticoagulant treatment (OAT), e.g. warfarin PT standardised as International Normalised Ratio (INR) – used to monitor OAT

Question 46

Activated partial thromboplastin time: what does it do, what is the normal time, and when is it prolonged?

Answer 46

APTT measures factors VIII, IX, XI and XII as well as X, V, prothrombin and fibrinogen Normal 3-40 seconds Prolonged in haemophilia, heparin therapy

Question 47

Which measuring technique measures which haemolytic pathway?

Answer 47

Extrinsic pathway - PT Intrinsic - APTT Contains a little overlap when it comes to factors X, V, and prothrombin and fibrinogen This allows for some diagnosis as if either PT or APTT is fine but the other isn't then you know there is an issue with the other