Week 3

Question 1

Define coagulation

Answer 1

The conversion of liquid blood to a gel like solid.

Question 2

Define haemostasis

Answer 2

The stopping of bleeding (not just clotting)

Question 3

Define protease.

Answer 3

An enzyme which cleaves peptide bonds in their substrates.

Question 4

Define proteolysis.

Answer 4

Cleavage of peptide bonds in proteins.

Question 5

What are the three phases of haemostasis?

Answer 5

1 - vasoconstriction
2 - formation of platelet plug
3 - coagulation

Question 6

Describe the normal plasma used in the clotting lab.

Answer 6

Citrated plasma (i.e. sodium citrate added to prevent coagulation in storage) from a bulk sample of plasmas obtained from at least 100 donors.

Question 7

Why is a phospholipid/kaolin used in the clotting lab?

Answer 7

The reactions of the clotting cascade all take place on negatively charged phospholipid surfaces so kaolin is required for this experiment. Kaolin is an aluminium silicate clay on which a suspension of negatively charged phospholipids are adsorbed in order to initiate the cascade.

Question 8

What reagents are used in the clotting lab?

Answer 8

• normal plasma (with sodium citrate)
• kaolin with adsorbed phospholipids
• calcium chloride solution
• chelated calcium chloride solution
• heparin solution
• thrombin
• benzamidine

Question 9

What is Benzamidine?

Answer 9

A protease inhibitor.

Question 10

What is heparin?

Answer 10

A naturally occurring glycosaminoglycan of varying chain lengths which is released from mast cells. It is an anticoagulant.

Question 11

What is the APTT and what does it evaluate?

Answer 11

Activated Partial Thromboplastin Time - evaluates the intrinsic and common pathways of the clotting cascade.

Question 12

What are the stages of an APTT?

Answer 12

1. Plasma incubated with phospholipid and kaolin at 37C
2. Calcium chloride added and incubation continues
3. Time to clot formation measured

Question 13

In an APTT, what surface does the phospholipid/kaolin mimic?

Answer 13

An activated platelet.

Question 14

In the APTT, why is sodium citrate added to the plasma?

Answer 14

To prevent coagulation in storage.

Question 15

What is the difference between blood serum and blood plasma?

Answer 15

Serum is plasma with fibrinogen removed aka the portion of plasma remaining after coagulation of blood.

Question 16

When blood clots form naturally they are red. Why is this?

Answer 16

RBCs are trapped in the clot. (Plasma is used in the lab so no RBCs get trapped)

Question 17

Why does benzamidine slow the time for plasma to clot?

Answer 17

It is a competitive protease inhibitor. As most of the coagulation factors are serine proteases, the benzamidine will compete for the active site of the enzymes. Binding to the active site is reversible so benzamidine will slow the process but not stop it completely.

Question 18

Is calcium needed for thrombin action?

Answer 18

No. A clot will form without calcium. Calcium is needed to form thrombin but is not needed to convert fibrinogen to fibrin by thrombin.

Question 19

Which pathway is the target for heparin action?

Answer 19

The final common pathway because it inhibits the action of thrombin so with thrombin inhibited the final common pathway will not occur.

Question 20

What are the key stages of fibrinolysis?

Answer 20

• plasminogen becomes trapped in clot
• plasminogen is activated by serine protease tPA (tissue plasminogen activator)
• tPA converts plasminogen to plasmin
• plasmin breaks down fibrin mesh

Question 21

After treatment with heparin has been started, why is it important to check the APTT?

Answer 21

• individual variation in coagulation means that effects of heparin are not predictable
• effects on the APTT predict the therapeutic effects well
• so checking the APTT ensures the dosage is appropriate
• too much heparin would increase the risk of bleeding/haemorrhage and too little would be ineffective

Question 22

Why might streptokinase be an advantage to bacteria?

Answer 22

It acts in a similar way to tPA so can digest fibrin strands and break down a clot thus allowing the bacteria to gain access to tissues and bloodstream.

Question 23

Will a patient with a deficiency of factor VII have a prolonged APTT?

Answer 23

No - factor VII is part of the extrinsic pathway so will not affect APTT.

Question 24

Will a patient with a deficiency in factor VIII have a prolonged APTT?

Answer 24

Yes - factor VIII is part of the intrinsic pathway so a deficiency in this will lead to an increased APTT. (Haemophilia A)

Question 25

How are the intrinsic and extrinsic pathways activated?

Answer 25

Intrinsic - blood trauma, blood sticks to exposed collagen of damaged endothelial cells or damaged platelets release phospholipids.

Extrinsic - tissue/cell trauma, tissue factor (TF/thromboplastin) released from damaged cells into blood.

Question 26

Describe the role of vitamin K in blood clotting.

Answer 26

Vitamin K is fat-soluble and absorbed through the intestine along with lipids. It is required by the liver for the synthesis of clotting factors. Therefore, a vitamin K deficiency can lead to uncontrollable bleeding due to a lack of clotting factors and no clot formation.

Question 27

How does prostacyclin impact haemostasis?

Answer 27

It is a prostaglandin that is synthesise and released by endothelial cells and WBCs. It opposes the action of thromboxane A2 (involved in platelet activation) so inhibits the adhesion of platelets.

Question 28

How do antithrombin, heparin and activated protein C inhibit blood clotting?

Answer 28

Antithrombin - inhibits clotting factors II, X and XII
Heparin - inhibits thrombin
Activated protein C - inactivates major clotting factors and stimulates plasminogen activators

Question 29

What is the mode of action of heparin?

Answer 29

It is an activator of antithrombin III which goes on to inhibit thrombin and other clotting factors.