Blood Coagulation, Haemostasis and its Investigation

Question 1

What is haemostasis?

Answer 1

It is a protective process evolved in order to maintain a stable physiology.

“An explosive reaction designed to curtail blood loss, restore vascular integrity and ultimately preserve life.”

Question 2

Normal haemostasis is a state of equilibrium.

What is it an equilibrium between?

Answer 2

Fibrinolytic factors and anticoagulant proteins are in equilibrium with coagulation factors and platelets.

Question 3

What occurs as a result of chronic venous insufficiency?

Answer 3

• atrophic changes
• hyperpigmentation
• ulceration
• infection

Question 4

What are the three stages of the haemostatic system?

Answer 4

PRIMARY HAEMOSTASIS:

• vasoconstriction (immediate)
• platelet adhesion (within seconds)
• platelet aggregation and contraction (within minutes)

SECONDARY HAEMOSTASIS:

• activation of coagulation factors (within seconds)
• formation of fibrin (within minutes)

FIBRINOLYSIS:

• activation of fibrinolysis (within minutes)
• lysis of the plug (within hours)

Question 5

As a recap, describe primary haemostasis.

Answer 5

With haemostasis at rest, the triggers and cofactors are separated by endothelial cells.
If the endothelial tissue is gone, we expose the blood to components of the subendothelial tissue. When collagen, which was in the subendothelial tissue, is exposed, it attracts Von Willebrand factors (vWFs).
Normally, vWFs circulate in the blood as globules, but when there is an injury, it exposed a sticky part of its molecule which recognises collagen, so it will stick to the site of injury.
The vWFs bind platelets. When the platelets bind, they change morphologically (become activated); they become flatter for a larger surface area. They also release important mediators of coagulation.
More platelets bind and get activated, and eventually, coagulation occurs on the platelet surfaces.
This plug of platelets, localised to the site of injury, provides the phospholipid surface upon which secondary haemostasis takes place.

We say that vWF activity occurs under shear stress because, in areas of injury, the blood flow becomes more turbulent.

Question 6

Describe platelets during haemostasis.

Answer 6

For ADHESION, vWF binds to extracellular collagen and GPIb-IX-V complex.
For ACTIVATION, GPIIb-III undergoes conformational change.

With activation, there are structural platelet shape changes and the ‘release’ reaction.

Platelets also expose negatively charged phospholipids and provide coagulant surfaces.

Question 7

How can we tell that the classical concept of coagulation may not be completely right?

Answer 7

The classical concept of distinct intrinsic, extrinsic then common pathways is NOT physiologically correct; however, it is useful to interpret a routine coagulation screen.

Nevertheless, it FAILS to explain why:

• FXII deficient patients do not bleed
• FXI deficiency is usually only a mild disorder
• FVIII/IX deficiency causes severe bleeding disorder but should be bypassed by FVII activity
• thrombin generation has a lag phase then rapid reproduction

Question 8

What are some features of blood sample collection for haemostasis testing?

Answer 8

• the accuracy of the haemostasis laboratory depends on the quality of the specimen submitted
• blood is anticoagulated with 3.2% (0.109 M) sodium citrate (it leeches calcium out of the cell)
• most tubes contain 0.3 mL anticoagulant and require 2.7 mL of blood
• underfilling the tube yields grossly inaccurate results

Question 9

What are some example of pre-analytical variables that can affect the blood testing?

Answer 9

PROBLEMS WITH BLUE-TOP TUBE:

• partial fill tubes
• vaccum leak and citrate evaporation

PROBLEMS WITH PHLEBOTOMY:

• heparin contamination
• wrong label
• slow fill
• underfill
• vigorous shaking
• difficult venepuncture

BIOLOGICAL EFFECTS:

• Hct >55 or <15
• lipaemia, hyperbilluribinaemia, haemolysis

LABORATORY ERRORS:

• delay in testing
• prolonged incubation at 37 C
• freeze/thaw deterioration

Question 10

What are some principles of clotting tests?

Answer 10

You incubate plasma with reagents necessary for coagulation.

• phospholipid, co-factors
• trigger or activator
• calcium

You also measure the time taken to form the fibrin clot.

Question 11

How would you perform a mixing study?

Answer 11

• mix the patient and normal plasma in equal volumes (50:50 mix)
• repeat abnormal coagulation test

Results:

• if the test normalises, there are factor deficiencies
• if the test remains abnormal, there is something interfering with the coagulation (an inhibitor, usually an antibody)

Question 12

Describe D-dimer testing.

Answer 12

• it’s a measure of the D-dimer, a fibrin degradation product
• it’s found elevated in the situation of enhanced fibrinolysis (thrombosis, DIC)
• it’s not specific for thrombosis also elevated as an acute phase reactant
• a negative result is useful if clinical suspicion of VTE is low