Normal haemostasis involves platelet plug formation, coagulation cascade, and fibrin clot formation to stop bleeding at the site of vascular injury. It is regulated by the endothelium, platelets, and coagulation factors. The coagulation cascade can be initiated through the intrinsic or extrinsic pathway, both leading to thrombin generation and fibrin clot formation. Counter-regulatory mechanisms like fibrinolysis and natural anticoagulants limit clot formation to the site of injury. Laboratory tests monitor components of the coagulation cascade like platelet count and function, prothrombin time, activated partial thromboplastin time, and fibrinogen levels.

1. NORMAL HAEMOSTASIS &
COAGULATION CASCADE
DR YOGESH AGRAWAL
RESIDENT
MD TRANSFUSION
MEDICINE
SMS MEDICAL COLLEGE

2. HAEMOSTASIS: (blood+ stable)
Haemostasis is defined as a set of well-regulated processes designed to
rapidly and specifically form a clot at sites of vascular injury.
It is the first stage of wound healing.
• Normal haemostasis accomplish two important functions:
• Maintenance of blood in a fluid, clot free state.
• A rapid & localized haemostatic plug formation at a site of vascular injury.

3. SEQUENCE OF EVENTS IN HEMOSTASIS:
1. VASOCONSTRICTION: After initial
injury, there is brief period of
arteriolar vasoconstriction
attributable to:
1. Reflex neurogenic mechanisms.
2. Local factors derived from
endothelium like ENDOTHELIN, a
potent vasoconstrictor.
This effect is transient.

4. 2. PRIMARY HEMOSTASIS/PLATELET PLUG
FORMATION
 Endothelial injury exposes highly
thrombogenic sub-endothelial extracellular
matrix (ECM).
 Platelets adhere to ECM & become
activated.
 Platelets undergo shape change & release
secretory granules, which recruit additional
platelets (aggregation) to form haemostatic
plug k/a primary haemostasis.

5. 3. SECONDARY HEMOSTASIS/COAGULATION
Tissue factor, pro-coagulant factor
synthesized by endothelium is exposed
at the site of injury. Activate
coagulation cascade, resulting in
activation of thrombin.
Thrombin converts circulating soluble
fibrinogen to insoluble fibrin, causing
local fibrin deposition.
This is k/a secondary hemostasis.

6. 4. CLOT STABILIZATION AND RESORPTION
 Polymerised fibrin & platelet
aggregates form solid permanent
plug to prevent any further
haemorrhage.
 At this stage, counter-regulatory
mechanisms e.g. t-PA are set into
motion to limit the haemostatic plug
at the site of injury.

7. HAEMOSTASIS contd…
Normal haemostasis involves the interaction of
1. Blood vessel wall (endothelium)
2. Platelets
3. Coagulation factors

8. ROLE OF ENDOTHELIUM
Maintains normal flow of blood by
1. Anti-platelet effect,
2. Anti-coagulant effect, and
3. Fibrinolytic properties.

9. 1. ANTI-PLATELET EFFECTS
 Prevents spontaneous platelet aggregation
 Produces :
1. Antithrombin.
2. Von Willebrand's factor (vWF).
3. Vasodilators like NO and prostacyclin (PGI2) which inhibits
platelet aggregation.

10. 2. ANTI-COAGULANT EFFECTS
 Mediated by:
• Heparin-like molecules.
• Thrombomodulin.
• Tissue factor pathway inhibitor (TFPI).
 Heparin-like molecules interact with antithrombin III to inactivate
thrombin , factor Xa & several other coagulation factors

11. ANTI-COAGULANT EFFECTS CONTD..
Thrombomodulin binds thrombin
activate protein C
proteolytic cleavage of factors Va & VIIIa
Inhibits clotting

12. ANTI-COAGULANT EFFECTS CONTD..
• Endothelium is also a major source for tissue factor pathway
inhibitor, that complexes & inhibits activated tissue factor- factor
VIIa and factor Xa molecules.

13. 3. FIBRINOLYTIC EFFECTS
• Endothelium synthesizes tissue-type Plasminogen Activator
(t-PA), promoting fibrinolytic activity to clear fibrin deposits
from endothelial surfaces.

15. ROLE OF PLATELETS:
•After vascular injury, platelets encounter ECM constituents &
undergo 3 general reactions:
1) Platelet adhesion
2) Secretion (Release Reaction)
3) Platelet aggregation

16. 1. PLATELET ADHESION
• Platelets adhere to extra-cellular
matrix (ECM) via vWF which acts as
bridge b/w plt. Surface receptors
(e.g. GP 1b) & exposed collagen.

18. 2. SECRETION
Release of contents of α and δ-granules.
Dense body contents are especially important because:
I. Calcium is required in the coagulation Cascade.
II. ADP is a potent mediator of plt. Aggregation and
“Recruitment”

20. 3. PLATELET AGGREGATION
Important Stimulus for plt. aggregation:
I. ADP
II. TXA2
III. Thrombin

21. ROLE OF COAGULATION SYSTEM:
1. Constitutes 3rd component of the haemostatic process & is a major
contributor to thrombosis.
2. Results in formation of thrombin.
3. Each reaction step involves an enzyme (an activated coagulation factor), a
substrate (an inactive proenzyme form of a coagulation factor), and a
cofactor (a reaction accelerator).
4. Blood coagulation pathway has been divided into extrinsic & intrinsic
pathways, converging where factor X is activated.

22. There are major 13 factors which are involved in the coagulation
cascade. All these factors are blood proteins or their derivatives.
Even if one of the factor is defective, the whole clotting process is
impaired leading to hemorrhage. These factors are F-I to F-XIII.

24. • There are 3 major stages in the coagulation cascade:
• Stage 1: Formation of Prothrombinase Complex (Prothrombin Activator)
• Prothrombinase is formed in two ways:
• Extrinsic Pathway (also known as Tissue Factor Pathway)
• Intrinsic Pathway (also known as Contact Activation Pathway)
• Stage 2: Conversion of Prothrombin into Thrombin.
• Stage 3: Conversion of Fibrinogen into Fibrin
• Stage 2 & Stage 3 is collectively called as Final Common Pathway

27. Extrinsic Pathway
• In this pathway, the formation of prothrombinase complex is initiated by the tissue thromboplastin
Mechanism:
•It begins with trauma to blood vessel or tissues outside the blood vessel. It releases F-VII and tissue
phosholipids.F-VII comes in contact with F- III (TF or Thromboplastin) expressed on TF-bearing cells
(stromal fibroblasts & leukocytes) forming an activated complex (TF-VIIa)
•TF-VIIa activates F-X in presence of Ca++ and tissue phospholipids
•F-Xa acts on F-V and activates it
•F-Xa complexes with tissue phospholipids, F-Va, Ca++ and forms a complex called prothrombinase
complex or prothrombin

28. INTRINSIC PATHWAY ;
• In this pathway, the formation of prothrombinase complex is initiated by platelets which are within the
blood itself .(starts with S.E.C.)
• MECHANISM: Begins with the formation of the primary complex on collagen by HMWK, prekallikrein and F-XII
• Prekallikrein is converted to kallikrein and F-XII gets activated.
• Damaged platelets adhere to the wet surface of blood vessel and release platelet phospholipids.
• F-XIIa acts enzymatically on F-XI (Plasma Thromboplastin Antecedent) and activates it
• F-XIa acts enzymatically on F-IX and activates it in presence of Ca++
• F-IXa activates F-VIII (Anti Haemophilic Factor)
• F-VIIIa and F-IXa activate F-X
• F-Xa acts enzymatically on F-V (Proaccelerin) and activates it in presence of Ca++
• F-Va,F-Xa, Phospholipid and Ca++ form a complex called prothrombin complex

29. • • In the presence of prothrombin activator or prothrombinase complex and
calcium prothrombin is converted to thrombin
• •Thrombin then activates other components of the coagulation cascade,
including F-V and F- VIII (which activates F-XI,which in turn activates F-
IX)and activates and releases F-VIII from being bound to vWF
• •F-VIIIa is the co-factor of F-IXa,and together they form the "tenase"
complex, which activates F-X and so the cycle continues.("Tenase" is a
contraction of "ten" and the suffix "-ase" used for enzymes)

30. • • Thrombin converts fibrinogen (plasma protein produced by the liver) to
fibrin
• • Thrombin also activates F-XIII (Fibrin Stabilizing Factor) which in
presence of Ca++ stabilizes the fibrin polymer through covalent bonding
of fibrin monomers

31. • Once activated, the coagulation cascade must be restricted to the local
site of vascular injury to prevent clotting of the entire vascular tree.
• Clotting is also regulated by 3 types of natural anti-coagulants:
1. Anti-thrombin
2. Protein C & protein S
3. Tissue factor pathway inhibitor (TFPI)
• Clotting cascade also sets into motion a fibrinolytic cascade that limits the
size of final clot. This is accomplished by the generation of plasmin from
plasminogen.

32. Factors limiting Coagulation
Once initiated, coagulation must be restricted to the site of vascular injury to
prevent deleterious consequences.
1. One limiting factor is simple dilution; blood flowing past the site of injury
washes out activated coagulation factors, which are rapidly removed by the
liver.
2. A second is the requirement for negatively charged phospholipids, which, as
mentioned, are mainly provided by platelets that have been activated by
contact with subendothelial matrix at sites of vascular injury.
However, the most important counterregulatory mechanisms involve factors that
are expressed by intact endothelium adjacent to the site of injury.
Activation of the coagulation cascade also sets into motion a fibrinolytic cascade
that limits the size of the clot and contributes to its later dissolution. Fibrinolysis is
largely accomplished through the enzymatic activity of plasmin, which breaks down
fibrin and interferes with its polymerization. An elevated level of breakdown
products of fibrinogen (often called fibrin split products), most notably fibrin-
derived D-dimers, are a useful clinical markers of several thrombotic states.

33. • Plasmin is generated by enzymatic catabolism of the inactive circulating
precursor plasminogen, either by a factor XII–dependent pathway (possibly
explaining the association of factor XII deficiency and thrombosis) or by
plasminogen activators. The most important plasminogen activator is t-PA; it is
synthesized principally by endothelium and is most active when bound to fibrin.
This characteristic makes t-PA a useful therapeutic agent, since its fibrinolytic
activity is largely confined to sites of recent thrombosis. Once activated, plasmin
is in turn tightly controlled by counterregulatory factors such as α2-plasmin
inhibitor, a plasma protein that binds and rapidly inhibits free plasmin.

35. Coagulation Monitoring – Conventional Tests ( details in separate
session)
* Platelets – Number and Function
• Clotting studies= PT(PT-INR) ,APTT ,Thrombin time, Fibrinogen levels
etc.