Overview
This lecture covers the coagulation arm of Haemostasis: the coagulation factors and their sources, the intrinsic/extrinsic/common pathway model and how it maps onto the PT and APTT laboratory tests, the cofactor, calcium and phospholipid requirements for the cascade, fibrin formation and cross-linking, vitamin K dependence (illustrated by a warfarin overdose case), the inhibitors that keep coagulation in balance, and fibrinolysis as the clot-clearing counterpart.
Haemostasis overview
- Bleeding triggers three parallel responses that converge on “bleeding stopped”: (1) vessel constriction, (2) platelet activation and platelet plug formation, (3) activation of the coagulation cascade to generate fibrin.
Coagulation factors
- Factor I, Fibrinogen, plasma protein, made in liver, common pathway, converted to fibrin.
- Factor II, Prothrombin, plasma protein, liver (vitamin K dependent), common pathway, converted to thrombin.
- Factor III, tissue thromboplastin/tissue factor (TF), lipoprotein mixture, from damaged cells and platelets, extrinsic pathway.
- Factor V, Proaccelerin, plasma protein, liver and platelets, common pathway.
- Factor VII, Proconvertin, plasma protein, liver (vitamin K dependent), extrinsic pathway.
- Factor VIII, antihaemolytic factor A, plasma protein, platelets and endothelium, intrinsic pathway.
- Factor IX, antihaemolytic factor B, plasma protein, liver (vitamin K dependent), intrinsic pathway.
- Factor X, Stuart-Power factor, protein, liver (vitamin K dependent), common pathway.
- Factor XI, antihaemolytic factor C, plasma protein, liver, intrinsic pathway.
- Factor XII, Hageman factor, plasma protein, liver, intrinsic pathway.
- Factor XIII, fibrin-stabilizing factor, plasma protein, liver and platelets, stabilises fibrin and slows fibrinolysis.
The coagulation pathways
- Intrinsic pathway: factors XII, XI, IX, VIII; also called the contact activation pathway; measured by the activated partial thromboplastin time (APTT).
- Extrinsic pathway: tissue factor and factor VII; also called the tissue factor pathway; measured by the prothrombin time (PT).
- Common pathway: factors X, V, prothrombin → thrombin, fibrinogen → fibrin; both intrinsic and extrinsic pathways converge here.
- Simplest cascade: XII → XI → IX → X (VII also feeds into X); X → Xa; prothrombin + Xa → thrombin; fibrinogen + thrombin → fibrin.
- Extrinsic (tissue factor) pathway steps: (1) exposure and expression of tissue factor after endothelial damage or activation; (2) TF binds FVIIa; (3) the TF/FVIIa complex (“tenase”) activates FX; (4) Xa activates prothrombin; (5) thrombin activates fibrinogen. The extrinsic pathway initiates coagulation after endothelial damage; tissue factor is “extrinsic” to blood, especially present in the vessel wall.
- Intrinsic (APTT) pathway is activated experimentally by kaolin or silica; it proceeds XII → XI → IX → X without involving factor VII.
- All factors are activated by proteolytic cleavage: XII→XIIa, XI→XIa, VII→VIIa, IX→IXa, X→Xa, prothrombin (II)→thrombin (IIa), fibrinogen→fibrin.
- Cofactors are enzymatic proteins that enhance activity of coagulation factors rather than being enzymes themselves: FV and FVIII are cofactors. FV helps (accelerates) FX activity (as Va with Xa); FVIII helps FIX activity (as VIIIa with IXa).
- Coagulation factors require calcium (Ca2+) and phospholipid membranes for activity, at both the IX/X/VIIIa step and the Xa/prothrombin/thrombin step.
- Coagulation occurs anchored on phospholipid membranes, especially on platelets: the “Xase” complex (FVIIIa, FIXa, FX) on the membrane converts FX to FXa; FXa then combines with FVa and prothrombin (FII) in the “prothrombinase” complex to generate thrombin. Platelets expose phosphatidylserine on their surface, creating the negatively charged phospholipid surface coagulation reactions occur on.
Laboratory tests: PT and APTT
- Prothrombin time (PT): plasma + Ca2+, phospholipid and tissue factor activates the extrinsic pathway; a fibrin clot forms after about 10 seconds; PT ≈ 10 sec. Only the extrinsic/common pathway is active in this test (intrinsic factors not involved).
- Activated partial thromboplastin time (APTT): plasma + Ca2+, phospholipid and silica/kaolin activates the intrinsic pathway; a fibrin clot forms after about 25 seconds; APTT ≈ 25 sec. Only the intrinsic/common pathway is active (factor VII not involved).
Fibrin structure, cross-linking and degradation
- Fibrinogen structure: D-domain–E-domain–D-domain, with fibrinopeptide A and fibrinopeptide B cleavage sites at the E-domain.
- Thrombin cleaves fibrinopeptides A and B from fibrinogen to produce fibrin.
- Fibrin monomers polymerise into a fibrin polymer.
- FXIII (FXIIIa) cross-links the fibrin polymer, which is essential for the strength of the fibrin clot; fibrin provides structural strength for the haemostatic plug and anchors it to adjacent tissue.
- Fibrin is constantly degraded by plasmin into fibrin degradation products (FDPs).
- D-dimers are one of these degradation products and are measured diagnostically to assess clot formation/turnover.
Source and vitamin K dependence of coagulation factors
- Liver produces: fibrinogen; FII, FVII, FIX and FX (FII = prothrombin); FV; FXI; FXII. Liver failure can therefore cause major haemostatic problems.
- Endothelium produces: FVIII (from liver endothelium) and von Willebrand factor.
- Factors II, VII, IX, X (and proteins C and S) require post-translational modification to bind Ca2+: γ-carboxylation of glutamate residues, a vitamin K-dependent process carried out by vitamin K-dependent γ-glutamyl carboxylase.
- Vitamin K cycle: vitamin K → vitamin K hydroquinone (via the carboxylase, using CO2) → carboxylates the inactive factors (II, VII, IX, X, protein C, protein S) into their activated, Ca2+-binding forms, generating vitamin K epoxide, which is recycled back to vitamin K.
- Warfarin blocks the recycling steps of this cycle (vitamin K epoxide → vitamin K, and vitamin K → vitamin K hydroquinone), preventing γ-carboxylation of factors II, VII, IX and X.
Clinical case: warfarin overanticoagulation
- 87-year-old male on warfarin for pulmonary embolism and atrial fibrillation.
- Results over three dates showed marked prolongation: PT rose to >100.0 sec (reference 9.0–13.0), INR rose to >10.0 (reference 0.8–1.2), APTT rose to 71 sec (reference 20–31); fibrinogen remained roughly normal (2.1–2.2 g/L, reference 1.7–4.3).
- Warfarin interferes with γ-carboxylation of factors II, VII, IX, X; overdose leads to very high PT and APTT.
- Management: rescue with vitamin K.
Inhibitors of coagulation
A clot forms in about 10 seconds, so inhibitors are essential to prevent uncontrolled ("giant") clot formation — coagulation is a balance between activators and inhibitors.
- Antithrombin binds irreversibly to and inactivates factor XIa, factor IXa, factor Xa and thrombin (an older name for antithrombin is antithrombin III).
- Protein C (assisted by protein S) inactivates factor VIIIa and factor Va.
- Both mechanisms ultimately block fibrin formation.
- Balance: activation triggers coagulation; inhibitors are needed to stop it. Too little coagulation causes bleeding risk; too little inhibition or too much coagulation causes thrombosis risk. Thrombolysis (breakdown of fibrin clots) is the third arm of this balance.
- Families with antithrombin, protein C or protein S deficiency have a high rate of venous thrombosis (deep vein thrombosis and pulmonary embolism) at a young age.
Fibrinolysis
- Fibrinolysis degrades fibrin clots and exists in a constant equilibrium with clot formation.
- Endothelium releases tissue plasminogen activator (tPA), which is inhibited by plasminogen activator inhibitor (PAI).
- tPA converts plasminogen (bound to fibrin) into plasmin; plasmin breaks down the fibrin clot into fibrin degradation products (FDPs); plasmin itself is inhibited by plasmin inhibitor.
- Fibrin is readily lysed via tPA from normal endothelial cells; healthy endothelium secretes tPA in response to fibrin and can clear excess fibrin inside vessels.
- Synthetic tPA is given therapeutically for arterial thrombosis, e.g. heart attack (treatment window about 12 hours) or stroke (treatment window about 3 hours) — the quicker the better.
- The overall haemostasis/coagulation cycle: haemorrhage → platelet activation (via collagen/tissue factor exposure and platelet recruitment) → clotting cascade (intrinsic and extrinsic pathways converge at factor X → Xa → prothrombin → thrombin → fibrinogen → fibrin) → platelet stabilisation (stable clot) → fibrinolysis (plasminogen → plasmin, fibrin degradation) → back to haemorrhage.
Self-test
- Name the three parallel responses triggered by bleeding that together achieve haemostasis.
- List the factors of the intrinsic pathway, the extrinsic pathway, and the common pathway.
- Describe the steps of the extrinsic (tissue factor) pathway from endothelial damage to fibrin formation.
- Distinguish the PT test from the APTT test in terms of what is added to the plasma, which pathway each measures, and the approximate clotting time for each.
- What are cofactors in the coagulation cascade, and which two coagulation factors act as cofactors, and for which enzymes?
- What two additional components (besides enzymes) are required for the coagulation reactions to proceed, and where anatomically do these reactions occur?
- Describe the steps by which fibrinogen is converted into a stable, cross-linked fibrin clot, including the enzyme and factor responsible for cross-linking.
- What are D-dimers and why are they measured clinically?
- List the coagulation factors produced by the liver and those produced by the endothelium.
- Describe the process and purpose of vitamin K-dependent post-translational modification of coagulation factors, naming the factors affected.
- Explain how warfarin interferes with coagulation at the molecular level.
- A patient on warfarin presents with PT >100 sec, INR >10, and APTT 71 sec. Explain why both PT and APTT are prolonged, and state the treatment to reverse this.
- List the three main inhibitors of coagulation and state which coagulation factors each inactivates.
- Explain why deficiency of antithrombin, protein C or protein S predisposes to venous thrombosis at a young age.
- Describe the steps of fibrinolysis, including the roles of tPA, PAI, plasminogen, plasmin and plasmin inhibitor.
- Why is synthetic tPA given for arterial thrombosis, and what are the approximate treatment windows for heart attack and stroke?
- Explain, using the intrinsic, extrinsic and common pathway concepts, why coagulation, once triggered, must be tightly balanced by both inhibitors and fibrinolysis to prevent inappropriate clotting or bleeding.
Answers
Reveal answers
- Vessel constriction, platelet activation and platelet plug formation, and activation of the coagulation cascade leading to fibrin formation; all three converge on bleeding being stopped.
- Intrinsic: factors XII, XI, IX, VIII. Extrinsic: tissue factor and factor VII. Common: factors X, V, prothrombin (converted to thrombin), fibrinogen (converted to fibrin).
- Tissue factor is exposed/expressed after endothelial damage or activation; TF binds FVIIa; the TF/FVIIa “tenase” complex activates FX to Xa; Xa activates prothrombin to thrombin; thrombin activates fibrinogen to fibrin.
- PT: plasma + Ca2+, phospholipid and tissue factor; measures the extrinsic/common pathway; clot forms in about 10 seconds. APTT: plasma + Ca2+, phospholipid and silica/kaolin; measures the intrinsic/common pathway; clot forms in about 25 seconds.
- Cofactors are enzymatic proteins that enhance the activity of coagulation factors rather than acting as enzymes themselves. FV (as Va) is a cofactor that helps FX(a); FVIII (as VIIIa) is a cofactor that helps FIX(a).
- Calcium (Ca2+) and phospholipid membranes; the reactions occur anchored to phospholipid membranes, especially on the surface of activated platelets (which expose phosphatidylserine).
- Thrombin cleaves fibrinopeptides A and B from fibrinogen to produce fibrin monomers; these polymerise into a fibrin polymer; FXIII (as FXIIIa) cross-links the polymer to form a strong, stable clot that anchors to adjacent tissue.
- D-dimers are fibrin degradation products generated when plasmin breaks down cross-linked fibrin; they are measured diagnostically to assess clot formation/turnover.
- Liver: fibrinogen, FII (prothrombin), FVII, FIX, FX, FV, FXI, FXII. Endothelium: FVIII (from liver endothelium) and von Willebrand factor.
- Factors II, VII, IX, X (and proteins C and S) undergo γ-carboxylation of glutamate residues, catalysed by a vitamin K-dependent γ-glutamyl carboxylase, which allows them to bind Ca2+ and become functional.
- Warfarin blocks the recycling of vitamin K epoxide back to vitamin K and of vitamin K to vitamin K hydroquinone, depleting the reduced vitamin K needed for γ-carboxylation of factors II, VII, IX and X, so these factors cannot bind Ca2+ properly.
- Both PT (extrinsic/common pathway, factors VII, X, prothrombin) and APTT (intrinsic/common pathway, factors IX, X, prothrombin) are prolonged because warfarin impairs γ-carboxylation of the vitamin K-dependent factors II, VII, IX and X, which are shared across both pathways via the common pathway and the extrinsic/intrinsic starting factors. Treatment: give vitamin K.
- Antithrombin inactivates factor XIa, factor IXa, factor Xa and thrombin. Protein C (assisted by protein S) inactivates factor VIIIa and factor Va.
- These proteins normally inhibit coagulation; their deficiency removes a brake on the cascade, tipping the activation/inhibition balance towards excess coagulation and thrombosis, presenting as venous thrombosis (DVT/PE) at a young age.
- Endothelium releases tPA (inhibited by PAI); tPA converts plasminogen bound to fibrin into plasmin; plasmin degrades fibrin into fibrin degradation products (FDPs); plasmin is itself inhibited by plasmin inhibitor.
- Synthetic tPA rapidly activates fibrinolysis to dissolve an arterial thrombus; treatment windows are approximately 12 hours for heart attack and approximately 3 hours for stroke, with earlier treatment being better.
- Coagulation is initiated rapidly (clot forms in ~10 seconds) via convergence of the intrinsic and extrinsic pathways on the common pathway, so without control it would propagate into a giant clot; antithrombin and the protein C/protein S system restrain the cascade by inactivating key activated factors, while fibrinolysis (tPA/plasmin) continuously clears fibrin, together maintaining the balance needed for effective haemostasis without excess bleeding or thrombosis.