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Case 1

  • Mr EJ
  • 63yo M
  • Haematemesis and malaena
  • Known liver disease secondary to heavy alcohol intake.
  • Why is he bleeding?
  • How can we measure it?
  • How can we help him?
  • Probable upper GI bleed
  • At risk of bleeding due to portal hypertension
    • Can lead to varicies
  • Could also be an ulcer or mallory-weiss tear
  • Could possibly also have liver disease
    • reduced clotting factors
    • want to test LFT to see if this is occuring
      • particularly albumin (albumin is a good marker of enzyme production)
    • also test INR (measure fn of extrinsic pathway)
    • Could also test for Vit. K deficiency (important factor in coagulation pathway)
  • Treatment:
    • Could give FFP (fresh frozen plasma) to fight anaemia
    • Could also give clotting factors and Vit K if needed.

Integrity of Circulatory System

  1. Maintain blood in a fluid state
  2. Limit blood loss from damaged blood vessels

through formation of clot (haemostasis)

  • Balance between procoagulant and anticoagulant mechanisms
  • Haemostasis: physiologic process outside damaged vessel
  • Thrombosis: pathological process within vessel lumen / heart


  • Quick, localised, regulated haemostatic response to breach in blood vessel.
  • Requires
  1. Vasoconstriction
  2. Platelet plug formation (Primary Haemostasis)
  3. Thrombin clot formation (Secondary Haemostasis)
  • Complex activating and inhibiting system
  • Tightly regulated
  • Five components
    • Vessels / Endothelium
    • Platelets
    • Coagulation factors (clot forming)
    • Coagulation inhibitors (clot controlling)
    • Fibrinolysis (clot dissolving)

Cell-based model

  1. Initiation
  2. Amplification, Propagation
  3. Stabilisation

Cellbased model.PNG

Vessel, Endothelium

  • Vasoconstriction
  • Surface
  • Tissue factor
    • Integral membrane protein present on cell membranes in subendothelium
    • Exposed by endothelium injury
  • Production & release of various factors
    • Haemostasis: vWF(von Willebrand Factor), TF (tissue factor)
    • Anticoagulant: AT (antithrombin), TM
    • Fibrinolysis: TPA (tissue plasminogen activator)


  • Anucleate small particulates in the blood
  • Essentially a little packet of granules
  • When activated, they release the granules
    • Granules cause other platelets to release their granules, and cause vasoconstriction


  • Adhesion to collagen
    • Via vWF, GPIb-IX receptor
    • Direct, GPIa receptor
  • Degranulation
    • Release of vasoconstrictors, coagulation factors and growth factors
  • Activation
    • Cytoskeletal rearrangement – morphological change
    • Conformational change of GPIIb-IIIa receptor: can now bind vWF, fibrinogen
  • Aggregation, via Arachidonic Acid pathway
  • Platelet plug
    • Platelets bridged: GPIIb-IIIa receptors bind fibrinogen
  • Thrombogenic surface critical role
    • Activated Factors Va, VIIa, XIa on platelet surface
  • Coagulation cascade initiation / amplification

Coagulation Factors

  • Exposed Tissue Factor
    • Circulating Factor VII binds rapidly
    • FVII activated (VIIa)
  • VIIa/TF complex activates
    • Factors V, VIII, XI
    • VIIIa/IXa Tenase complex
    • Va/Xa Prothrombinase complex
    • Activate II (Prothrombin to Thrombin)
  • Coagulation cascade amplified


  • Thrombin burst
    • Converst soluble fibrinogen to insoluble fibrin mesh
  • Haemostatic fibrin clot
  • FXIIIa stabilises clot further

Clotting Factors

  • Synthesis
    • Liver (most) or endothelium
    • Vitamin K-dependent synthesis:
      • Factors II, VII, IX, X; Protein S, C (γ-carboxylation is vital for biological activity)
    • vWF - megakaryocytes, endothelial cells
  • Factor VIII circulates complexed with vWF
    • Protection from proteolytic degradation

Initiation.PNG Amplification.PNG Amp2.PNG Stab1.PNG

The Anticoagulant System

  • Dilution of procoagulants
  • Secretion of PGI2 and nitric oxide by neighbouring endothelial cells (vasodilators)
  • Removal of particulate matter and activated factors by the reticuloendothelial system
  • Specific inhibitors:
    • Tissue Factor Pathway Inhibitor
    • Protein C and Protein S
    • Antithrombin

Protein C & Protein S

  • Protein C activated on endothelial cell surface by thrombin, in complex with thrombomodulin (endothelial cell receptor)
  • Activated Protein C (APC), combined with Protein S degrades FVa & FVIIIa (which are required to sustain thrombin formation)
  • Protein S produced by liver

APC pathway.PNG


  • Serine plasma protease
  • Major inhibitor of activated coagulation factors, including thrombin
  • Glycosaminoglycans (produced by endothelial cells, eg heparin) are high affinity binding sites for antithrombin
  • AT binds to AG, then rapid inactivation of thrombin


Tissue Factor Pathway Inhibitor

  • Bind and inhibits TF-FVIIa complex
  • Ensures that a small procoagulant stimulus does not elicit an uncontrolled burst of thrombin generation
  • Produced in endothelial cells

Tissuefactor inhibitor pathway.PNG


  • Prevents excessive fibrin deposition
  • Closely coupled with fibrin formation
    • catalysed by fibrin formation
  • Localised surface bound phenomenon


Back to Case 1

  • Haemostasis pathology
    • Endothelial defect
    • Platelet defect
    • Coagulation factor defect
    • Acquired or congenital?

Endothelial Defects

  • Congential
    • Ehlers-Danlos Syndrome
  • Acquired Vascular Purpura
    • Corticosteroid excess
    • Senile purpura
    • Leucocytoclastic vasculitis (immune complex deposition)
    • Leakage from terminal arterioles, capillaries, post-capillary venules
      • Scurvy (vitamin C required for normal collagen metabolism)

Platelet Disorders

  • Clinical
    • Petechiae
      • Capillary bleeding, esp lower limbs
    • Mucosal
    • Menses
  • Aetiology
    • Quantitative (Thrombocytopenia)
    • Qualitative


  • Reduced Production
    • Marrow aplasia:
      • Aplastic anaemia
      • Radiation, drugs
    • Marrow infiltration
      • Malignancy, fibrosis
    • Selectively impaired platelet production
      • Drugs: gold, sulphonamides, ethanol, thiazides
    • Ineffective megakaryopoiesis
      • B12 or folic acid deficiency,
      • Myeloproliferative disease
    • Congenital defects
  • Accelerated Plt Removal
    • Immune
      • ITP
      • SLE, NHL
      • Drugs: quinine, heparin
      • Infection: HIV, measles
      • Transfusion
    • Non-Immnune
      • Disseminated I’vascular Coagulopathy
      • Haemolytic-uraemic syndrome, TTP
      • Sepsis
      • Massive Haemorrhage
  • Sequestration
    • Hypersplenism
      • any cause of a large spleen
  • Idiopathic
    • Platelet-associated IgG
      • Directed against GPIb-IX or GPIIb-IIIa
    • Destruction of coated platelets in spleen
    • Self-limiting (‘Acute’)
      • Children, young adults
      • Frequently preceded by a viral illness
      • Spontaneous remission <3 months
    • Chronic
      • Adults
      • Idiopathic or secondary (LPD, AID, viral infection)
      • Management: corticosteroids, splenectomy, intravenous γ-globulin

Platelet Function Disorders

  • Hereditary
    • Membrane
    • Granules
    • Von Willebrand’s Disease
      • Failure of adhesion via GPIb-IX due to low vWF
  • Acquired
    • Drug-Induced (Aspirin, NSAIDs)
    • Bone marrow disorders (myelodysplasia)
    • Uraemia

Congenital Coagulation Factor Disorders

  • Haemophilia A (Factor VIII deficiency)
  • Haemophilia B (Factor IX deficiency)
  • Von Willebrand Disease

Acquired Coagulation Factor Disorders

  1. Decreased Production
    • Liver disease (F II, V, VII, IX, X, Protein C)
    • Vitamin K deficiency (F II, VII, IX, X)
      • Diet, warfarin
  2. Inhibition
    • Heparin, Low molecular weight heparin (LMWH)
    • New: Thrombin & Xa inhibitors
    • Circulating Inhibitors
      • Coagulation factor antibodies
      • Antiphospholipid syndrome
  3. Increased Consumption, Loss
    • Disseminated Intravascular Coagulopathy (DIC)
    • Massive haemorrhage / transfusion

Disseminated Intravascular Coagulopathy (DIC)

  • Causes
    • Massive tissue damage, tumour, amniotic fluid embolus, snake bite, sepsis
  • Pathogenesis
    • Thrombogenic material in circulation → activation of coagulation → widespread microthrombi →
      • Arterial occlusion at organ
      • Consumption of clotting factors and platelets → bleeding
  • Investigations
    • ↓platelets, ↑APTT, ↑INR, ↑D-dimer, ↓fibrinogen

Back to case 1 (again)

  • Investigations:
    • Platelet count, blood film
    • Platelet function tests
    • Activated Partial Thromboplastin Time (APTT)
    • Prothrombin Time (PT)
      • International Normalised Ratio (INR)
    • Fibrinogen, D-dimers
    • Factor Assays

Activated Partial Thromboplastin Time (APTT)

  • Measures the clotting time after the activation of contact factors, but without added tissue thromboplastin
  • Intrinsic pathway
  • Depends on contact factors, Fs V, VIII, IX, X, XI, XII, prothrombin & fibrinogen
  • Sensitive to circulating inhibitors & heparin
  • Not usually affected by LMWH

Prothrombin Time (PT)

  • Measures the clotting time of plasma in the presence of tissue extract (thromboplastin)
  • Extrinsic clotting system
  • Depends on prothrombin, Fs V, VII, X, fibrinogen concentration
  • Express results as INR
  • International Normalised Ratio


New notes

  • Want to end up with a clot (fibrin) quickly; but generalised clotting = lose all clotting factors and DIC. Needs to be carefully regulated. Therefore there are lots of redundancies. Really the clotting pathway isn't linear.
  • Extrinsic pathway: activate factor 7, feeds directly into the common pathway. Very quick - occurs when you expose blood to air.
  • Intrinsic pathway: less important for life. Activated by a lot of contact factors e.g. LPS. Much slower. All of it is calcium dependent.
    • Coag tests - has citrate in it when transporting the tube to lab, to deplete it of calcium. When you want to do the test, you add Ca.
  • Extrinsic pathway: break in blood vessel, exposes tissue factors to the circulation. Factor VII becomes activatedl. Platelets - stick to the exposed endothelium by von Willebrand's factor. So you have platelet stuck to endothelium and activated VII. When a platelet is stuck down, it secretes granules, which recruit other platelets (positive feedback). This forms the platelet plug.
  • Common pathway: activated factor 7 causes activation of factor 10, which binds to factor 5 and then lives on a platelet. Factor 8 accelerates the process by giving factor 10 somewhere to sit.
  • Intrinsic pathway: starts with factor 12; end up back at the common pathway again.

Formation of fibrin

  • Cleave fibrinogen --> fibrin, which is sticky
  • Factor 13 stabilises fibrin
  • Left over bits = fibrinogen degradation products; can be used to test for coagulation

APC pathway - contains 4 inherited thrombophilias

  • Negative feedback for clotting
  • Protein C sees thrombin (in common pathway) and becomes activated PC (=APC)
  • APC binds to protein S and inactivates factor 5 (which is needed for factor 10 to stick to platelets) --> this inactivates the common pathway.
  • Protein S deficiency --> can't turn off coagulation --> increased clotting
    • Similar with protein C deficiency
  • Factor 5 Leiden --> allows F5 to work, but it confers resistance to APC
  • Prothrombin excess: mutation can stabilise prothrombin RNA --> more prothrombin and more thrombin in circulation
    • This causes thrombophilia: more likely to clot

Heparin and antithrombin

  • Antithrombin doesn't do much until it binds to heparin - then it cleaves all the clotting factors. Antithrombin deficiency --> can't turn off clotting --> strong clotting predisposition
  • Heparin - activates antithrombin 3 to stop clotting. Doesn't work in antithrombin 3 deficiency - then you need to give AT3.
  • LMW heparin -- predominantly results in reduced factor 10; makes it harder to measure clotting


  • Fibrinogen degradation products fall off fibrinogen when you make fibrin
  • Fibrin itself can be dissolved --> fibrin degradation products.
  • Fibrin degraded by plasmin (which comes from plasminogen)
    • Needs a plasminogen activator to do this
    • Then it throws off FDPs
  • Measuring FDPs: if increased - recent clotting episode, or has had surgery
    • Absence: they haven't had a clot (very good at excluding recent thrombotic episode)
  • t-PA is an important cofactor to activate plasmin.
    • MI: we have fresh clot in coronary arteries - need to dissolve it quickly. So you give people t-PA to break down the clot to reduce damage.
    • Same thing can be done in strokes; but more controversial because it predisposes people to bleeding

Vital facts

NB: Factor 2 is prothrombin

  • Serine proteases complex with a cofactor on a membrane to produce activation and amplification of the next step in the sequence
  • Most procoagulants are synthesised in the liver
  • Some are vitamin K dependent
    • Factors 2, 7, 9 10; Anticoagulants: S, C (gamma carboxylation is vital for biological activity)
    • Vitamin K deficiency (e.g. not enough vegetables, or malabsorption) - warfarin is vitamin K antagonist. Prolongs both extrinsic and intrinsic pathway.
  • Factor 8 circulates complexed with vWF, which protects it from proteolytic degradation
    • vWF stacks up to link the platelet to the endothelium
    • vWF disease: 1) don't make it 2) mutations, so you can't stack it effectively.
    • vWF deficiency also causes low Factor 8 --> abnormal coagulation tests

Coagulation tests (i)

  • Activated partial thromboplastin time (APTT)
    • kaolin added to whole plasma to provide surface activation of factors 7, 6
    • platelet lipid surrogate is added, and the time to produce fibrin is recorded
    • affected by deficiencies of factors preceding activation of X: ie sensitive to action of heparin, and used to monitor heparin therapy
  • normal: 30-40 seconds
  • slowed down by things upstream of factor 10. fractionated heparin affects these things --> can measure efficacy of therapy with APTT
  • ddx for APTT deficiency
    • liver disease
    • inherited factor deficiency
    • global factor deficiency (e.g. DIC)

Coag test (ii)

  • Prothrombin time (PT)
    • thromboplastin/tissue factor added to plasma: time to produce fibrin is recorded
    • reported as a corrected prothrombin ratio (PR), expressed as an INR which is corrected to take account of differences in thromboplastins. (INR is so that different labs produce the same result)
    • sensitive to abnormalities of the extrinsic pathway (1, 2, V, VII, X) - used to monitor warfarin therapy
  • D-Dimer (FDPs) - latex agglutination

Coag test (iii)

  • Platelets aren't very reproducible, hard to measure function
  • Tests of platelet function
    • Blood film
    • Bleeding time
    • Aggregometry (this is what we do now)
      • sends test serum down a tube that contains collagen, ADP etc to make platelets sticky. Measure how long it takes for that to coagulate. It will coagulate quickly if it's responding well.
        • good screen for platelet abnormality

Too little coagulation

  • Thrombocytopenia
  • Platelet function defects (or not enough platelets)
  • Coagulation factor deficiencies
    • Inherited
    • Failure of synthesis
    • Consumption
  • Vascular purpura
  • Is it platelets? Is it blood factors? Is it vessels?

Thrombocytopenia (i)

  • Petechiae - capillary bleeding (lower limbs, buccal mucosa, abrasions)
  • Platelet production defets
    • Marrow aplasia (radiation, drug, primary)
    • Marrow infiltration (malignancy, fibrosis)
    • Selectively impaired platelet production (gold, sulphonamides, ethanol, thiazides, etc)
    • Ineffective megakaryopoeisis
  • Accelerated platelet removal
    • Immune (ITP, SLE, NHL, drug, heparin, infection, HIV, sepsis)
    • Non-immune (DIC, haemolytic, uremic syntdrome/TTP, transfusion)
  • Platelet sequestration
    • hypersplenism - any cause of a large spleen
  • DIC: prolonged INR, prolonged APTT, low fibrinogen, thrombocytopenia

Platelet function disorders

Drugs: NSAIDs, clopidogrel, renal failure

  • Hereditary
    • Membrane
    • Granules
    • vW disease
  • Acquired
    • Bone marrow disorders
    • Uremia
    • Drug-induced (see above)

vW disease

  • Deficiency of vWF
    • circulates as multimers of a 270 kd subunit
    • carrier for VIII
    • platelet adhesion via GPIb receptor
  • lifelong history of mucosal bleeding, severe bleeding during surgery, trauma
  • Ix:
    • increased bleeding time
    • decreased antigenic vWF
    • abnormal ristocetin-induced platelet aggregation
    • increased APTT
    • decreased VIII:C level
  • history:
    • personal and family history of bleeding
    • torrential bleeding after a minor operation (e.g. dental, tonsillectomy). Do you think you bleed excessively?
  • no easy way to screen for it

Haemophilia A

  • deficiency/defective factor VIII:C
  • x-chromosome (Xq28) - sex-linked
  • >600 mutations identified: point mutations, insertions, deletions
  • spontaneous/traumatic soft tissue haemorrhages, particularly into joints (arthropathy)

Acquired disorders of coagulation

  • Vitamin K deficiency (II, VII, IX, X)
    • diet, warfarin
    • also needed for PC and PS, which have short half lives. Warfarin - deplete yourself of PC and PS before you deplete of clotting factors. Therefore heparinise people before giving warfarin (don't increase clotting that way)
  • hepatic disease (2, 10, 7, 9, 10, protein C)
    • DIC massive tissue damage, tumour, amniotic fluid, embolus, sepsis, etc
    • thrombocytopenia, abnormal APTT, INR, fibrinogen, elevated D-dimer
  • inhibitors
    • antibodies to coagulation factors
    • antiphospholipid antibodies

Hypercoagulable states

  • antiphospholipid syndrome
  • deficiencies
    • antithrombin 3 deficiency (heparin resistance)
    • protein C, S (loss of ability to inactivate VIIIa, Va)
  • Trousseau's syndrome (migratory thrombophlebitis associated with malignancy)
    • Esp: mucoid tumours (pancreatic, gastric)
  • Obesity, immobility, oestrogen therapy, pregnancy
  • women who have had a previous clot shouldn't take oestrogen