Week 11

• Haemostasis

  "Global process whereby vessel integrity and patency is maintained throughout the body for its lifetime"
• Process of haemostasis

  1. Closely linked interaction between Blood vessel wall and Circulating platelets

  2. Blood coagulation factors

  3. Needs to be carefully controlled! So also important are:

  4. Coagulation inhibitors

  5. Fibrinolysis
• Primary Haemostasis

  Depends upon the response of the platelets and blood vessel wall to the injury. When the small blood vessels are injured, blood platelets adhere and aggregate at the site of injury, reducing and finally arresting bleeding.
• Secondary Haemostasis

  Starts when the cascade system of coagulation is activated by substances released at the time of blood vessel injury.
• Platelets

  • Made in the bone marrow from megakaryocytes
  • Production stimulated by thrombopoietin
  • Lifespan: 10 days
  • Normal count ~250 x 10^9/L (range: 150-400 x 10^9/L)
• Platelet glycoproteins important in platelet reactions

  • GPIa: binds to collagen
  • GPIb: binds to von Willebrand factor (VWF)
  • GPIIb/IIIa: binds to von Willebrand factor (VWF) and fibrinogen
• Von Willebrand Factor (VWF)

  • Large glycoprotein
  • Made in: endothelial cells – Weibel-Palade bodies, megakaryocytes – platelet a granules
  • Majority continuously secreted by endothelial cells
  • Minority stored in Weibel-Palade bodies
  • Secreted in response to: Stress, Exercise, Adrenaline, DDAVP
  • Two functions: Promotes platelet adhesion to subendothelium and each other, Carrier molecule for Factor VIII – protects it from premature destruction
• Platelet function

  • Primary haemostasis = formation of mechanical platelet plug
  • Provide a template for the coagulation cascade
• Primary haemostasis (1) VASOCONSTRICTION

  1. Of injured vessel and adjacent small arteries and arterioles

  2. Purpose: Initial slowing of blood to area of injury (prevent exsanguination)

  3. Allows contact activation of platelets and coagulation factors
• Primary haemostasis (2) PLATELET ADHESION

  1. Break in endothelial lining exposes subendothelial connective tissue (collagen)

  2. Platelets adhere via GPIa and GPIb receptors mediated by VWF
• Primary haemostasis (3) PLATELET ACTIVATION

  1. Prostaglandin synthesis – formation of thromboxane A2

  2. Triggers platelet granule release

  3. Potent vasoconstrictor

  4. ADP causes platelets to swell and aggregate
• Primary haemostasis (4) PLATELET AGGREGATION

  1. Platelet rolling in the direction of blood flow over exposed vWF

  2. GpIIa/IIIb binds to fibrinogen

  3. Forms inter-platelet bridges

  4. Promotes growth of haemostatic plug and provide temporary control of bleeding
• Primary haemostasis (5) AMPLIFICATION

  1. Of platelet activation by thromboxane

  2. Positive feedback loop
• PFA-100

  In vitro system for measuring platelet-VWF function
• PFA-100


  • Attempts to reproduce primary haemostasis
  • Aspirates a blood sample under constant vacuum from the sample reservoir through a capillary and a microscopic aperture cut in a membrane coated with collagen and adrenaline or ADP
  • Time to full occlusion of the aperture = 'closure time'
  • Prolonged in VWD and other disorders of platelet function
• PFA-100 components

  • Trigger solution container
  • Soft keys
  • LCD screen
  • Built-In Printer
  • Carousel
  • Cassette
  • Test Cartridge
• Platelet Aggregation Studies

  Measure the fall in light absorbance in platelet-rich plasma as platelets aggregate
• Agents used in primary platelet aggregation

  • ADP
  • Collagen
  • Ristocetin
  • Arachidonic acid
  • Adrenaline
• Secondary platelet response

  Due to agents released by the platelets themselves
• Pattern of response to each agent

  Helps make the diagnosis
• Flow cytometry

  Used to identify platelet glycoprotein defects
• Coagulation cascade

  Biological amplification system with few initiators and sequential activation by proteolysis of circulating precursor proteins (coagulation factor enzymes)
• Coagulation cascade

  1. Generates thrombin

  2. Converts soluble plasma fibrinogen to fibrin

  3. Forms a mesh over platelet aggregates

  4. Stable haemostatic plug
• Coagulation factors


  • Extrinsic Xase (VIIa, TF, PL, Ca2+)
  • Intrinsic Xase (IXa, VIIIa, PL, Ca2+)
  • Prothrombinase (Xa, Va, PL, Ca2+)
• Clotting cascade

  1. Prothrombin (FII)

  2. Thrombin (FIIa)

  3. FXa + FVa

  4. Fibrinogen (FI)

  5. Fibrin
• Clotting cascade - Initiation

  Small amounts of thrombin generated
• Clotting cascade - Amplification

  Second, million times larger, burst of thrombin production
• Tissue factor (TF)

  • Expressed constitutively by vascular adventitia, smooth muscle and epidermal cells
  • Membrane-bound
  • Forms a protective envelope around all blood vessels
  • After vascular injury, plasma FVII can bind to TF to form the extrinsic factor Xase complex
  • Activates FIX and FX
• Extrinsic Pathway

  1. Tissue Factor

  2. VII

  3. VIIa

  4. X

  5. Xa

  6. IX

  7. IXa

  8. Va

  9. VIIIa

  10. Common Pathway

  11. Thrombin

  12. Extrinsic Xase
• Intrinsic Pathway

  1. XII

  2. XIIa

  3. XI

  4. XIa

  5. IX

  6. IXa

  7. VIIIa

  8. X

  9. Xa

  10. Common Pathway

  11. Va

  12. Thrombin

  13. Intrinsic Xase

  14. Prothrombinase

  15. Thrombin
• Fibrin formation

  1. Cross-linked fibrin forms a mesh within the platelet plug

  2. Traps red cells

  3. Fused platelets degranulate and autolyse

  4. Solid mass of cross-linked fibrin within a few hours
• Clot retraction

  1. Edges of the blood vessel wall at the point of injury are slowly brought together again to repair the damage

  2. Mediated by GPIIb/IIIa receptors

  3. Link cytoplasmic actin to fibrinogen
• Physiological limitation of coagulation

  • Coagulation factor inhibitors
  • Blood flow
  • Fibrinolysis
• TFPI (Tissue factor pathway inhibitor)

  • Made in endothelial cells
  • Present in plasma and platelets
  • Inhibits Xa, VIIa, TF
• Antithrombin

  • Circulating inhibitor
  • Inhibits the serine proteases: IIa (thrombin), IXa, Xa and XIa
• Protein C and Protein S

  Inhibit the cofactors Va and VIIIa
• Blood flow

  • Rapidly dilutes and disperses activated factors at the periphery of damaged area before fibrin formation can occur
  • Activated factors destroyed by liver parenchymal cells
  • Particulate matter removed by liver Kupffer cells
• Fibrinolysis

  Normal haemostatic response to vascular injury
• Prothrombin Time (PT)

  Measures: Extrinsic pathway - VII, X, V, prothrombin and fibrinogen
• Prothrombin Time (PT)

  • Normal: 10-14 s
  • Tissue thromboplastin
  • Calcium
  • Citrated plasma