Blood coagulation

1. Blood coagulation
Dr. Sai Sailesh Kumar G
Associate Professor
Department of Physiology
RDGMC
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2. Learning objectives
 List the coagulation factors
 Describe the steps involved in the mechanism
of clot formation
 Describe the process of fibrinolysis
 List the disorders affecting blood coagulation
 List the anticoagulants and describe their
mechanism of action
 Describe the screening tests of hemostasis
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3. Introduction
 More than 50 substances were found in blood that
affect blood coagulation
 Promote coagulation – pro coagulants
 Inhibit coagulation – anti coagulants
 Blood coagulates or not – balance between these
two substances
 Normally anticoagulants predominate
 So blood does not coagulate while circulating
 When blood vessel ruptured, pro coagulants are
activated and over ride anticoagulants
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4. General mechanism
 Clotting takes place in three essential steps
1. In response to rupture of blood vessel, complex
cascade of chemical reactions occurs in the
blood involving clotting factors. The net result
is formation of prothrombin activator
2. Prothrombin activator catalyzes conversion of
prothrombin to thrombin
3. Thrombin acts as enzyme and converts
fibrinogen to fibrin that form clot
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5. Prothrombin to thrombin
 Prothrombin activator is formed as a result of
rupture of blood vessel
 Prothrombin activator in the presence of sufficient
amounts of calcium, converts prothrombin to
thrombin
 Thrombin causes polymerization of fibrinogen
molecules into fibrin fibers within 10-15 seconds
 The rate limiting step in the blood coagulation is
formation of prothrombin activator
 Platelets also plays important role in conversion of
prothrombin to thrombin
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6. Prothrombin and thrombin
 Prothrombin is a plasma protein
 Molecular weight 68,700
 Unstable protein that can split into smaller compounds
 One of the compound is thrombin
 Thrombin molecular weight – 33,700
 Prothrombin is formed continuously in the liver
 It is used continuously through out the body for
coagulation
 If liver fails to produce prothrombin
 Plasma concentration of prothrombin decreases
 Defect in the blood coagulation
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7. Prothrombin and thrombin
 Vitamin K is required by the liver for normal
activation of prothrombin and few other clotting
factors
 Lack of vitamin k or presence of liver disease
that prevent normal prothrombin formation can
decrease the prothrombin to low levels
 Bleeding tendency increases
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8. Prothrombin to thrombin
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9. Fibrinogen and fibrin
 Fibrinogen is a high molecular weight protein
 MW = 340,000
 Fibrinogen is formed in the liver
 Because of high molecular weight, a small amount
of fibrinogen leaks into the interstitial fluid
 When there is alter in the permeability of capillaries
in pathological conditions
 Large amounts of fibrinogen leaks into tissue fluid
and causes it to clot in the same way as blood
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10. Fibrinogen to fibrin
 Thrombin is a protein enzyme
 Thrombin has weak proteolytic capabilities
 Thrombin acts on fibrinogen
 Removes four low molecular weight peptides
 Forms fibrin monomers
 Fibrin has capability to polymerize with other
fibrin monomer
 Forms long fibrin fibers
 Constitute the reticulum of blood clot
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11. Fibrinogen to fibrin
 Early phase of polymerization
 fibrin monomers are held with weak
noncovalant hydrogen bonds
 Clot is weak
 Can be broken with ease
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12. Fibrin stabilizing factor
 late phase of polymerization
 platelets entrapped in clot releases fibrin
stabilizing factor
 Fibrin stabilizing factor is activated by thrombin
 Fibrin stabilizing factor cause covalent bonds
between fibrin monomers
 Also forms multiple cross linkages between
adjacent fibrin fibers
 Strengthens the fibrin meshwork
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13. Blood clot
 Composed of meshwork of fibrin fibers that
entraps
1. Blood cells
2. Platelets
3. plasma
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14. Clot retraction
 With in few minutes after a clot is formed
 It begins to contract
 Express most of the fluid from the clot with in
20-60 minutes
 The fluid expressed is called serum ( fibrinogen
and most of other clotting factors removed)
 Plasma with out fibrinogen and other clotting
factors is serum
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15. Clot retraction
 Will serum clots?
 Serum can not clot
 It lacks clotting factors
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16. Clot retraction
 Platelets are necessary for clot retraction
 Failure of clot retraction – decreased platelet count
 Platelets release thrombaxane A2 - clot retraction
 Platelets directly cause clot retraction
 Platelets activates thrombosthenin, actin, myosin
molecules
 These are contractile proteins and cause strong
contraction of platelets so that they attach firm to fibrin
 This action compress the fibrin meshwork into smaller
mass
 As clot retracts, the edges of the broken blood vessel
pulled together
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17. Positive feedback of clot
formation
 Clot initiate positive feedback to promote more
clotting
 Prothrombin is converted into thrombin
 Thrombin has proteolytic activity and acts on
prothrombin itself
 Forms and more thrombin
 Blood clot continues to grow until blood leakage
ceases
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18. Initiation of coagulation
1. Trauma to the vascular wall and adjacent tissues
2. Trauma to the blood
3. Contact of blood with damaged endothelial cells or
collagen
 All the above causes formation of prothrombin
activator
1. Extrinsic pathway – begins with trauma to vessel
wall and surrounding tissues
2. Intrinsic pathway – begins in the blood
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19. Initiation of coagulation
 Both intrinsic and extrinsic pathways, a series of
different proteins called blood clotting factors
plays a major role
 Most of the clotting factors are inactive forms of
proteolytic enzymes
 When converted to active forms, their enzymatic
actions cause cascade reactions of the clotting
process
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20. Clotting factors
 They are designated by Roman numerals
 To indicate the activated form of the factor, a
small letter “a” is added after the Roman
numeral
 such as factor VIIa indicates the activated state
of Factor VII
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22. Extrinsic pathway
 Begins with traumatized vascular wall
 or traumatized extra vascular tissues
 that come in contact with blood
 This leads to following steps
1. Release of tissue factor
2. Activation of factor X
3. Effect of Xa to form prothrombin activator
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23. Release of tissue factor
 Traumatized tissue releases several factors
 Tissue factor or tissue thromboplastin
 This is composed of
1. Phospholipids (from membranes/ platelets)
2. Lipoprotein complex – proteolytic enzyme
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24. Activation of factor X
1. Activated factor X combines with phospholipids
+ factor V
2. Formation of prothrombin activator
3. With in a sec in the presence of calcium,
prothrombin split into thrombin
4. Thrombin further activates factor V
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26. Intrinsic pathway
 Begin with trauma to the blood
 Exposure of blood to collagen
1. Blood trauma causes activation of factor XII
and release of platelet phospholipids
2. Activation of factor XI
3. Activation of factor IX
4. Activation of factor X
5. Formation of prothrombin activator
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27. Blood trauma
 Activates factor XII and platelets
 When factor XII comes in contact with collagen
or with a wettable surface such as glass in is
activated (XIIa)
 Activated platelets releases platelet
phospholipids
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28. Role of factor VIII
 Activates factor X
 Deficiency of factor VIII or platelets – this step is
deficient
 Hemophilia – factor VIII missing
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30. Role of calcium ions
 Except first two steps in intrinsic pathway,
 Calcium ions are required for rest all steps in both
pathways
 Lack of calcium – either pathway does not occur
 Blood when removed form body, can be stored and
prevent clotting by decreasing the calcium ion
concentration
 Common anti coagulant used- ACD, CPD
 ACD – acid citrate dextrose
 CPD – citrate phosphate dextrose
 Citrate ions – deionize calcium
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31. Role of calcium ions
 Intrinsic and extrinsic pathways occurs
simultaneously
 tissue factor initiates extrinsic pathway
 Contact of factor XII and platelets with collagen
in the vascular wall initiates intrinsic pathway
 Extrinsic pathway can be explosive
 Clotting occurs in 15 seconds
 Intrinsic pathway is slower
 Clotting occurs in 1-6 minutes
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32. Excessive bleeding in humans
 Vitamin K deficiency
 Hemophilia
 Thrombocytopenia (platelet deficiency)
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33. Vitamin K
 Vitamin K is essential in the formation of clotting
factors II,VII,IX,X and protein C
 Vitamin K add carboxyl group to glutamic acid
residues
 After this step, vitamin k is oxidized and become
inactive
 Vitamin k epoxide reductase complex 1 converts
vitamin k again to its active form
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34. Vitamin K deficiency
 Absence of vitamin K in diet
 Absence of intestinal bacterial flora
 Obstructive jaundice
 Gastro intestinal disease (poor absorption)
1. Vitamin K is injected to the surgical patients with liver
disease or obstructive bile ducts before the surgical
procedure
2. Effective only when half of the liver cells are functional
3. Produce sufficient clotting factors to prevent excess
bleeding
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35. Hemophilia
 Bleeding disease
 Occurs almost exclusively in males
 85% cases, deficiency of factor VIII – Hemophilia A or
classical hemophilia
 15% cases, deficiency of factor IX
 Both these factors are transmitted genetically by the way of
female chromosome
 Women will almost never have hemophilia
 If one of the x chromosome is deficient, she acts as carrier
(other x chromosome have appropriate genes)
 Transmits disease to half of her male offspring
 Transmits carrier state to half of her female offspring
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36. Hemophilia
 Severity depends on genetic deficiency
 small injury also cause bleeding
 Bleeding lasts for several days
 Example: tooth extraction
 Factor VIII has large and small components
 Smaller component is important for intrinsic
pathway
 Smaller component deficiency – classical
hemophilia
 Large component deficiency – von willebrand
disease
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37. Hemophilia
 Therapy – injection of factor VIII
 Cost is high as it is collected from human blood
and only extremely small quantities
 Hemophilia is sometimes referred to as “the
royal disease,” because it affected the royal
families of England, Germany, Russia and Spain
in the 19th and 20th centuries.
 Queen Victoria of England, who ruled from 1837-
1901, is believed to have been the carrier of
hemophilia B, or factor IX deficiency.
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38. Anticoagulants for clinical use
 Thromboembolism
 desired to delay the coagulation
 Most effective anticoagulants for this purpose
1. Heparin – inactivates thrombin and factor Xa
2. Coumarins (Warfarin) – inhibits vitamin K
3. Anti platelet agents – Asprin
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39. Heparin
 Powerful anticoagulant
 produced by many different cells of the body
 Large quantities are produced by basophils and
mast cells
 Concentration in blood is low
 Injection of 0.5-1 mg/kg of body weight prolongs
clotting time about 30 or more minutes
 Immediate action and Action last for 1.5-4 hours
 Destroyed by heparinase
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40. Coumarins
 Warfarin
 inhibits vitamin K epoxide reductase complex 1
 Decreased active vitamin K levels
 Factor II,VII,IX,X levels fall
 Action is not immediate
 After 12 hours of injection, coagulant activity
decreases 50%
 Normal coagulation results 1-3 days after
discontinuing the dose
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41. Prevention of blood coagulation
outside the body
 Blood collected in glass tubes clot in 6 minutes
(intrinsic pathway)
 Blood collected in silicon containers clot in one
hour or more
 Silicon surface prevents activation of platelets
and factor XII
 Important factors that initiate intrinsic pathway
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42. Prevention of blood coagulation
outside the body
 Heparin can be used to prevent coagulation of
blood outside the body as well inside the body
 Especially used in surgical procedures
 Blood must be passed through heart lung
machine
 Artificial kidney machine
 And back to the person
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43. Prevention of blood coagulation
outside the body
 Oxalate anticoagulants
 Precipitates calcium
 Forms calcium oxalates
 Decrease ionic calcium levels
 Blocks blood coagulation
 Oxalates are toxic
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44. Prevention of blood coagulation
outside the body
 Citrate anticoagulants
 Sodium citrate, ammonium citrate, potassium citrate
 Forms unionized calcium (deionizes calcium)
 Decrease ionic calcium levels
 Blocks blood coagulation
 Citrate anticoagulants are non toxic and can be
removed easily from the body by liver
 In case of liver disease, long lasting action of
citrates – depress calcium levels - tetany and death
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45. Blood coagulation tests
 Clotting time
 Blood is collected in a chemically clean glass tube
 Tip the tube back and forth about every 30 seconds
 Until the blood has clotted
 Normal value 6-10 minutes
 Clotting time varies widely, depending on the
method used
 No longer used in many clinics
 Measurement of clotting factors themselves are
made using sophisticated chemical procedures
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46. Blood coagulation tests
 prothrombin time
 Indication of concentration in the blood
 Blood removed from patient is oxalated
 Prothrombin can not convert to thrombin
 Then large excess of calcium ions and tissue factors
added the blood
 Excess calcium nullifies the effect of oxalates
 Tissue factor activates extrinsic pathway
 Prothrombin converts to thrombin
 The time required for coagulation – prothrombin time
 Normal time – 12 seconds
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48. THANK YOU
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