Transcript Lecture 17 - New CombinedPrimary and secondary hemostasis

HEMOSTASIS
Primary and Secondary Hemostasis
HEMOSTASIS
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Hemostasis
The process by which the body stops bleeding upon
injury and maintains blood in the fluid state in the
vascular compartment
 Process is rapid and localized
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HEMOSTASIS
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The primary players in hemostasis include
Blood vessels
 Platlets
 Plasma proteins
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Coagulation proteins – involved in clot formation
 Fibrinolysis – involved in clot dissolution
 Serine protease inhibitors
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Other minor players include
Kinin system
 Complement system
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HEMOSTASIS
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Defects
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In blood vessels, platlets or serum proteins can be
corrected by utilization of the other 2 players
In 2 of the 3 players results in pathologic bleeding
Blood Vessels
Platlets
Plasma Proteins
HEMOSTASIS
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Hemostasis can be divided into two stages
 Primary hemostasis
Response to vascular injury
 Formation of the “platelet plug” adhering to the endothelial wall
 Limits bleeding immediately
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Secondary Hemostasis
Results in formation of a stable clot
 Involves the enzymatic activation of coagulation proteins that
function to produce fibrin as a reinforcement of the platelet plug
 Gradually the stable plug will be dissolved by fibrinolysis
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FORMATION OF A STABLE PLUG
VASCULAR SYSTEM
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Smooth and continuous endothelial lining is designed to
facilitate blood flow
Intact endothelial cells inhibit platelet adherence and blood
coagulation
Injury to endothelial cells promotes localized clot formation
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Vasoconstriction
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Narrows the lumen of the vessel to minimize the loss of blood
Brings the hemostatic components of the blood (platelets and plasma
proteins) into closer proximity to the vessel wall
Enhances contact activation of platlets
 Von Willebrand factor
 Collagen fibers
 Platlet membrane glycoprotein Ib
Activated platlets enhance activation of coagulation proteins
PRIMARY HEMOSTASIS
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Platelets
Interact with injured vessel wall
 Interact with each other
 Produce the primary hemostatic plug
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Primary platelet plug
Fragile
 Can easily be dislodged from the vessel wall
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PLATELETS
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Platelets
Small, anucleated cytoplasmic fragments
 Released from megakaryocytes in the BM
 Megakaryocyte proliferation is stimulated by thrombopoietin
(TPO)
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Normal platlet count is 150-400 x 109/L
Survive 9-12 days
Nonviable or aged platelets removed by spleen & liver
2/3 of platelets circulate in the peripheral blood
1/3 are sequestered in the spleen
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Humoral factor
Produced primarily by liver, kidney, spleen, BM
Produced at a relatively constant rate
These 2 pools are in equilibrium and constantly exchanging
Spontaneous hemorrhaging occurs when platlet count gets below
10 x 109/L
PLATLETS
MATURE MAGAKARYOCYTE
PLATLET RELEASE
PLATLET FUNCTION
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Platlets function to
Provide negatively charged surface for factor X and
prothrombin activation
 Release substances that mediate vasoconstriction,
platlet aggregation, coagulation, and vascular repair
 Provide surface membrane proteins to attach to other
platlets, bind collagen, and subendothelium
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PLATELETS
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Are the primary defense against bleeding
Circulate in resting state
Have minimal interaction with other blood components or the
vessel wall
 Morphology of resting platelet is smooth, discoid
 When stimulated by endothelial damage, platlets become
activated and they
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Become round and ‘sticky’
Build a hemostatic plug
Provide reaction surface for proteins that make fibrin
Aid in wound healing
Platlet activation and plug formation involves
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Adhesion
Shape change
Secretion
Aggregation
FORMATION OF PRIMARY HEMOSTATIC
PLUG
PLATELETS AND
SECONDARY HEMOSTASIS
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Primary platelet plug is
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Unstable and easily dislodged
Secondary hemostasis
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Fibrin formation stabilizes and reinforces the platelet plug
Proteins interact to form fibrin assemble on negatively
charged membrane phospholipids of activated platelets
System mediated by many coagulation factors present in an
inactive form in blood.
Factors are assigned Roman numerals, I through XIII
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All are produced in the liver. The von Willebrand factor is also
produced in endothelial cells and megakaryocytes.
SECONDARY HEMOSTASIS
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Coagulation factors are divided into three
categories based on hemostatic function
Substrate –fibrinogen (Factor I), which is the main
substrate used to make fibrin
 Co-factors – accelerate enzymatic reactions
 Enzymes
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Coagulation factors are also classified by
physical properties
 Contact proteins
 Involved
in earliest phases of clotting
 Partially consumed during coagulation
 Found in serum
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Prothrombin Group
Vitamin-K Dependant Clotting Factors
 Most are found in serum
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SECONDARY HEMOSTASIS
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Fibrinogen Group
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Thrombin-Sensitive Clotting Factors
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Some drugs prevent clotting by acting as
antagonists to Vitamin K (Warfarin and
Coumadin)
All are acted upon by thrombin in the process of blood
coagulation
None found in serum
The cascade theory of blood coagulation
Involves a series of biochemical reactions
Transforms circulating substances into an insoluble
gel through conversion of fibrinogen to fibrin
Requires
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Plasma proteins
Phospholipids
calcium
CASCADE THEORY OF COAGULATION
Each coagulation factor is converted to an active
form by the preceeding factor in the cascade
 Calcium participates in some of the reactions as a
co-factor
 The blood coagulation cascade occurs on cell
surface membranes.
 The membrane localizes the reaction to the site of
injury
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SECONDARY HEMOSTASIS
Three different complexes assemble on the
phospholipid membrane
 The pathways for the formation of these
complexes are
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Intrinsic
 Extrinsic
 Common -Both intrinsic and extrinsic pathways
converge to share factors in the common pathway
 Both intrinsic and extrinsic pathways require
initiation
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Intrinsic - all factors involved in clot formation are in the
vascular compartment
 Extrinsic- is initiated when a tissue factor not found in
blood enters the vascular system
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COMPLEXES ON MEMBRANE
Extrinsic pathway
Intrinsic pathway
Common pathway
Fibrin formation
EXTRINSIC PATHWAY
INTRINSIC PATHWAY
COMMON PATHWAY
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Intrinsic and extrinsic pathways
Converge on the common pathway
 In the final steps thrombin converts fibrinogen to soluble
fibrin and the fibrin monomers are crosslinked to form a
stable fibrin polymer.
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COMMON PATHWAY
COAGULATION CASCADE
INHIBITION OF COAGULATION
Antithrombin (AT) is a potent physiologic
inhibitor of thrombin, and several other factors
involved in coagulation
 In the presence of heparin, the inactivation of
thrombin by AT is significantly increased
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INHIBITOR PATHWAY OF
COAGULATION
SUMMARY OF PRIMARY AND
SECONDARY HEMOSTASIS
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Sequence after vessel injury
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Vasoconstriction
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Platelet adhesion
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Controlled by vessel smooth muscle; enhanced by chemicals
secreted by platelets
Adhesion to exposed subendothelial connective tissue
Platelet aggregation
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Interaction and adhesion of platelets to one another to form initial
plug at injury site
SUMMARY OF PRIMARY AND
SECONDARY HEMOSTASIS
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Sequence cont’d
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Fibrin-platelet plug
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Coagulation factors interact on platelet surface to produce fibrin;
fibrin-platelet plug then forms at site of vessel injury
Fibrin stabilization
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Fibrin clot must be stabilized by F-XIIIa
FIBRINOLYSIS
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Activation of coagulation also activates fibrin lysis
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Fibrinolysis results in a gradual enzymatic cleavage of
fibrin to soluble fragments
Due to the activity of plasmin which is responsible for degradation
of fibrin
 Limits the extent of the hemostatic process
 Reestablishes normal blood flow
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PLASMIN ACTION
FDP= fibrin
degradation
products
KININ AND COMPLEMENT SYSTEMS
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The kinin system is also activated by both
coagulation and fibrinolytic systems
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The kinin system is important in inflammation,
vascular permeability, and chemotaxis
The complement system is activated by plasmin
INTERRELATIONSHIP OF COAGULATION,
FIBRINOLYTIC, KININ, AND COMPLEMENT
SYSTEMS
HEMOSTATIC BALANCE
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The regulation of hemostatic and fibrinolytic
processes is dynamic
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Balance between
Pro- and anti-hemostatic mediators
 Pro- and anti-fibrinolytic mediators
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Balance can be upset if any components are
Inadequate
 Excessive
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Development of thrombi
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Excessive local or systemic activation of coagulation
Sustained bleeding
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Excessive local or systemic fibrinolytic activity
HEMOSTATIC BALANCE
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When hemostasis is delayed
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Either platelet disorder or a coagulation defect
Bleeding episode may be prolonged
 Imbalance created between
 An abnormally slow hemostatic rate
 A normal rate of fibrinolysis
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An inadequate fibrinolytic response
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May retard lysis of a thrombus and even contribute to its
extension
BALANCE OF CLOTTING AND
FIBRINOLYSIS
DIAGNOSIS OF BLEEDING PROBLEMS
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Questions to address:
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Is a bleeding tendency present?
Is the condition familial or acquired?
Is the disorder one affecting
Primary hemostasis (platelet or blood vessel wall problems)
 Secondary hemostasis (coagulation problems)
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Is there another disorder present that could be the cause of
or might exacerbate any bleeding tendency?
Principal Presentations of bleeding disorders
Easy bruising
 Spontaneous bleeding from mucous membranes
 Menorrhagia – excessive bleeding during menstruation
 Excessive bleeding after trauma
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LABORATORY EVALUATION OF
HEMOSTASIS
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Three different categories of disorders may be
found
Vascular and platlet disorders
 Coagulation factor deficiencies or specific inhibitors
 Fibrinolytic disorders
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Bleeding disorders present differently depending
upon the causative problem
Platlet disorders present as petechiae and bleeding
into mucous membranes because of failure to form
the platlet plug
 Patients with coagulation defects (includes those
with hemophilia) may develop deep spreading
hematomas and bleeding into the joints with evident
hematuria because of failure to reinforce the platlet
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LABORATORY EVALUATION
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Tests to differentiate between these include
Platlet count
 Peripheral blood smear evaluation
 Ivy bleeding time (N=2.5-9.5 min) or platlet function
analyzer (PFA)
 Prothrombin time (PT) – test contains
thromboplastin and calcium chloride and measures
measures the extrinsic and common pathways
(Normal=11-13 sec)
 Activated partial thromboplastin time (APTT) contact activators and a platlet substitute and
calcium chloride are added to measure the intrinsic
and common pathways (Normal usually 23-35 sec,
may vary depending upon analyzer used, reagents
used, and patient population)
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LABORATORY EVALUATION
Thrombin time (TT) – add thrombin and measure
the time required for thrombin to convert fibrinogen
to fibrin (common pathway) (N=15-22 sec)
 Mixing studies with PT and APTT abnormal results patient plasma is mixed with normal plasma to
distinguish between factor deficiencies and
coagulation inhibitors
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If assay is corrected – due to factor deficiency
 If partially corrected or uncorrected – due to inhibitor
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Coagulation factor assays
 Assays for fibrin degradation products – evidence of
fibrinolysis
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INHERITED QUALITATIVE
PLATELET DISORDERS
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Defects in platelet-vessel wall interaction
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Most common disorder is von Willebrand disease
 Deficiency or defect in plasma VWF
Defects in platelet-platelet interaction
 Defects of platelet secretion and signal transduction
 Abnormalities of platelet granules
 Defects in platlet coagulant activity
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LAB TESTS IN DISORDERS OF
PRIMARY HEMOSTASIS
Platlet
count
PT
APTT
Bleeding
time
Vascular disorder
Normal
Normal
Normal
Normal or
abnormal
Thrombocytopenia
Decreased Normal
Normal
Abnormal
Platlet
Dysfunction
Usually
Normal
Normal
Normal or
Abnormal
Normal
DRUGS THAT ALTER PLATELET
FUNCTION
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A variety of drugs alter platelet function
Some are used therapeutically for their antithrombotic
activity
 For others, abnormal platelet function is an unwanted side
effect
 Effect on platelet function
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Defined by an abnormality of bleeding time or platelet aggregation
 Aspirin
 Inhibits platlet aggregation
 Inhibits platlet secretion
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DISORDERS OF SECONDARY
HEMOSTASIS
Hereditary vs acquired
 Quantitative vs qualitative deficiencies
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Laboratory screening tests (PT, APTT)
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Does not differentiate quantitative vs qualitative disorders
Qualitative abnormal proteins will
Prolong clotting test
 Be recognized by immunologically-based procedures
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Activity assays
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Essential when screening for deficiencies
VON WILLEBRAND DISEASE
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Inherited hemorrhagic disorder
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Genetically and clinically heterogeneous
Caused by a deficiency/dysfunction of VWF
Most common hereditary bleeding disorder
VWF
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Multimeric blood protein
Performs two major roles in hemostasis
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Mediates adhesion of platelets to sites of vascular injury
Is a carrier protein for F-VIII
Inherited defects in VWF may
Interfere with biosynthetic processing or disrupt specific ligand
binding sites
 Cause bleeding by impairing either platelet adhesion or blood
clotting
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HEMOPHILIAS
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Hemophilia A
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Factor VIII Deficiency
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Hemophilia B
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Factor IX Deficiency
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Antihemophilic Factor
X-linked recessive disorder
Most common type of hemophilia
Christmas Factor (from family of first patients diagnosed with the
disorder)
X-linked recessive disorder
Hemophilia C
Factor XI Deficiency
Autosomal recessive disorder seen primarily in the Ashkenazi
Jewish population
 Symptoms range from mild to severe
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HEMOPHILIA
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Insufficient generation of thrombin by
F-IXa/VIIIa complex through the intrinsic pathway of
coagulation cascade
 Bleeding severity complicated by excessive fibrinolysis
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Clinical severity corresponds with level of factor
activity
 Severe hemophilia
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Factor coagulant activity <1% of normal
 Frequent spontaneous bleeding into joints and soft tissues
 Prolonged bleeding with trauma or surgery
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HEMOPHILIA
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Moderate hemophilia
Factor coagulant activity 1-5% of normal
 Occasional spontaneous bleeding
 Excessive bleeding with surgery or trauma
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Mild hemophilia
Factor coagulant activity >5% of normal
 Usually no spontaneous bleeding
 Excessive bleeding with surgery or trauma
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HEMOPHILIA –
CLINICAL PRESENTATION
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Readily diagnosed
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In severe disease and patients with prior family history
Diagnosis based on
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Unusual bleeding symptoms early in life
Age of first bleeding varies with severity of disease
Family history
Physical exam
Laboratory evaluation
HEMOPHILIA – TREATMENT
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Replacement of clotting factor to achieve hemostasis
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Annual cost for patient with severe hemophilia
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$20,000-100,000
Various products available
Plasma-derived low, intermediate and high purity products
 Plasma-derived ultrapure products
 Ultrapure recombinant products
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Replacement products – benefits vs risks
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Blood-born pathogens
Hepatitis A, B, C, G; HIV, Parvovirus B-19
 Thrombotic complications with some F-IX concentrations
 Development of alloantibody inhibitors
 Neutralize coagulant effects of replacement therapy
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COAGULATION SCREENING TESTS
IN CONGENITAL DEFICIENCIES
Platlet PT
count
APTT
PFA
TT
Congenital Deficiency
N
N
N
N
N
XIII, mild deficiency of any factor,
plasminogen activator inhibitor-1, α2 antiplasmin
N
A
N
N
N
VII – (extrinsic pathway)
N
N
A
N
N
XII, XI, IX, VIII, prekallikrein, high
molecular weight kininogen (intrinsic
pathway – includes hemophilias)
N
A
A
N
N
X, V, II (common pathway)
N
A
A
N
A
Fibrinogen (last part of common pathway)
N
N
A or N
A or N
N
Von Willibrands