Special Hematologic Tests

Special Hematologic Tests

• Peripheral Blood Smear
• RBC Indices
• Clotting Time
• Bleeding Time
• ESR
• Malarial Smear
• LE Preparation
• Coagulation Studies: PT, APTT
 Peripheral Blood Smear
• Glass microscope slide coated on 1 side w/ a thin layer
of venous blood
• Stained w/ Wright’s stain and examined under a
microscope
• Used to supplement information provided by
automated hematology analyzers (quantitative
information about blood cells and even ID specimens
w/ abnormal cells – “flag”/ warning signs)
• Evaluates RBC morphology, relative(estimate) and
differential counts of WBC and estimate of platelets,
presence of atypical, abnormal cells and parasites
 PBS Preparation

1.Place small amount of venous blood on a glass slide 4. Spreader slide is further pulled out, leaving a thin layer
of blood behind

2. A spreader slide has been positioned at an angle and

slowly drawn toward the drop of blood

End Result – a well-formed blood film dried for about 10

3. Spreader brought in contact w/ drop of blood and
mins
drawn away
PBS
 Evaluation of PBS

• RBC morphology: anisocytosis (size),

poikilocytosis (shape), hgb content (normo-
/hypochromia), immature RBCs  anemias
• WBC: estimate and differential counts,
atypical and immature cells  leukemias
• Platelet estimate  dengue, ITP
• Blood parasites: malaria, filaria
RBC
WBC
Malaria in PBS
Several red blood cells have ring stages inside
them. Close to the center there is a schizont and
on the left a trophozoite.
 Red Blood Cell Indices
• MCV (Mean corpuscular volume): average size of an RBC
MCV = Hct x 10 Normal range: 80-100 fl
RBC

• MCH (Mean corpuscular hemoglobin): average amount of Hb per RBC

MCH = Hb x 10 Normal range: 27-31 pg/cell
RBC

• MCHC (Mean corpuscular hemoglobin concentration): average

concentration of Hb per unit volume of RBCs
MCHC = Hb x 100 Normal range: 32-36 g/dl
Hct
 Erythrocyte Sedimentation Rate
• Rate at which rbcs settle and is
measured as the number of
millimeters of clear plasma present at
the top of the column after one hour
(mm/hr)
• Non-specific measure of inflammation
• Anticoagulated blood placed in upright
tube (Westergren or Wintrobe tube)
• governed by balance between pro-
sedimentation factors (fibrinogen) and
factors resisting sedimentation, namely
negative charge of RBCs (zeta potential)
• In inflammatory process, high
proportion of fibrinogen causes RBCs
stick to each other  rouleaux
formation (stack of coins)
- in lymphoproliferative disorders: Igs
secreted in high amounts
 ESR
• INCREASED
- pregnancy • DECREASED
- inflammation - polycythemia
- anemia - sickle cell anemia
- rheumatoid arthritis
- renal CA
- hereditary
- slanting the tube
spherocytosis
- presence of acute phase - CHF
reactants - excess anticoagulant
- increased levels of Ig
- plasma CHONs
- presence of
- wrong use of anticoagulant poikilocytes
- fibrogenemia
 ESR
WESTERGREN METHOD
WINTROBE METHOD

NORMAL VALUES: Adult male: 0-10 mm/hr

Adult female: 0-15 mm/hr
Children: 0-5 mm/hr
 Relation with CRP

• CRP (C-reactive Protein)

- an acute phase CHON produced by liver
during inflammatory reaction
- levels rise more quickly after inflammatory
or infective process begins
- direct measure of inflammatory response
 L.E. Preparation
• In systemic lupus erythematosus (SLE): L.E. cell in BM
and peripheral blood
• Caused by Ig (L.E. factor) present in patient’s plasma
• Reagents and Equipments
1. Wire sieve and pestle
2. Petri dish
3. Wintrobe sedimentation rate tubes, three or four.
4. Clean glass slides
5. Wright stain and buffer.
6. Disposable dropper
• Specimen: clotted whole blood, 10 ml
 Principle

• Clotted blood is allowed to sit at room temperature

for two hours.
• Forcing it through a sieve then macerates the clot.
• The trauma, produced when the blood is forced
through the strainer, causes extrusion of nuclei from
the polymorphonuclear cells.
• The L.E. factor present in the blood lyses the
nuclear material, which is then phagocytized by
other neutrophils. This forms the L.E.cell.
• Result: POSITIVE or NEGATIVE
L.E. Cell
 Clotting Time
• Time required for blood to form a clot in vitro
• Whole blood will form a solid clot when exposed
to a foreign surface such as a glass tube
• Rough measure of all intrinsic clotting factors in
the absence of tissue factors – variations are wide
and test sensitivity is limited
• Replaced by PTT (least effective in diagnosis of
actual hemostasis failure)
• Was used as a screening test to measure all
stages in intrinsic coagulation system and
monitor heparin therapy
 Clotting Time
• Time-consuming test, poor reproducibility
• Sensitive only to extreme factor deficiencies
• Conditions accompanied by ↑clotting time:
- Factors V, VII, VIII, IX, XI, XII deficiencies
- HDN
- Vitamin K deficiency
- Heparin therapy
- Presence of circulating Abs
- Anemia and leukemias
- Afibrinogenemia, Pneumonia
 Methods

• Capillary Method
• Slide Method
• Tube Method
 Tube Method (Lee and White)
• Equipments
- water bath, 37 oC
- glass test tube (10x75
mm)
- stopwatch
- plastic syringe
• Specimen: fresh whole
blood 4 ml
 Procedure
• Label 3 glass test tubes w/ patient’s name and number them 1, 2, 3
• Perform a clean atraumatic venipuncture and draw 4 ml of blood
• Remove needle from syringe and fill each of 3 tubes w/ 1 ml of blood
- last 1 ml may be discarded
- start stopwatch as soon as blood enters syringe
• Place 3 test tubes in 37oC water bath
• At exactly 3 mins, remove 1st tube from water bath and tilt gently to a
45 o angle to see whether blood has clotted
• If blood has not clotted , return to water bath and examine it to 30 sec
intervals
• After blood in the 1st tube has clotted, examine 2nd tube immediately
then examine 3rd tube
• Record time it took the blood in 3rd test tube to clot
• NORMAL RANGE: 5 -12 mins
 Factors Affecting Clotting Time
• Poor venipuncture technique – causes hemolysis of tissue
thromboplastin to mix w/ blood, shortens CT
• Bubbles entering syringe when blood is drawn –↑ rate of
coagulation
• Tilting all tubes in the same direction and angle
• Excessive agitation of blood (transfer of blood from syringe
to test tube)
• Dirty tubes
• Coagulation will be retarded by the ff:
- temp below 35oC
- temp above 45oC
- diameter of tubes (larger diameter, slower clot formation)
Clotting Time – Capillary Method

• Materials
- sterile disposable lancet
- stop watch
- dry glass capillary tube (2 mm diameter, 10
cm long)
- cotton swab
- 70% ethyl alcohol
Procedure
• Sterilize finger w/ 70% alcohol
and allow to dry naturally
• Prick finger and remove 1st drop
• Dip 1 end of capillary tube into
blood drop gently w/o pressure
(3-4 tubes used)
• Allow to fill tube w/ blood by
lowering fitted capillary tube
• After 30 secs, break a small piece
of capillary tube
• Repeat breaking at regular time
intervals till fibrin thread appears
at broken end of tube
• Record time interval between
pricking finger and 1st appearance
of fibrin thread at broken ends of
tube
Clotting Time: Slide Method
 Bleeding Time
• Useful tool to test for platelet plug and capillary
integrity
• Occasionally requested for patients for surgery
• Depends on efficiency of tissue fluid accelerating
coagulation process, capillary function, # of
platelets present and their ability to form a
platelet plug
• Prolonged: platelet count <50,000/ul and in
platelet dysfunction
• Used as a screening test for defects of primary
hemostasis
 Methods

• Duke method
• Ivy method
• Mielke method
• Simplate or Surgicutt
method
 Duke Method
• A standardized puncture of
the ear lobe is made.
• The length of time required for
bleeding to cease while the
blood is being blotted every 30
seconds is recorded.
• A lancet is used to make the
puncture.
• No repeat testing is allowed
due to space.
• Causes apprehension in the
patient.
• This test method is the easiest
to perform, but is the least
standardized and has the
worst precision and accuracy.
Ivy Method
• A blood pressure cuff is used to
maintain constant pressure within
the capillaries to help standardize
the procedure.
• The cuff is inflated to 40 mm Hg on
the upper arm to control capillary
tone and to improve the sensitivity
and reproducibility.
• The forearm is the bleeding time site
used.
• A sterile, disposable blood lancet is
used and the length of time required
for bleeding to cease is recorded.
• The greatest source of variation in
this test is largely due to difficulty in
performing a standardized puncture.
This usually leads to erroneously low
results.
 Simplate/ Surgicutt Method

• Modification of the Ivy Method

• The first bleeding time device introduced
was the Simplate.
• The Simplate device has a trigger and
spring method for the blade. The blade
has a depth of 1.0 mm and a width of 5.0
mm.
• Another brand name is the Surgicutt.
 Overview of Procedure
• Select a site on the patient’s forearm approximately 3 fingers widths
below the bend in the elbow that is free of visible subcutaneous veins.
• Avoid surface veins, scars, bruises and edematous areas. Apply the blood
pressure cuff and inflate to 40 mm Hg.
• Perform the incision after the cuff has been in place for 30-60 seconds.
• Apply the bleeding time device to the arm in a horizontal position. Do
NOT apply undue pressure as this will falsely increase the depth of the
incision. Start the stop watch as soon as the incision is made.
• The blood is wicked onto the filter paper at 30 second intervals taking
great care that the wound is NOT directly touched with the filter paper.
• Once blood no longer stains the filter paper the timing is stopped. The
bleeding time is reported out to the nearest 30 seconds.
• If bleeding continues for more than 15 minutes, the procedure should be
discontinued, and pressure should be applied to the wound sites. The
bleeding time should be repeated on the other arm. If bleeding has again
not ceased within 15 minutes, the results are reported as greater than 15
minutes.
Simplate/ Surgicutt Method
 Simplate/ Surgicutt Method
• Advantages of this method • Disadvantages of this
include: method include:
– Instrument is a sterile, – Slight scarring can occur so
standardized, easy to use patient should be informed.
device that makes a uniform
incision.
– Instrument is a spring
activated surgical steel blade
which is housed in a plastic
unit. This eliminates
variability of blade incision.
– This method is the most
standardized method of all
the bleeding time
procedures.
– Inexpensive
 Mielke Method
• Modification of the Ivy Method.
• A Bard-Parker or similar disposable
blade is used, along with a
rectangular polystyrene or plastic
template that contains a
standardized slit.
• The blade is placed in a special
handle containing a gauge in order
to standardize the depth of the
incision.
• The same procedure as described
for the Surgicutt method is
employed.
 Mielke Method
• Advantages of this • Disadvantages of this
method include: method include:
– That the surgical incision – Patient apprehension, due
more closely approximates to unconcealed scalpel.
the patient’s hemostatic – Small scars might form.
response to surgery, when
compared to the puncture
in the Ivy Method.
– The depth of the incision
can be controlled
 Results for Bleeding Time
• Adults:
– Simplate (template) method: 2-9.5 minutes
– Mielke (template) method: <10 minutes
– Duke (ear lobe) method: 1-3 minutes
• Newborn to 8 years (Mielke [modified] method):
3.4 +/- 1.3 minutes
• Children 8 to 18 years (Mielke [modified] method):
2.8 +/- 1.6 minutes
 Bleeding Time
• Results increased in:
– Anxiety
– Females
– Improper technique/operator error (eg, increased cuff pressure,
increased length or depth of cut, antecubital fossa cut instead of volar
surface of the arm)
• Results decreased in:
– Advancing age
– Repeating the test within 4 hours of first bleeding time
• Other factors affecting the results
– Handedness
– Ethnicity
– Serum triglycerides
– Skin fold thickness
– Social class
– Weight to height ratio
 Malarial Smear
• Mainstay in diagnosis of malaria
• Microscopic examination of blood utilizing
blood films
• More modern techniques (Ag tests and
polymerase chain reaction/ PCR) have been
used but not in endemic areas
• Most economic, preferred and reliable tool in
the diagnosis for malaria
• Uses 2 types of smears: thick and thin
 Malarial Smear
• Remains the gold standard for diagnosis
Giemsa stain
- distinguishes between species and life cycle stages
- parasitemia is quantifiable
• Threshold of detection
- Thin film: 100 parasites/tl
- Thick film: 5-20 parasites/tk
• Requirements:
- equipment, training, reagents, supervision
• Simple, inexpensive yet labor-intensive
• Accuracy depends on microscopist’s skills
 Interpreting Thick and Thin
Smears
• THICK FILM • THIN FILM
- lysed RBCs - fixed RBCs, single layer
- larger volume - smaller volume
- 0.25 ul blood/100 fields - 0.005 ul blood/ 100
- blood elements more fields
concentrated - good species
- good screening test differentiation
- negative or positive - requires more time to
- parasite density read
- more difficult to - low density infections
diagnose species can be missed
 Malarial Smear
• Prepare smears as soon as possible after
collecting venous blood to avoid:
- changes in parasite morphology
- staining characteristics
• Take care to avoid fixing the thick smear
- risk of fixing thick when thin is fixed w/
methanol if both smears on same slide
- let alcohol on finger dry to avoid fixing thick
smear
 Stages of Development
• SPOROZOITES: (“Sporos” – seeds) the
infectious form injected by the mosquitoes
• MEROZOITES: (“Meros” – piece) stage
invading RBCs
• TROPHOZOITES: (“Trophes” – nourishment)
the form multiplying in RBCs
• GAMETOCYTES: sexual stages
 Phases of the Life Cycle
SPOROGONY SCHIZOGONY
• Sexual phase of the • Asexual reproduction in the
parasite’s life cycle occurs human host
a. MACROGAMETOCYTES – a. Pre-erythrocytic/Exo-
female erythrocytic – liver
b. MICROGAMETOCYTES - b. Erythrocytic – rbcs
male
(Malignant tertian)

FM VS OJ MZ
 Morphology of malaria parasites.
P. vivax P. ovale P. malariae P. falciparum P. knowlesi COLUMN 1 (left to right): Plasmodium vivax (note enlarged infected
RBCs). (1) Early trophozoite (ring form) (note one RBC contains 2
rings—not that uncommon); (2) older ring, note ameboid nature of
rings; (3) late trophozoite with Schüffner’s dots (note enlarged RBC);
(4) developing schizont; (5) mature schizont with 18 merozoites and
clumped pigment; (6) microgametocyte with dispersed chromatin.
COLUMN 2: Plasmodium ovale (note enlarged infected RBCs). (1)
Early trophozoite (ring form) with Schüffner’s dots (RBC has
fimbriated edges); (2) early trophozoite (note enlarged RBC,
Schüffner’s dots, and RBC oval in shape); (3) late trophozoite in RBC
with fimbriated edges; (4) developing schizont with irregular-shaped
RBC; (5) mature schizont with 8 merozoites arranged irregularly; (6)
microgametocyte with dispersed chromatin.
COLUMN 3: Plasmodium malariae (note normal or smaller than
normal infected RBCs). (1) Early trophozoite (ring form); (2) early
trophozoite with thick cytoplasm; (3) late trophozoite (band form);
(4) developing schizont; (5) mature schizont with 9 merozoites
arranged in a rosette; (6) microgametocyte with compact
chromatin.
COLUMN 4: Plasmodium falciparum. (1) Early trophozoites (the
rings are in the headphone configuration with double chromatin
dots); (2) early trophozoite (accolé or appliqué form); (3) early
trophozoites (note the multiple rings/cell); (4) late trophozoite with
larger ring (accolé or appliqué form); (5) crescent-shaped
gametocyte; (6) crescent-shaped gametocyte.
COLUMN 5: Plasmodium knowlesi—with the exception of
image 5, these were photographed at a higher magnification (note
normal or smaller than normal infected RBCs). (1) Early trophozoite
(ring form); (2) early trophozoite with slim band form; (3) late
trophozoite (band form); (4) developing schizont; (5) mature
schizont with merozoites arranged in a rosette; (6) microgametocyte
with dispersed chromatin. Note: Without the appliqué form,
Schüffner’s dots, multiple rings per cell, and other developing
stages, differentiation among the species can be very difficult. It is
obvious that the early rings of all five species can mimic one another
very easily. Remember: One set of negative blood films cannot rule
out a malaria infection. (From Garcia LS: Malaria Clin Lab
Med 30:93-129, 2010,with permission. Column 5 courtesy CDC.)
The morphology of malaria parasites.
Plasmodium vivax: 1, Early trophozoite (ring form). 2, Late
trophozoite with Schüffner’s dots (note enlarged red blood
cell). 3, Late trophozoite with ameboid cytoplasm (very typical of P.
vivax). 4, Late trophozoite with ameboid cytoplasm. 5, Mature schizont
with merozoites (18) and clumped pigment. 6, Microgametocyte with
dispersed chromatin. 7, Macrogametocyte with compact chromatin.
Plasmodium malariae: 1, Early trophozoite (ring form). 2,Early
trophozoite with thick cytoplasm. 3, Early trophozoite (band
form). 4, Late trophozoite (band form) with heavy pigment. 5,Mature
schizont with merozoites (9) arranged in rosette. 6, Microgametocyte
with dispersed chromatin. 7, Macrogametocyte with compact
chromatin.
Plasmodium ovale: 1, Early trophozoite (ring form) with Schüffner’s
dots. 2, Early trophozoite (note enlarged red blood cell). 3, Late
trophozoite in red blood cell with fimbriated edges. 4, Developing
schizont with irregularly shaped red blood cell. 5, Mature schizont with
merozoites (8) arranged irregularly. 6, Microgametocyte with dispersed
chromatin. 7,Macrogametocyte with compact chromatin.
Plasmodium falciparum: 1, Early trophozoite (accolé or appliqué
form). 2, Early trophozoite (one ring is in headphone
configuration/double chromatin dots). 3, Early trophozoite with
Maurer’s dots. 4, Late trophozoite with larger ring and Maurer’s
dots. 5, Mature schizont with merozoites (24). 6, Microgametocyte
with dispersed chromatin. 7, Macrogametocyte with compact
chromatin.
Note: Without the appliqué form, Schüffner’s dots, multiple rings/cell,
and other developing stages, differentiation among the species can be
difficult. It is obvious that the early rings of all four species can mimic
one another very easily. Remember: One set of negative blood films
cannot rule out a malarial infection. (Reprinted by permission of the
publisher from Garcia LS: Diagnostic medical parasitology, ed 5,
Washington, DC, 2007, Copyright by American Society for
Microbiology.)
Schuffner’s Dot Schuffner’s Dot
COAGULATION DISORDERS
HEMOSTASIS
 Complex process: changes blood from a fluid to a solid
state
 multiple components of the blood clotting system are
activated in response to BV injury to control bleeding
 Composed of 4 events:
1. Primary hemostasis
2. Secondary hemostasis
3. Fibrin clot formation and stabilization
4. Inhibition of coagulation
PRIMARY HEMOSTASIS:
Vasoconstriction and Platelet Plug Formation
 The key component of primary hemostasis is the platelet.
 Triggered by injury to the vessel wall, exposing subendothelial
collagen.
 Vasoconstriction occurs at the site of injury to reduce blood flow.
 Adhesion: vWf adheres platelets to exposed subendothelial
collagen via the platelet receptor glycoprotein Ib (GPIb). Platelets
also adhere directly to collagen by other receptors.
 Aggregation: Platelets aggregate with each other with the help of
fibrinogen that binds to activated glycoprotein IIb-IIIa (GPIIb /
IIIa), forming a platelet plug. Platelet aggregates also provide the
phospholipid surface necessary for coagulation factor activation.
PRIMARY HEMOSTASIS
 Characterized by vascular contraction, platelet
adhesion and formation of a soft aggregate plug.
 Begins immediately after endothelial disruption.
 Injury causes temporary local contraction of vascular
smooth muscle. Vasoconstriction slows blood flow,
enhancing platelet adhesion and activation.
 Short lived: The immediate post injury vascular
constriction abates quickly.
 If flow is allowed to increase, the soft plug could be
sheared from the injured surface, possibly creating
emboli.
PRIMARY HEMOSTASIS
SECONDARY HEMOSTASIS: Activation of
Coagulation Factors and Generation of
Thrombin
 Responsible for stabilizing the soft clot and maintaining
vasoconstriction.
 Vasoconstriction is maintained by platelet secretion of serotonin,
prostaglandin and thromboxane.
 The soft plug is solidified through a complex interaction between
platelet membrane, enzymes, and coagulation factors.
 Goal: generation of sufficient thrombin to convert fibrinogen 
fibrin clot
 Involves activation of intrinsic, extrinsic and common
coagulation pathway factors
 PHASE IN 2o HEMOSTASIS FEATURES
 Tissue factor (TF) is released from injured tissue cells,
INITIATION OF COAGULATION endothelial cells and monocytes.
 TF and Factor VIIa form the TF / Factor VIIa complex.
 TF / Factor VIIa activates a small amount of Factor IX and X
to generate a small amount of thrombin.
 Factor XII (and other “contact” factors) play a minor role in
the activation of Factor XI.

 Thrombin activates Factor V to Va, Factor VIII to VIIIa and

PROPAGATION PHASE activates more platelets.
 Thrombin also activates FXI to FXIa.
 Additional Factor Xa is produced when TF / Factor VIIa
complex activates Factor IX. The resultant Factor IXa along
with Factor VIIIa forms the tenase complex which then
converts more Factor X to Xa.
 Factor Xa and Va along with calcium and a phospholipid (PL)
surface (activated platelets) form the prothrombinase
complex which converts prothrombin (Factor II) to large
amounts of thrombin (Factor IIa)
FIBRIN CLOT FORMATION
AND STABILIZATION

 Thrombin converts fibrinogen to fibrin

monomers which polymerize to form a soluble
clot.
 Thrombin then activates Factor XIII which cross-
links the fibrin monomers and stabilizes the clot.
COAGULATION FACTORS
 Produced in the liver and circulate in an inactive
form until coagulation cascade is initiated.
 Cascade occurs in steps.
 Completion of each step activates another
coagulation factor in a chain reaction leading to
conversion of fibrinogen  fibrin
Coagulation Factors
  I: Foolish  Fibrinogen
People  Prothrombin
MNEMONIC
 II :
 III : Try  Thromboplastin
 IV : Climbing  Calcium
 V: Long  Labile Factor (Proaccelerin)
 VI: Unassigned ( No longer used)
 VII : Slopes  Stable Factor
 VIII : After  Anti-Hemophilic Factor A
 IX : Christmas  Christmas Factor (AHF –
B)
 X : Some  Stuart-Prower Factor
 XI: People  Plasma Thromboplastin
Antecedent (AHF-C)
 XII: Have  Hageman Factor
 XIII: Fallen  Fibrin Stabilizing Factor
COAGULATION FACTORS
 Enzyme precursors or zymogens found in the plasma, along with
nonenzymatic cofactors and calcium
 Zymogens: substrates having no biologic activity until converted
by enzymes to active forms called serine proteases.
 Zymogens include II, VII, IX, X, XI, XII, and prekallikrein.
 Serine proteases are IIa, VIIa, IXa, Xa, XIa, Xlla, and kallikrein.
 Cofactors assist in the activation of zymogens and include V, VIII,
tissue factor, and high molecular weight kininogen (HMWK).
 In its active form, factor XIII is a transglutaminase.
 Fibrinogen: only substrate in the cascade that does not become
an activated enzyme.
THE COAGULATION GROUPS

 CONTACT GROUP
 PROTHROMBIN GROUP
 FIBRINOGEN GROUP
COAGULATION GROUP FEATURES

CONTACT GROUP a. Includes prekallikrein, HMWK, and factors XI and XII

b. Produced in the liver
c. Requires contact with a foreign surface for activation
(e.g., collagen in vivo, kaolin in vitro)
d. Functions of the contact group:
1) XII and prekallikrein reciprocally activate each other;
HMWK is a cofactor for this process.
2) All play a role in intrinsic coagulation activation.
3) Xlla, kallikrein, and HMWK play a role in the
inflammatory response, intrinsic fibrinolytic activation,
kinin formation, and activation of the complement
system.
PROTHROMBIN GROUP a. Includes factors II, VII, IX, and X
b. Produced in the liver
c. Vitamin K is required for synthesis of functional factors,
with calcium binding sites necessary for binding to
phospholipid (PF3) surfaces.
d. Causes for synthesis of nonfunctional factors:
1) Vitamin K deficiency or antibiotics that kill the intestinal
bacterial flora responsible for vitamin K synthesis
2) Oral anticoagulants (warfarin) that interfere with the
metabolism of vitamin K (vitamin K antagonists)
COAGULATION GROUP FEATURES
FIBRINOGEN GROUP a. Includes factors I, V, VIII, and XIII
b. Produced in the liver
c. Consumed in the clotting process
d. Factors I, V, and VIII serve as substrates for the fibrinolytic
enzyme plasmin.
e. Factors I and V are found in platelets.
f. Thrombin feedback on fibrinogen group factors depends on its
concentration.
1) Low thrombin levels activate factors V, VIII (positive feedback
on the cascade), and XIII and induce platelet aggregation.
2) When thrombin levels are high, thrombin binds to
thrombomodulin on the endothelial cell surface and activates
the protein C pathway.
3) Activated protein C and its cofactor, protein S, inhibit factors V
and VIII (negative feedback on the cascade).
g. Conversion of fibrinogen to fibrin is a three-step process.
1) Fibrinogen alpha and beta fibrinopeptides are cleaved by
thrombin, forming soluble fibrin monomers.
2) Fibrin monomers spontaneously polymerize, forming soluble
fibrin polymers. This is the endpoint for clot-based tests.
3) Clot stabilization occurs, requiring thrombin activation of XIII
and calcium.
 INTRINSIC
PTT
Heparin EXTRINSIC
12 Slower PT
11 More #s INSIDE it Coumadin (Warfarin)
10 Faster
Less EXTRA #
9
8
7
10
=
5 “CLOTS”
X
2
COMMON MATH PROBLEM = COMMON PATHWAY
I 1. Blood trauma causes (a) activation of
N Factor XII and (b) release of platelet
T phospholipids (containing platelet factor
3)
R
2. Activated Factor XII (XIIa) enzymatically
I activates Factor XI (XIa) which needs
N kininogen and is accelerated by
S prekallikrein
I 3. Factor XIa enzymatically activates Factor
IX to IXa
C
4. Factor IXa+ Factor VIIIa +platelet
phospholipids and factor 3 activate
P Factor X to Xa
A 5. Factor Xa+Factor V+ platelet or tissue
T phospholipid form the complex called
H prothrombin activator
W 6. The prothrombin activator in turn
A initiates within seconds the cleavage of
prothrombin to form thrombin
Y
 Extrinsic Pathway
1. Release of tissue factor: Traumatized
tissue releases a complex of several
factors called tissue factor or tissue
thromboplastin
2. Activation of Factor X: Tissue factor
further complexes with Factor VII and,
in the presence of calcium ions, acts
enzymatically on Factor X to form
activated Factor X
 Factor Xa forms prothrombin
activator— Factor Xa combines with
tissue or platelet phospholipids as well
as with Factor V to form the complex
called prothrombin activator
Thrombin and Fibrin
Formation
PHASE OF INHIBITION OF COAGULATION FEATURES

INHIBITION OF THROMBIN GENERATION  At the same time that a clot is being formed, the clotting
process also starts to shut itself off to limit the extent of the
thrombus formed.
 Thrombin binds to the membrane receptor thrombomodulin
and activates Protein C to Activated Protein C (APC).
 APC combines with its cofactor Protein S which then inhibits
Factors Va and VIIIa, slowing down the coagulation process.
 Thrombin bound to thrombomodulin becomes inactive and
can no longer activate procoagulant factors or platelets.
 The endogenous anticoagulant, antithrombin inhibits the
activity of thrombin as well as several of the other activated
factors, primarily Factor Xa.

FIBRINOLYSIS  Tissue plasminogen activator (t-PA) converts plasminogen to

plasmin which breaks down cross-linked fibrin to several fibrin
degradation products, the smallest of which is D-dimer.
 Thrombin activatable fibrinolysis inhibitor (TAFI) prevents the
formation of plasmin.
 Anti-plasmin and plasminogen activator inhibitor-1 (PAI-1)
inhibit plasmin and t-PA respectively
FIBRINOLYSIS
D-dimer
 a fibrin degradation product (or
FDP), a small protein fragment
present in the blood after a blood
clot is degraded by fibrinolysis
 is so named because it contains
two D fragments of the fibrin
protein joined by a cross-link.
 used to help rule out the presence
of an inappropriate blood clot
(thrombus) where D-dimer is ↑:
Deep vein thrombosis (DVT)
Pulmonary embolism (PE)
ABNORMALITIES IN
CLOTTING FACTORS
Clinical Features of Bleeding Disorders
Platelet Coagulation
disorders factor disorders

Site of bleeding Skin Deep in soft tissues

Mucous membranes (joints, muscles)
(epistaxis, gum,
vaginal, GI tract)
Petechiae Yes No
Ecchymoses (“bruises”) Small, superficial Large, deep
Hemarthrosis / muscle bleeding Extremely rare Common
Bleeding after cuts & scratches Yes No
Bleeding after surgery or trauma Immediate, Delayed (1-2 days),
usually mild often severe
COAGULATION FACTOR
DISORDERS
 Inherited bleeding  Acquired bleeding
disorders disorders
 Hemophilia A and B  Liver disease
 vonWillebrands  Vitamin K
disease deficiency/warfarin
 Other factor overdose
deficiencies  DIC
 ACQUIRED DISORDERS
 Usually characterized by multiple clotting
abnormalities
 Vitamin K deficiency results in impaired
synthesis of factors II, VII, IX, X and protein C
 Severe parenchymal liver disease can be
associated w/ hemorrhagic diathesis
 DIC produces a deficiency of multiple coagulation
factors
HEREDITARY DEFICIENCIES
 Sex-Linked Recessive Disorders
- Deficiencies of:
1. factor VIII (Hemophilia A)
2. factor IX (Christmas disease or Hemophilia B)
 Mostly others follow autosomal patterns of
transmission
HEREDITARY DEFICIENCIES
Deficiencies of Factor VIII-vWF
Complex
 Hemophilia A and von Willebrand disease:
- 2 of the most common inherited disorders of
bleeding
- caused by qualitative or quantitative defects
involving factor VIII-vWF complex
von Willebrand factor
 an adhesive plasma glycoprotein which performs its
hemostatic functions through binding to FVIII, to
platelets surface glycoproteins, and to constituents of
connective tissue
 VWF acts as a stabilizer of FVIII in the circulation.
 Synthesis in endothelium and megakaryocytes
 Forms large multimer
 Carrier of factor VIII
 Anchors platelets to subendothelium
 Bridge between platelets
von Willebrand Disease: Clinical
Features

 Inheritance - autosomal dominant

 Incidence - 1/10,000
 Clinical features - mucocutaneous bleeding
 Factor VIII – liver and kidney
 vWF – endothelial cells and megakaryocytes
 Both associate to form a complex in circulation
 come together and circulate in the plasma as a unit that serves to
promote clotting as well as platelet-vessel wall interactions necessary
to ensure hemostasis.
von Willebrand Disease
 vWF – determined through Ristocetin Agglutination
Test: immunoassay between vWF and platelet
membrane glycoprotein Ib
 1% in frequency
 Autosomal dominant but may have rare recessive
variant
 Spontaneous bleeding with normal platelet count
and prolonged bleeding time
 Can be due to reduced # or quality defects of vWF
Reduced Quantity of Circulating
vWF
 Type 1: Autosomal dominant
- approximately 70% of all cases, relatively mild
- (+) reduced penetrance and variable expressivity  varied
clinical manifestations
- poorly defined mutations
 Type 3: Autosomal recessive
- w/ extremely low levels of functioning vWF
- markedly affect stability of factor VIII
- severe manifestations: may resemble Hemophilia
- associated w/ deletions or frameshift mutations
Qualitative Defects in vWF
 Type 2 (2A): most common subtype
- autosomal dominant disorder
- due to missense mutations: vWF formed is
abnormal  defective multimer assembly
- most active forms of vWF (large and medium
multimers) are missing
- accounts for 25% of all cases w/ mild to moderate
bleeding
 Laboratory Evaluation of
von Willebrand disease
 Classification
 Type 1 Partial quantitative deficiency
 Type 2 Qualitative deficiency
 Type 3 Total quantitative deficiency
 Diagnostic tests:

vonWillebrand type
Assay 1 2 3

vWF  Normal 
vWF activity   
Multimer analysis Normal Normal Absent
Treatment of von Willebrand Disease
 Cryoprecipitate
 Source of fibrinogen, factor VIII
and VWF
 Only plasma fraction that consistently contains VWF multimers

 DDAVP (deamino-8-arginine vasopressin)

 plasma VWF levels by stimulating secretion from
endothelium
 Duration of response is variable
 Not generally used in type 2 disease

 Factor VIII concentrate (Intermediate purity)

 Virally inactivated product
Hemophilia A
 Most common hereditary disease
associated w/ serious bleeding
 Caused by reduced amount or
activity of factor VIII (cofactor for
factor IX in the activation of factor
X)
 Inherited as an X-linked recessive
trait: occurs in males, homozygous
females
- unusual inversion involving X
chromosome; point mutations
 Wide range of clinical severity
based degree of factor deficiency
 Normal plt, BT, PT, prolonged PTT
Hemophilia B
 Christmas Disease or
Factor IX deficiency
 Clinically
indistinguishable from
Hemophilia A (factors VIII
and IX work together to
activate factor X)
 Inherited as X-linked
recessive trait
 Variable clinical severity
 Hemophilia A and B
Hemophilia A Hemophilia B

Coagulation factor deficiency Factor VIII Factor IX

Inheritance X-linked X-linked

recessive recessive

Incidence 1/10,000 males 1/50,000 males

Severity Related to factor level

<1% - Severe - spontaneous bleeding
1-5% - Moderate - bleeding with mild injury
5-25% - Mild - bleeding with surgery or trauma

Complications Soft tissue bleeding

Hemophilia
Clinical manifestations (hemophilia A & B are
indistinguishable)
Hemarthrosis (most common)
Fixed joints
Soft tissue hematomas (e.g., muscle)
Muscle atrophy
Shortened tendons
Other sites of bleeding
Urinary tract
CNS, neck (may be life-threatening)
Prolonged bleeding after surgery or dental
extractions
Hemarthrosis (acute)
Hematoma
Treatment of Hemophilia A & B
 HEMOPHILIA A Treatment  HEMOPHILIA B Treatment
 Intermediate purity plasma  Agent
products
 High purity factor IX
 Virucidally treated
 Recombinant human factor IX
 May contain von Willebrand
factor
 High purity (monoclonal) plasma  Dose
products  Initial dose: 100U/kg
 Virucidally treated  Subsequent: 50U/kg every 24
 No functional von Willebrand hours
factor
 Recombinant factor VIII
 Virus free/No apparent risk
 No functional von Willebrand
factor
Complications of therapy
 Formation of inhibitors (antibodies)
 10-15% of severe hemophilia A patients
 1-2% of severe hemophilia B patients

 Viral infections
 Hepatitis B Human parvovirus
 Hepatitis C Hepatitis A
 HIV Other
INHIBITORS
 30% of people with hemophilia develop an
antibody to the clotting factor they are receiving
for treatment. These antibodies are known as
inhibitors.
 Long term management involves attempting to
eradicate inhibitors by administering high dose
FVIII (or FIX) in a process called immune
tolerance
FACTOR DEFICIENCIES
 HEMOPHILIA A (Classic Hemophilia)
80-85% of all Hemophiliacs
Deficiency of Factor VIII
Lab Results - Prolonged PTT
 HEMOPHILIA B (Christmas Disease)
 10-15% of all Hemophiliacs

 Deficiency of Factor IX

 Lab Test - Prolonged PTT

 VON WILLEBRAND’S DISEASE

 Deficiency of VWF & amount of Factor VIII

 Lab Results - Prolonged BT, PTT

ACQUIRED DISORDERS OF
COAGULATION
 Vitamin K deficiency
 Source of vitamin K Green vegetables
Synthesized by intestinal flora

 Required for synthesis Factors II, VII, IX ,X

Protein C and S

 Causes of deficiency Malnutrition

Biliary obstruction
Malabsorption
Antibiotic therapy

 Treatment Vitamin K
Fresh frozen plasma
Disseminated Intravascular
Coagulation (DIC)
 An acute, subacute, chronic thrombohemorrhagic
disease occurring as a secondary complication in a
variety of diseases
 Characterized by activation of coagulation sequence 
formation of microthrombi throughout microcirculation
in an unequal distribution
 Coagulapathy could be localized to specific organ or
tissue
DIC
 As a consequence of the thrombotic diathesis,
there is consumption of platelets, fibrin, and
coagulation factors and, secondarily, activation
of fibrinolytic mechanisms.
 Can present w/ s/s of tissue hypoxia and
infarction due to microthrombi or hemorrhagic
disorder related to depletion of hemostatic
elements (consumption coagulopathy)
Common clinical conditions associated with
DIC
Activation of both coagulation and fibrinolysis
Triggered by

 Sepsis  Obstetrical complications

 Amniotic fluid embolism
 Trauma  Abruptio placentae
 Head injury
 Vascular disorders
 Fat embolism
 Reaction to toxin (e.g. snake
 Malignancy venom, drugs)

 Immunologic disorders
 Severe allergic reaction
 Transplant rejection
 Disseminated Intravascular Coagulation (DIC)
Mechanism

Systemic activation
of coagulation

Intravascular Depletion of platelets

deposition of fibrin and coagulation factors

Thrombosis of small Bleeding

and midsize vessels
with organ failure
 Pathogenesis of DIC

Release of
thromboplastic
material into Consumption of
circulation coagulation factors;
presence of FDPs
Coagulation Fibrinolysis
 aPTT
 PT
Fibrinogen  TT
Thrombin Plasmin  Fibrinogen

Presence of plasmin
Fibrin  FDP
Monomers Fibrin(ogen)
Degradation Intravascular clot
Products  Platelets
Fibrin Schistocytes
Clot
(intravascular) Plasmin
Disseminated Intravascular Coagulation
Treatment approaches

 Treatment of underlying disorder

 Anticoagulation with heparin

 Platelet transfusion

 Fresh frozen plasma

 Coagulation inhibitor concentrate (ATIII)

 Liver Disease and Hemostasis
1. Decreased synthesis of II, VII, IX, X, XI, and
fibrinogen
2. Dietary Vitamin K deficiency (Inadequate intake or
malabsorption)
3. Dysfibrinogenemia
4. Enhanced fibrinolysis (Decreased alpha-2-
antiplasmin)
5. DIC
6. Thrombocytopenia due to hypersplenism
 Laboratory Evaluation of Bleeding
Overview
CBC and smear Platelet count Thrombocytopenia
RBC and platelet morphology TTP, DIC, etc.

Coagulation Prothrombin time Extrinsic/common pathways

Partial thromboplastin time Intrinsic/common pathways
Coagulation factor assays Specific factor deficiencies
50:50 mix Inhibitors (e.g., antibodies)
Fibrinogen assay Decreased fibrinogen
Thrombin time Qualitative/quantitative
fibrinogen defects
FDPs or D-dimer Fibrinolysis (DIC)

Platelet function von Willebrand factor vWD

Bleeding time In vivo test (non-specific)
Platelet function analyzer (PFA) Qualitative platelet disorders
and vWD
Platelet function tests Qualitative platelet disorders
Coagulation factor deficiencies
Summary
Sex-linked recessive
 Factors VIII and IX deficiencies cause bleeding
Prolonged PTT; PT normal

Autosomal recessive (rare)

 Factors II, V, VII, X, XI, fibrinogen deficiencies cause bleeding
Prolonged PT and/or PTT

 Factor XIII deficiency is associated with bleeding and

impaired wound healing
PT/ PTT normal; clot solubility abnormal

 Factor XII, prekallikrein, HMWK deficiencies

do not cause bleeding
BLEEDING & COAGULATION
DISORDERS
 Part 1:
BLEEDING DISORDERS
 HEMOSTASIS
 The body’s intrinsic ability to slow down or stop bleeding
 normal hemostasis involves a delicate balance between factors
that promote blood coagulation and thrombus stabilization and
factors that inhibit blood coagulation and promote thrombus
dissolution

FORMATION clot RESOLUTION

NORMAL HEMOSTATIC
RESPONSE
 Involves:  Evaluated by common
a. Blood vessel wall laboratory tests:
b. Platelets a. Platelet count
c. Clotting cascade b. Bleeding time
c. Prothrombin time (PT)
d. Partial thromboplastin
time (PTT)
 HEMOSTASIS

DEPENDENT UPON:
 Vessel Wall Integrity
 Adequate Numbers of Platelets
 Proper Functioning Platelets
 Adequate Levels of Clotting Factors
 Proper Function of Fibrinolytic Pathway
 NORMAL CLOTTING
 Response to vessel injury
 1. VASCULAR PHASE
 1. Vasoconstriction to reduce
 2. PLATELET PHASE blood flow

 3. COAGULATION PHASE  2. Platelet plug formation (von

Willebrand factor binds
 4. FIBRINOLYTIC PHASE damaged vessel and platelets)
 3. Activation of clotting cascade
with generation of fibrin clot
formation
 4. Fibrinolysis (clot breakdown)
Hemostasis Lab Tests
•CBC-Plt
•BT,(CT)
BV Injury •PT
Tissue Factor
•PTT
Neural

Blood Vessel Platelet Coagulation

Constriction Aggregation Cascade
Primary hemostatic plug

Reduced Platelet
Activation Fibrin
Blood flow formation
Plt Study
Morphology
Stable Hemostatic Plug
Function
Antibody
Stages of Hemostasis
 COAGULATION PATHWAY
Normally the components, called coagulation factors,
act like a row of dominoes toppling against each other
to create a chain reaction.
If one of the factors is missing, this chain reaction
cannot proceed.
LABORATORY EVALUATION

 PLATELET COUNT
 BLEEDING TIME (BT)
 PROTHROMBIN TIME (PT)
 PARTIAL THROMBOPLASTIN TIME (PTT)
 THROMBIN TIME (TT)
PLATELET COUNT

 NORMAL 150,000 - 450,000 cells/mm3

< 150,000 Thrombocytopenia

50,000 - 100,000 Mild Thrombocytopenia

< 50,000 Severe Thrombocytopenia

BLEEDING TIME

 Provides in vivo assessment of platelet

response to limited vascular injury
(PLATELET FUNCTION)

NORMAL VALUE
2-9 MINUTES
 PROTHROMBIN TIME (PT)
 Measures Effectiveness of the Extrinsic Pathway
and Common Pathway
 Sensitive to deficiencies of factors II, V, VII and
X
 To monitor WARFARIN (COUMADIN) therapy
 Mnemonic – PET

NORMAL VALUE
10-15 SECS
 PARTIAL THROMBOPLASTIN TIME
(PTT)
 Activated partial thromboplastin time (aPTT)
 Measures Effectiveness of the Intrinsic Pathway and
Common Pathway
 Sensitive to deficiencies of factor II, V, VIII, IX, X, XI, XII
 To monitor HEPARIN therapy: binds to antithrombin III
and ↑its ability to inactivate thrombin, factor Xa and
others
 Mnemonic – PITT
NORMAL VALUE
25-40 SECS
 THROMBIN TIME

 Time for Thrombin To Convert

Fibrinogen Fibrin
 A Measure of Fibrinolytic Pathway

NORMAL VALUE
< 20 SECS (15 – 19)
International Normalized Ratio (INR)

 The degree of prolongation of PT by warfarin

depends in the strength of the reagents used in
the lab w/c could vary among laboratories
 INR: created to standardize the variations and
allow for global application of anticoagulant
recommendations
 INR = patient PT/mean control PT
 MIXING STUDIES
 Also known as Circulating inhibitor screen
 Principle:
- Patient plasma is mixed or diluted w/ normal pooled plasma to
demonstrate factor levels
- the normal plasma provides the missing factor in the patient
plasma
- 50% activity is generally ample to produce a normal PT or APTT
- clotting time tends to increase w/ time and incubation due to
loss of labile factors so it is important to compare the results of
the patient’s diluted sample w/ the normal pooled plasma
- Correction indicates a factor deficiency. NO or PARTIAL
correction indicates a factor inhibitor
What if PT or APTT or BOTH are
prolonged?
So What Causes Bleeding
Disorders?
?

?
BLEEDING DISORDERS

 HEMORRGHAGIC DIATHESIS
 Excessive bleeding can result from:
a. Increased fragility of blood vessels (Vessel Defects)
b. Platelet deficiency or dysfunction (Platelet
Disorders)
c. Derangement of coagulation (Factor Deficiencies)
d. Combination of these (Other Disorders)
VESSEL DEFECTS
Infections
Drug Reactions
Scurvy and Ehlers-Danlos syndrome
Henoch-Schonlein purpura
Hereditary hemorrhagic telangiectasia
Amyloid infiltration of blood vessels
VESSEL DEFECTS
 Infections:  Drug reactions: may
meningococcemia, other induce bleeding w/o
forms of septicemia, causing
infective endocarditis thrombocytopenia
and rickettsioses - mediated by drug-
- presumably due to induced Abs and
microbial damage to deposition of immune
microvasculature complexes in vessel wall
(vasculitis) or DIC  hypersensitivity
vasculitis
 VESSEL DEFECTS
 Henoch-Schonlein
purpura (IgA vasculitis)
 Scurvy and Ehlers-Danlos - an idiopathic, systemic
syndrome hypersensitivity disease
- associated w/ - characterized by
microvascular bleeding purpuric rash, colicky
from impaired formation abdominal pain,
of collagens needed to polyarthralgia, AGN
support vessel walls
- result from deposition
of circulating immune
complexes
VESSEL DEFECTS

 Hereditary hemorrhagic telangiectasia

- autosomal dominant disorder
- characterized by dilated tortuous BVs w/ thin
walls that bleed readily
- bleeding can occur anywhere: mucous
membranes of nose, tongue, mouth, eyes, GIT
VESSEL DEFECTS

 Amyloid infiltration of blood vessels

- Systemic amyloidosis: associated w/
perivascular deposition of amyloid and
consequent weakening of BV wall
- most commonly observed in plasma cell
dyscrasias
PLATELET DISORDERS

 THROMBOCYTOPENIA
 THROMBOCYTOPATHY
THROMBOCYTOPENIA
 Decreased platelet count
 <150,000/ul
 Spontaneous bleeding not evident until count
falls <20,000/ul
- usually involves small vessels affecting skin,
mucous membrane of GIT and GUT, intracranial
THROMBOCYTOPENIA
 many causes can be classified into 4 major
categories:
1. Decreased platelet production
2. Decreased platelet survival
3. Sequestration
4. Dilutional
 Decreased Platelet Production
 Accompany generalized diseases of BM (aplastic
anemia and leukemias)
 Or result from diseases affecting megakaryocytes
 In Vitamin B12 and folic acid deficiency: (+) poor
development and accelerated destruction of
megakaryocytes (ineffective megakaryopoiesis)
w/in BM due to impaired DNA synthesis
DECREASED PLATELET
PRODUCTION
1. Generalized diseases of BM
- Aplastic anemia
- Marrow infiltration: leukemia, metastatic CA
2. Selective impairment of platelet production
- Drug-induced: alcohol, thiazides
- Infections: measles, HIV
3. Ineffective megakaryopoiesis
- Megaloblastic anemia, myelodysplastic syndrome
Decreased Platelet Survival
 Immunologic or nonimmunologic in etiology
 Immunologic: due to circulating antiplatelet Abs
and immune complexes
- Abs directed against self-Ags on plts (AutoAbs)
- Abs against plt Ags that differ among different
individuals (AlloAbs)
 Nonimmunologic: due to mechanical injury
DECREASED PLATELET
SURVIVAL
 Immunologic destruction  Nonimmunologic
- Autoimmune: ITP, SLE destruction
- Isoimmune: post- - DIC
transfusion, neonatal - Thrombotic
- Drug-associated: thrombocytopenic
quinidine, heparin, sulfa purpura(TTP)
compounds
- Giant hemangiomas
- Microangiopathic
hemolytic anemias
Sequestration
 Spleen: normally sequesters 30-40% of the
body’s platelets w/c remain in equilibrium w/
circulating pool
 Seen in patients w/ marked splenomegaly
(hypersplenism)
 Can be ameliorated by splenectomy when
necessary
 IMMUNE THROMBOCYTOPENIA
(ITP)
 Rare autoimmune disorder
 Syndrome: platelets become coated w/ AutoAbs to
platelet membrane Ags  splenic sequestration and
phagocytosis by mononuclear macrophages
 The resulting shortened lifespan of plts in
circulation + incomplete compensation by ↑plt
production by BM megakaryocytes results in a ↓ #
of circulating plts
ITP
 Primary (Idiopathic) ITP – in the absence of any known risk
factors
- 2 Clinical Subtypes:
1. Acute
2. Chronic
- caused by formation of autoAbs against platelet
membrane glycoproteins IIb-IIIa or Ib-IX
 Secondary ITP – can occur in the setting of a variety of
conditions and exposures (SLE, AIDS, after viral infections,
complication of drug therapy)
Idiopathic Thrombocytopenic
Purpura
Dilutional
 Massive transfusions can produce dilutional
thrombocytopenia
 Blood stored for > 24 hrs contains virtually NO
viable platelets, thus plasma volume and red cell
mass are reconstituted by transfusion but # of
circulating plts is relatively reduced
DEFECTIVE PLATELET
FUNCTION
 Qualitative defects of platelet function can be
congenital or acquired
 Congenital defects of platelet function can be classified
into 3 groups on the basis of specific functional
abnormality:
1. defects of adhesion
2. defects of aggregation
3. disorders of platelet secretion (release reaction)
Defective Adhesion of Platelets
to Subendothelial Matrix
 Bernard-Soulier syndrome
- - Autosomal recessive
- - thrombocytopenia and giant platelets
(macrothrombocytopenia)
- - Inherited deficiency of platelet membrane
glycoprotein complex IB-IX: a receptor for vWF and
essential for normal platelet adhesion to
subendothelial matrix
Defective Platelet Aggregation
 Glanzmann’s thrombasthenia
- autosomal recessive trait
- thrombasthenic platelets fail to aggregate in
response to ADP, collagen, epinephrine, thrombin
due to deficiency or dysfunction of glycoprotein IIb-
IIIa
> a protein complex involved in formation of
“bridges” between platelets by binding fibrinogen
and vWF
Disorders of Platelet Secretion
 Storage pool disorders
 Characterized by normal initial aggregation w/
collagen or ADP, but subsequent responses
(secretion of thromboxanes and release of
granule-bound ADP) are impaired
ACQUIRED DEFECTS OF
PLATELET FUNCTION

 Ingestion of Aspirin and other NSAIDs

 Uremia
ASPIRIN

 a potent, irreversible inhibitor of the enzyme

cyclooxygenase
- required for the synthesis of thromboxane A2
and prostaglandins (play important role in
platelet reaction and subsequent release
reactions)
 The antiplatelet effects of aspirin form the basis
for its use in the prophylaxis of thrombosis
Petechiae
(typical of platelet disorders)

Do not blanch with pressure

(vs angiomas)
Not palpable (vs vasculitis)
Purpura