FOR SECONDARY

IEIVIQSTASIS
 TEST FOR SECONDARY
HEMOSTASIS
i. Clotting Time

2. Prothrombin Time

Activated Partial Thromboplastin Time = f

w

(aPTT)

Stypven Time (Russel Viper Venom Time)

S

Thrombin Time
DW

Reptilase Time ~ .

Duckert’s Test/SM Urea Solubility Test

oN

Substitution Studies
Clotting Time

* Measure’s period
required for free
formation of blood to
clot after it has been
removed from the body
* Used to screen for
problems in the blood
clotting or coagulation
mechanism
 Clotting Time
* METHODS:
1. Capillary blood method
a. Slide Drop Method:
NORMAL VALUE: 2- 4 minutes
b. Capillary method/ Dale and Laidlaw’s Method
-uses Capillary tube (plain)
2. Whole blood (Lee and White Method)
~ NORMAL VALUE; 7-15 minutes
PROCEDURE:
1. 3 test tubes (labeled as 1,2, and 3) in 37 C water bath
2. 1 mL of blood into each of the test tubes
3. Start the timer as soon as blood enters tube 1
4, Gently tilt the tube 3 every 30 seconds ad observe for
clotting
5 After clotting is observed in tube 3, observe tube 2. When
clotting is observed in tube 2 observe tube 1,
 Prothrombin Time (PT)
* Monitors the coagulation factors of the EXTRINSIC and of the
COMMON pathways Catan , throrediaipdentin

* Used to monitor COUMARIN/COUMADIN/WARFARIN therapy siamo

bhp hohe

* NORMAL VALUE: 10-13 seconds

i a n
* Platelet poor plasma (PPP)+Thromboplastin=addition of CaCi2
(begin timing for clot formation)
* INR(International Normalized Ratio) —_> > | >
* Standardized way of reporting PT
* Introduced to minimize the difference in PT results due
to different reagent-instrument combinations
* [SR=International Sensitivity Index
* INR of 2.0-3.0 must be achieved bans cate = iain clat
 * Used to detect deficiencies in fibrinogen, prothrombin,
and factor V and X
Sty pven * Differs from the PT in that deficiencies in factor Vil are

Time(Russel Viper *
acre
Detects coagulation deficiencies in COMMON Pathway

Venom Time) * NORMAL VALUE: 6-10 seconds

* Reagent: East Indian Viper (Viperarusselli)
Platelet Poor Plasma [PPP]
+ Thrombin [Bovine or Human]
Thrombin Time
— Record Clotting Time
Detects fibrinogen deficiency
Sensitive test in detecting Heparin
inhibition
Prolonged in:
* Fibrinogen Deficiency
* Presence of thrombolytic agent
* Presence of FDP and FSP
* Presence of Heparin
NORMAL VALUE: 10-14 seconds
 Detects fibrinogen deficiency

REPTILASE- enzyme found in the venom of the Bothropsatrox snake

Reptilase can convert fibrinogen to fibrin and is unaffected by heparin

Reptilase Time Prolonged in:

+ Fibrinogen deficiency
* Presence of FDP and FSP
* Presence of thrombolytic agent

NORMAL VALUE: 10-15 seconds

Duckert’s Test/5M Urea Solubility
test
* Detects Factor XIll deficiency
* Reagent:
* 5M urea
* 1% monochloroacetic acid
* 2% acetic acid
* Factor XIII deficiency: solubility to 5M urea at room
temperature
 Substitution Tests Using the PT and APTT in the Identification of Single
Substitution Factor Deficiencies

studies Tests
Without
Substitu-
¢ Used to identify factor tions PT APTT
deficiency by mixing
Adsorbed Adsorbed
correction reagents with Plasma Serum Plasma Serum
patient’s plasma and then PT APTT Reagent Reagent Reagent Reagent Factor Deficiency
performing PT and aPTT
N P ¢C NC OVE—
N Pp Cc C XT, X11, HMWK Fletcher*
N P NC C IX
P Pp G NC V,[*
P P NC ¢ X
POP NC NC II, circulating anticoagu
P N NC C Vil

N, mortal time P, prolonged time; C, corrected; NC, not corrected

* Additional testing must be done to differentiate
TEST FOR FIBRINOLYSIS
 TEST FOR FIBRINOLYSIS

Whole blood clot lysis time{WBCLT)

P

Euglobulin clot lysis Time

YP

Protamine Sulfate Test

Ethanol Gelation Test
weP

Latex D-Dimer Test

 Clot should remain intact for approximately
48 hours after 37 C

Whole blood Clot lysis of dissolution prior to 48 hours is

indicative of excessive primary fibrinolysis

clot lysis
time(WBCLT) Clot lysis in48 hours =NORMAL
Clot lysis < 48 hours = INCREASED
FIBRINOLYSIS
 Euglobulin clot lysis Time

* EUGLOBULIN: are protein that participates when plasma is

diluted with water and acidified.
* It consists of plasminogen, plasmin, fibrinogen and
fibrinogen activators

* Plasma + acetic acid = to precipitate euglobulin

* Euglobulin dissolved in buffer + thrombin = TO CLOET
EUGLOBULIN

* No clot lysis > 2hours =NORMAL

* <2 Hours of Lysis: INCREASED FIBRINOLYSIS
Protamine >
Sulfate test
|
—S=E™

* Detects the presence of

FIBRINOGEN MONOMERS 4 77) ee

* PRINCIPLE: Plasma + Protime PROTAMINE

Sulfate = SULFATE
INJECTION, US?
PARACOAGULATION (gel-like
clot)
10 g/m.)

for 1V Use Oanty

* NORMAL:No gel formation
25 mL
* ABNORMAL: Gel formation Seye Toe Ve
© omy
 Ethanol Gelation Test
Less sensitive test but more specific test than protime sulfate
Detects the presence of fibrinogen Monomers
Reagents:
* Plasma
* Ethanol
* NaOH

NORMAL: No gel formation

ABNORMAL: Gel formation
Latex D-Dimer Test

* Used to evaluate specific fragments (D-

dimer)
* To monitor if the fibrinolytic pathway is
functional

* PRINCIPLE: based on Enzyme Immunoassay

* The test uses latex bead coated with
monoclonal antibody specific for D-
dimer but not to fibrinogen and FDP
* The test has no specificity for fibrinogen
and therefore identifies plasmin action
 PRIMARY FIBRINOLYSIS SECONDARY FIBRINOLYSIS

WBCLT <48 HOURS <48 HOURS

EUGLOBULIN CLOT LYSIS TIME <2 HOURS < 2 HOURS

PROTAMINE SULFATE - +

ETHANOL GELATION TEST - +

LATEX D-DIMER TEST = +

SUMMARY OF TESTS FOR FIBRINOLYSIS

 54
Laboratory
Evaluation
of
Fibrinolysis
Robert L. Baglini

jpfERMINATION OF LYSIS TIME Laboratory evaluation of fibrinolysis

has been evolving
Historical Aspects for the past 30 years. It began with
a simple assay, the
Euglobulin Lysis Test whole-blood clot lysis test (WBCLT
), and has progressed
UCTS to assays of fibrin(ogen) degradation
gTERMINATION OF LYSIS PROD D-dimers by latex particle methodolo
products
gy as well
(FDP) and
jstorical Aspects minogen assays using chromogenic substrat as to plas-
ee Sulfate Dilution Test es.
This chapter discusses some assays
Ethanol Gelation Test commonly used for
the evaluation of the fibrinolytic mech
Latex FDP Assay principles, expected
anism. It outlines the
results, and technical difficulties
Latex D-Dimer Assay ciated with their performance. asso-
‘The reader should already
DETERMINATION OF PROTEINS INVOLVED IN LYSIS have a clear understanding of the fibrinol
ytic mechanism
Plasminogen Assay and is referred to Chapter 50 for a detailed
discussion.
Plasminogen Activators
CASE STUDY 54-1
DETERMINATION OF LYSIS TIME
Historical Aspects
The WBCLT is significant from a historical
even though it has been replaced by more perspective,
sophisticated
methods, which will be discussed later in this
chapter. The
WBCLT is based on the fact that whole bloo
d will clot
spon tancously when collected in a glass tube
without anti-
coagulant. This clot should remain intact for appr
oximately
48 hours at 37°C; clot lysis or dissolution prior
to 48 hours is
indicative of excessive systemic fibrinolysis,
This test de-
tects only grossly increased activity.’
Modifications of the WBCLT include the dilut
e whole-
blood clot lysis time, which is said to be a more
sensitive
assay. It is based on the principle that dilution of
blood or
plasma with a buffer will decrease the activity of inhib
itors
to plasmin(ogen),. thus leaving plasminogen activator
or
plasmin free to lyse the fibrin clot.'! Another modification
of the WBCLT is the plasma clot lysis test. In this assay,
blood is collected in an anticoagulant, centr
ifuged, and the

621

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 622 Hemostasis

TS ~
plasma is recalcified. The resultant plasma clot is then incu- DETERMINATION OF LYSIS PRODUC
bated at 37°C and observed for lysis over a 48-hour period. ft n or fibrinogen by plasmin
The WBCLT, as well as its modifications, are empiric The degradation of either er fibri
r eferred to synonymously a,
procedures for the detection of lytic activity in the blood. results in fragments of protein (or split) products (FDP o,
Because of the technical difficulties, as well as the poor either fibrin(ogen) degradation
correlation of the results with clinical conditions, these tests FSP).
are not recommended for use in the modern laboratory.

Historical Aspects hatinat

EUGLOBULIN LYSIS TIME
The euglobulin lysis time avoids the problems of plasmin(ogen) ears, the tanned red-cell hemagglutination inhi.
inhibitors in the assay system. The result is a more rapid and halon text (TRCHN) was the standard against which all
sensitive assay of lytic activity. other quantitative assays of FDP were measured.” Al-
though the TRCHII is still used in research laboratories, its
Principle usefulness in the routine clinical laboratory is limited be-
Euglobulins arc proteins that precipitate when plasma is diluted cause of its complexity. Historically, the reagents used to
with water and acidified. They include plasminogen, plasmin, perform the TRCHII assay needed to be prepared by the
fibrinogen, and plasminogen activators. The inhibitors of fi-
brinolysis (antiplasmin and antiplasminogen activators) remain laboratory; however, today, they are available commer-
in the supernatant fluid and therefore can be removed from the
plasma. oe assay is based on the fact that rabbit-derived antihu-
man fibrinogen antisera, when mixed with the patient’s
Procedure serum, will combine with FDP. After incubation,
The patient’s citrated, platelet-poor plasma is diluted with wa- formalin-fixed, fibrinogen-coated, human group O red
ter, acidified, ‘and refrigerated. The euglobulins will precipitate, cells (tanned RBC) are added to the mixture. These tanned
whereas inhibitors of fibrinolysis remain in the supernate, RBC will then react’with the residual antihuman fibrinogen
which is carefully decanted. The precipitate is then redissolved antisera (antisera that has not combined with FDP). The
and clotted with thrombin. Ifthe plasminogen. the euglobulin mixture is evaluated for inhibition of red cell agglutination,
fraction is converted to plasmin, it will lyse the fibrin clot. The which is inversely proportional to the amount of FDP in the
time needed for complete lysis at 37°C is recorded as the eu- patient serum. Normally, there should be less than 12 wg of
globulin lysis time. fibrinogen-like antigen per milliliter of plasma.
* Quality Control The staphylococcal clumping test’? is of little use today
even though it is a sensitive assay for detecting the presence
Parallel testing of a presumed normal specimen can serve as a
crude quality control measure. Strict adherence to a stan- of split products X and Y (early breakdown products),
dardized procedure is necessary. soluble fibrin monomers, and fibrinogen. To perform the
test, whole blood is collected and allowed to clot, thus
Reference Range removing all fibrinogen. The remaining serum is diluted
The time required for complete clot lysis should be greater than then mixed with a suspension of Staphylococcus aureus strain
2 hours.* Lysis in less chan 2 hours is indicative of increased Newman D2C (coagulase negative) in a microtiter plate,
fibrinolytic activity. High-molecular-weight split products X and Y as well as
monomers will complex with the bacteria, resting mac-
Comments roscopic agglutination, The highest dilution of patient's
The procedure will detect increased fibrinolysis as a result of serum that induces agglutination is determined. The sensi-
surgery, obstetric complications, various medical problems, tivity of the bacterial suspension is quantitated using a fi-
and disseminated intravascular coagulation. This test does not brinogen standard plasma and the final result is reported in
detect fibrin(ogen) split products (X, Y, D, E). fibrinogen equivalents/mL. Normally, there should be 0to
The euglobulin lysis time is performed by many specialized 8 pg of fibrinogen equivalents/mL of serum.
coagulation laboratories but has relatively little application in
the clinical routine laboratory because of the associated techni-
cal problems. For example, the fibrinolytic activity of the eu- PROTAMINE SULFATE DILUTION TEST
globulin fraction is greatly influenced by the precipitation pro- The action of thrombin on fibrinogen results in the formation
cedure, pH, ionic strength, and protein concentration of the test of fibrin monomers that spontaneously polymerize to form a
plasma.* fibrin mesh. When FDP are present in large amounts, they may
If the plasma fibrinogen concentration is less than interfere with this polymerization reaction, thus forming solu-
80 mg/dL, or ifa significant concentration of FDP is present, a ble fibrin monomers. Primary (larger) degradation products xX
poor clot will form, resulting in a false shortening of the lysis and Y may spontaneously polymerize and are also inhibited by
time, hence a false positive. In the case of greatly reduced complexing with fibrinogen, as well as with the secondary
plasminogen concentration, as in disseminated intravascular (smaller) degradation products D and E,'2
coagulation, there is insufficient plasminogen in the euglobulin
fraction for activation and hence a false-negative result.*! Principle
Reference ranges will depend to some extent on the anticoag-
When protamine sulfate is added to plasma, it displaces the
ulant used, as sodium citrate enhances fibrinolysis.
secondary (smaller) degradation products from fibrin mone
Excessive agitation of the tube containing the clot may result
mers and primary (larger) FDP, which will then polymerz
in an erroneous endpoint.
The sample must be tested within 1 hour of collection.
spontaneously. This phenomenon is referred to as “par?”
agulation.’””5

_—
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 Laboratory Evaluation of Fib
rinolysis 623
Pr cedure
v js per formed by adding pati
lent’s 1 Principle
Theassty1 ia dilutions. At the end OF 3a ‘0 protam ;
sulfat
gijutieons are examinined for gel of 30 mi ine Latex particles coated wit
-_ation of fibrin monomegelrs form atioi n, whicIchi
h getes» the h antibody against either
fibrin(ogen)
and early FDP, Indicates: poly. or hu ma n fibrinogen are mixed with patien
mrt difficult to detect, and scrum, Macrosco t
c tread ing of the Tesults erlis p
Tequires some
some~ cates the presence IC agglutination of the latex Particles indi-
experience. of degradation products of either
or fibrin, fibrinogen
Quality Control
cis no adequate quality Specimen Requirem
contro] Procedure for thi ents
ee and negative controls Certain Precautions are req
should b uired
as ¢ run ms met
al ong hod.th
with
Ccause trace amounts of fib during blood collection, First,
rinogen will cause false-pos
atient sample. A positive control may be obtained from ia. reaction
s, it is essential that the itive
tients with known fibrinolytic activity. Strict adherence to pro- with thrombin, Second, sample be thoroughly clo
if heparin is present in tted
cedure is essential. the sample, a
Reference Range
Normally, no gel formation is seen, f
clotting process. Therefore, during the
Comments a trypsin inhibitor should be
to the collection tube to assu added
re the measurement of in
Not in vitro split prod
ucts. Tubes containi vivo and
If fibrinogen . preston high concentrations, an amorphous inhibitor, and venom or ng thrombin, a trypsin
a combination of these com
precipitatereaction.
positive wil develop. This should not be mistaken for a commercially available. ponents are
Because this test is insensiti
ve to fibrinogen and its deg Procedure
tion products, it readily dis rad
tinguishes between primary a-
secondary fibrinolysis. Primar and The patient’s serum in various
y fibrinolysis (degradation of dilutions is mixed with the late
fibrinogen) results in a neg Particles, and the high
est dilution in which macroscopic x
ative Teaction, whereas agglu-
fibrinolysis (degradation of fibr secondary tination is present represents the
in) yields a POSitive reaction: titer of FDP. Depending on
A positive result may be expect which manufacturer’s kit is used
ed in disseminated intrayas- , the sensitivity of the latex
cular coagulation, pulmonary emboli Particles may be to degradation products D and E‘or
sm, and during thrombo- molecular-weight fragments X to high-
lytic therapy, as well as in any and Y.:
other clinical conditions which
lead to the formation of FDP. This rea Quality Control
ction ¢ an be completely
‘inhibited by high levels of plasmin, often
foun d in patients with Positive and negative controls shou
pulmonary embolism. ld
patient specimens. These controls may be run along with the
False-positive reactions may be seen in be supplied by the man-
healthy women im- ufacturer of the kit. If not,
mediately before and during menstruat a positive control can be made
ion and in patients with diluting normal plasma, which conta by
advanced cirrhosis or metastatic can ins: fibrinogen. A normal
cer.!2 serum sample will serve as a negative
Generally speaking, this test is of limite control. The activity of
d value unless fre- the latex particles should be checked
quently performed, because experience in with dilutions of fibrino-
interpretation is of gen or plasma.
prime importance.
*
Comments
Comparisons of these assays have been perf
ormed.®:24.25
ETHANOL GELATION TEST Positive results have been reported in the
nated intravascular coagulation,
following: dissemi-
The ethanol gel test is said to be less sensitive but pulmonary embolism, deep
more specific vein thrombosis, acute myocardial infar
than the protamine sulfate dilution test in detect ction, abruptio placen-
ing soluble tae, preeclampsia, eclampsia, fetal death
fibrin monomers and polymers in plasm in utero, postpartum
a.” hemorrhage, malignant neoplasms, ovari
Principle an tumor, polycystic
disease, renal failure, hydronephrosis, lupus
nephritis, prolifer-
In the presence ofa 50% solution of ethanol, any sol ative glomerulonephritis, renal trans
uble fibrin plantation, cirrhosis, after
monomer complexes present will dissociate, result surgical complications, and during thro
ing in poly- mbolytic therapy, and
menization of the monomers and subsequent gel format in healthy normal women during mens
ion. truation. Fluctuating
his is the same paracoagulation reaction involved levels have been observed during normal
in the pro- pregnancy,
tamine sulfate test. False-positive results have been observed
have not been clotted properly (e.g., hepar in samples that
Procedure in contamination or
dysfibrinogenemia) or from patients with
. circulating rheama-
The test is performed by adding a 50% solution of ethanol to toid factor. Also, if the latex-serum
mixture is not read at the
Platelet-poor plasma and observing for gel formatio prescribed time, a false-positive reaction
n. may result, as pro-
Reference Range longed exposure to air will cause evap
oration and apparent
.. clumping.
There should be no gel formation under normal conditions.
Comments
This test has been used in conjunction with, LATEX D-DIMER ASSAY
or abe A more recent development in the evaluation
the protamine sulfate dilution test, however, it suffers fr of fibrinolytic
Similar interpretation problems. activity is the D-dimer (D-D) assay, which measu
res a specific
fragment arising from the degradation of cross-
linked
fibrin (D-dimer) and not fragments X, Y, D, or E. Becau
se it
measures fibrinolysis and not fibrinogenolysis, the presence
LATEX FDP ASSAY D-dimers is specific evidence of intravascular fibrin formation?
of
€cause of the need for a convenient, rapid .
, and sensitive assay as opposed to primary fibrinolysis (i.e., lysis in the absence of
for FDP, the latex FDP test was developed. fibrin formation).

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 624 Hemostasis

the same pri nciple: an excess of eit) activator Such

Specimen Requirements —— ase is added to a ee sample. The re lat
This assay may be performed on fresh itr? as streptokin
Se inize
vred, } en-streptokinase complex gener ates Plasmin _
EDTA plasma specimens as well as scrum. It 7 atic plasminog ill react with a synthetic (chromogenic) tie
however, that D-dimer levels are somewhat ° er eration tivity that vwenale is a color change
st
proportiona|
than in plasma.”6 This needs to be taken into ¢ oaen level of the plasma. These assays are os to the
when interpreting test results. pias c Mme,
Procedure cially oie ae inherent
to the synthetic substr
There are basically two methods to assay this product: ae iad ier a spectrophotometer OF fluoromete ate toc
dure r is te ie
immunoassay and latex bead. The enzyme immunoa y Pont aca, Dade Protopath); (2) technique
usefull when levels of D-dimer are low and avery sensitive a | : isCrt
(.g21 Du ofspecific reactions as well as the
is required. It does, however, require special technique an oe the on use in which the assays are perfo NAITOW ten,
expensive and time consuming. A busy clinical ser ingatlean rmed must
need for a rapid, simple, yet specific method, For this rea a Pevorstely monitored; and (3) icteric or hemolyzed plasm,
the latex bead method has gained popularity despite the fact tha
it is not as sensitive as the enzyme immunoassay. will interfere with the spectrophotometric Measuremen,
This method utilizes latex beads coated with monoclonal
antibody which is specific for D-dimer but not to ae
degradation products or early degradation products X and Plasminogen Activators
: ys of these proteins hav
The plasma or serum in various dilutions is mixed with e been introduced onl
y recen i
suspension of sensitized latex beads, and the highest dilution
plasma or scrum that causes macroscopic agglutinatio o de clinical Inboratory. A method using 125 | al,
n of these casein has been proposed to be accurate and precis 17
beads is determined. e
Reference Range Chromogenic assays for Tissue Plasminogen Activaty
(t-PA) and Plasminogen Activator (a modificat
Under normal circumstanced, less than
eug
ion of th.
present.
200 ng/mL should be lobulin lysis time) as well as an enzyme immunoasyy
(EIA) procedure for t-PA are now available commercially
Comments The kits, however, are expensive and the Int
erp ation of
Positive results may be seen in pati
ents with DIC, pulmonary the data derived from the assays is difficult to ret
apply to is
and cerebral embolism, phlebitis, thrombosis,
and postopera- relationship to thrombotic risk in the patient,
tively, prethrombotic tisk,
sickle cell disease, and clot
ution secondary to reso-
thrombolytic therapy.* The pres
theumatoid factor, a heterologous antibody, may ence of
Positive reaction.!© cause a false-
There is new evidence to suggest
ness of the D-dimer assay as a screening test for the the useful- CHAPTER SUMMARY
venous thrombosis, diagnosis of
4:22
Fresh citrated, heparinized, The laboratory assays described
or EDTA plasma specimens in this chapter measure some
be used, as well as serum. may components of the fibrinolytic syst
It shou
ld be noted, however, em. They are divided into
D-dimer levels are somewhat that three major categories: measureme
lower in serum than in plasma;? nt of the time for lysis to
this needs to be taken into cons * occur, of the products of lysis,
ideration when interpreting or of specific reactants in the
results. test fibrinolytic system. The
author
tests most commonly employedhas selected for discussion those
tories.
in routine hemostasis labor
DETERMINATION OF PROT The most important requirement for
EINS any of these tests is that
INVOLVED IN LYSIS - they measure i vivo and
not in vitro lysis. To acc
the instruction per taining omplish this,
Plasminogen Assays and assay must be Stri to specimen collection, preparation,
ctly
these assays will aid in the followed. If carefully performed,
Over diagnosis and monitoring of
the years, many methods associated with activatio even's
assay plasminogen, among the have been developed to n of the fibrinolytic
system.
m are the fibrin plate assay?
and the caseinolytic method.!
There exist other methodolo-
gies having wider acccptanc
e and application, Assays of CASE STUDY 54-1
plasminogen by radial immunodif
modification!® are ava
fusion (RID)? and its A35-year-old man was admitted through
ilable commer the emergency depa nt
the patient’s plasma is added and cially. In these tests, with
wouldan beante rior-wal myocardial infarction. It was determined rtme
a favorablle candidat that he
allowed to diffuse from a e for clot
well cut into an agarose mat
rix containing plasminogen therapy (streptokinas ). A coagulation dissolution with thrombolytic
antibody. The interaction of patien profile was performed before
antibody results in an immunopre
t’s plasminogen with the thena'é cardiac
hours cath
aftereterinduction of the therapy. It was later determin
ization laboratory that the clot ed in
cipitation reaction, the was not effectively
diameter of which is measured and lysed. and the patient was Subsequently treated by convention thet-
by control plasma. Although the test
compared to that caused PY using anticoagulants, Coagulation results were as follo al
is simple to perform, ws:
48 hours are required to obtain
results. Also, it measures
only the concentration of the plasmi
nogen molecule immu- PT (9-13 sec) PRETHERAPY _POST-THERAP Y
nologically, not taking into account its functi APTT (24_37 sec)
ities, onal capabil- 12.2 sec 17.0 sec
Fibrinogen (150-409 mg/dL)
300
Plasminogen should be assayed by both imm . FDP (latex) (< 10 pg/ml) g/dl = mg/dl
unologic.
[
and functional assays. The functional assays D-Dimer (< 209 ng/mL) S10 g/mL > 160% "
are all based on Plasminogen (68%~150%) <200 ng/mi 300 ngim
Fog" ca

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 a
52
Routine
—_____ Labo
-a bora
rato
tory
ry
—_____ Eval
Pv aluauatitionon
———__ of vCooaagguuilaattiioon
eo n
Cheryl S. Cook

gENCE RANGES
INTRINSIC AND COMMON bineg centuries, man
1s FOR THE Whol PaTHways has been intrigued by
e Soe coagulation and has the process of
i) and White ation BlTioomed Coagulation Time Precise methods for evalua strived to perfect accurate and
oa necalifi Tine illiam Henson’s ting the complex Process.
dis Since
acti: vated Clo ligated in the Veins of covery in the late 1700s that blood
boplastin+ Tiome me a
-oRThr om
THE EX TRINSIC Vestigators have been
AND COMMON avidly experimentin
PATHWAYS
Measure the blood’s g with ways
i
prothrom bin Time doe ge of coagul
other Tests Orward: when Paul Morawitz, ation took a significant step
published a compre a young German scient
he nsive explanation ist.
Coagulation.23 His obs of the chemistry of
ervations that coagul
occur when plasma was ation did not
placed in containers with
“nonwet-
h as paraffin-lined tubes,
din contact with “wetta but did so
ble” surfaces
led to the
the measurement of coagul development of methods for
ation.?
Among the first researche
rs
Morawitz were Drs. Roger Lee to expand on the theory of
ing the numerous procedure
and Paul White.18 Modify-
s described in the literatur
the early 1900s, these investiga e in
tors developed a procedure
known as the Lee and White
whole blood clotting time
(L-W). Today, more sensitive and
precise procedures have
replaced this test in most institutions
. It Was not until the
mid 1930s that the next breakthrough
in coagulation testing
came about. At this time, Dr. Armand Quick?!
procedure called the prothrombin time (PT) test.described a
With mi-
nor modifications, this test has become a
routine coagula-
tion procedure for evaluating the extrinsic coagulat
ion sys-
tem. In the 1950s, studies of blood from. hemophiliacs,
whose blood yielded normal prothrombin times despite its

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 eee OQGOD

60
and Xs Omm ‘
6 Hemostasis
‘on of 4 fibrin ar atevit
d by activating bre
sti mul
roduction Jude factor the pres Sod
ely char ed surfaces in
Extrinsic System ic sY> reagenr a |
: lcts Or other
Jate
factor, Thy |
thways include all
2
kp supPpo!!on tone |
phosph hway 1s recognized

insithec patintrhwainsiy chavandingcoeay} nt | |

Tissue Factor ro
1c int ene
Vn Catt IT hith c h ep
int ne wit
en

Vil, + Tissue Factor t o s routin

ologie
j n a n 52-
i ‘ij e
as Figur
d
ory metho
Jo domi
gtatus.
s or Jaborat
x, athway
r i ng incinwie an
u o tm
s: thosee. me e tes"* the Browped int ,”
X —— a s
Va r c o r i e h
| rn today i
y
W
discusse T h o s e cmk
ng he
egarsouurnid otf as
s f coagulationays.andThe ba t
Prothrombin eee———-p Thrombin n athwsa
e o cussion
a
saie i
an he resent moen , ee sith a gescncusrasilLondisbelow), n OFing,the
(sce di
test will b m Crerence rangess of CrrOt, and comments a
roccdurc. results, so ur ce So
in
Fibrinogen —$>—=$—_——— Fibr retation 0
f d
. te .
FIG 52-1 is monitored will be presen
> 92-1,
by prothrombin ti trinsic system of coagulation that
Extrinsi
s.
olipid; Cat*, calcium ion
rombin time, PL, phosph
n oes
REFEREN' will be given f
reference ranges roximat e . T
a
7
cha pter, it must be stre sseq
tt di sc us s ed in this affecte
inabili a
1n hemostasis are significantly
led to the development of the
sarah Ee ciate that reference ranges reagent systems
(PTT) test.!7 Although proce” tio ns, methodology, Gon
dural modifications ha fetimesince been-made, this test is still
pop ula
combinations thereof. refetenc,
by pati ent
t sys tem s, and own
one of the bestariilabi instrumen must establish its
tion system. ¢ for evaluating the intrinsic coagula- quently, every laboratory of 20 to 40 deter,
: es. Thi s will inv olv e the performance in the same
Separati proccss into the extrinsic and rang from healthy volunteers
tntrieic i pe na ti on s on pl as ma
in the mi ation of
from observations made samp Jes are to0 bbe tested. Calcul
laboratory.> The extri ved manner as patient’
the addition of a ce mits produces a reference
the mean and 95% confiden uatlied any time changes are
requires
tissue extract to th rinsic system coagulation. Dam-
to induce reeval
aged tissue will sti . Plasma range, which must be instru
extrinsic system by releasing
a
mnade in the met hod (¢.g ., changes in reagents or ils oe
lipoprotein substance ate the referred to as tissue throm- and 62 for further deta
boplastin. This lipo . commonly
in conjunction with factor VII, ments). Refer to Chapters 46
activates Bick > protein,
or X in the common pathway and leads to the establishing reference ranges.

Intrinsic System

HMWK
Kallikrein qc Prexallikrein

HMWK

catt”

| Villa
PL
Catt

X mmm / ’
Xa Va

PL : f
Ca** fo

Prothrombin
ff
FIG. 52-2 .w holeIntrbloinsi
itored by 244. ntris od eae of coagulation that is mon-
Fibrinogegen Plastin time (e; ing time or partial thrombo-
acti
molecular-weight Kininoe nonactivated). HMWK,
—————) Fibrin
high- (either
Ig
Sue en.

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 Routine Laboratory Evaluation of Coagulation 607

THE INTRINSIC AND

look for better methods of monitoring
57). Although by 1964, the PTT test heparin Chere ‘despre
OS ad
Be
sts FOMP
(0 f whiteATHW
WholAYS
e Blood Coagulation Sensitive to heparin, the test had not
was recogn
Time acceptance as the heparin-monitoring yet gainc a ee a
yo procedure.
velopment in the carly 1900s of safe
ith iheie plood by venipuncture, : and new procedure based loosely on the L-W test, Dr, - I Hat-
many y casy tersley developed the activated Pau 1&7
renipl A aria tions on clotting time (ACT) test.
Ay8 toleobta ood coagulation time were tried, [y 1913, The ACT uses diatoraccous earth f
' Lee the contact factors and requires the (diatomi te)
blood to be askept
an enter fe
Wi
iy oO
whit N F sescribed
that a procedu re for the whole blood warmed to
wh te de was a modification Coag-
of the numerous
4 Constant 37°C by taking a special incubator to the patient's
of! on rime bed in the bedside.
vod dese the fact literatu re at that time,
that when venous blood 18 The L-w Principle
me ig base (foreign surface), is putintoa
oe ube ed for this responsit will form a solid clot. The Whole blood contains all the components neces
sary to produce
e isa measure of the Overall a clot when removed from the veins and put
er seguir’ common pathwa into a glass tube.
ys of coagulation, By adding an activator and keeping the blood
ons 7 7 ig insensitive at a constant
to factor deficiencies, 37°C, a more reliable and rapid screen of the intrinsic and
the £ ay be present in the and moder-
face common pathways is achieved.
se defects | 2% of the of normal results,4
normal level of factor Reagents
‘aly! . a] L-W clotting VI will result
time when mixtures Two evacuated tubes containing 12 mg of diatomite are needed.
in a 10 ophiliac plasmas of normal
are tested. Technical
and hemo agitation errors, such Equipment
cess" of the tube when checking for
sed a clot
=iex shortene times. Moreover, there is no suitable qual- A portable heat block, thermometer, and two stopwatches are
measure for this ure.'§ For
proced used.
these
Wis n0 longer-considered an adequate test. reasons, Procedure
je L- - The two tubes containing diatomite are brought
to 37°C in a
heat block at the patient’s bedside. Using good venipu
jasma Recalcification Time technique, at least 2 mL of blood is drawn into ncture
a tube and
~P modification of the L-W discarded. The tourniquet is removed,
pode g citrated plasmais the recalcification time of
and the first tube
with
diatomite is attached to the needle. When blood
(instead af whole blood), starts to How
oa "glass or siliconized tubes, into the tube, the first stopwatch is started.
The tube is filled,
Cal a (PRP), platelet-poor pla and either platelet-rich mixed, and placed in the heat block. The proced
ure is repeated
sma (PPP), or both.!4 This with the second tube, and the second stopwatch
Con ase don the fact that except 60 seconds, the first tube is observed by tilting
is started. After
for calcium, normal PRP it ac 5-second
leant all the components of intervals until a clot is formed, at which time
the coagulation mechanism the second tube is
ann for generating a fibrin clot. Rem observed using the same procedure. The appropriate stopw
an the clot easier to see. The oval of red cells 4s stopped at the first appearance of a clot atch
time required for blood to in each tube. The
pm after Cat? is added is a genera -duplicates should agree within 10 seconds.
l measure of the intrinsic The average time is
ind common pathways. By usi reported.®
ng a parallel test on PPP,
screening for a platelet function defect may Reference Range
also be accom-
plished. Reference ranges for PRP and PPP are 100 to 150 The referenee range is 75 to 120 seconds.
The target
range
seconds and 130 to 240 seconds, respectively. Platelet-ric during heparin therapy is 140 to 185 seconds.
h
plasma should clot at least 20 seconds faster than platelet- Interpretation
poor plasma.? a Prolongation of the ACT is indicative of one or more
factor
The principal disadvantages of the plasma recalcification defects in the intrinsic or common pathways or the presen
ce ofa
time are the difficulty in standardizing the number of plate- circulating anticoagulant such as heparin.
lets in the PRP and the length of time necessary to perform Comments
the test, which moreover is insensitive to moderate factor
Temperature control is critical: all testing must be maintained at
deficiencies. In addition, errors in collection technique can 37°C. The primary use of the ACT is during extracorporeal
significantly affect the results, as will the amount of glass circulation, where frequent testing and rapid turnaround time is
contact. It is important that the same size tube always be required.® Other than the duplicate testing, there is no suitable
used for testing and that the specimens be tilted uniformly. method of quality control for this procedure.
Because the procedure cannot be standardized, it is impor-
tant to keep as many variables constant as possible. The PARTIAL THROMBOPLASTIN TIME
sensitivity of this test is somewhat improved by diluting the Several researchers observed that’ blood samples from classic
phsma, This accomplishes three things: (1) it adjusts the hemophiliacs have a prolonged clotting time, but that when
closer to the actual in vivo platelet count; (2) it increases tissue thromboplastin is added, as in the PT, the plasma clots
re sensitivity of the test system to factor deficiencies; and just as normal plasma does.'? Expanded studies showed that
it dilutes the natural inhibitors to coagulation that are thromboplastins, which are lipoproteins (protein + phospho-
lipid), may be classified as’complete or partial (only phospho-
sent. A normal control should be run with each test. lipid): From these conclusions, investigators developed the
nonactivated PTT. This test is more sensitive to abnormalities :
VATED CLOTTING TIME 7 in the early stages than were previous tests of the intrinsic
Y the miq 1960s, the lack of sensitivity and puctision of the system. An important refinement of the PTT was the Fee
of negatively charged activators to the system, resulting ir,
|W and the plasma recalcification test had led resear

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 d by the APTT is less th
60
8 Hemostasis s¢any fact? x meastire "40, ty
‘on is the
prt itary
ft of norma ; al
ee ; ification
| Significantly shorter clotting times.” This moilifict method is APTT may be grouped int
activated
ated partiali thromboplastinin time
time (APTT),
( ‘reen"9°for facto r Commer
0 ri n the-tion and prep aration Dy TeaBent °p th ee
how used exclusively, It is the test of choice to ane and also for Sources o ample collec Improper collection and To “Pata,
deficiencies of the intrinsic and common a assays categories 5 umentation: tly affect the results. The : eesing
Monitoring heparin therapy. It also is the basis 10" ° tion, and si nificant ed for individuals with hes

sit ep eps em am Pe ran 220g

Per a(S
a ne 7 APTT
the intrinsic
reagent system
consists (sec
of Fig.
two TEcomp ait ‘ a platelet
nhios- of spe?
t volu{ a should55 an otI Iess thanfin0.20 L/L toavoign
» ety,

Pholipids, eit and an activator. . Kaolin, ’ Celite, micron?

; fac-
nanufac vt by an cau secause
a falsely 5: - Plata) t
ellagic acid are used as the activator, depending on the! . sedHemolysis+. ™ may Y may erratic resultsIts or falsely sho,let 5
furer, Bt) plasma - d heparin contamination (e.g,, from he
jn the
the APT:
pias Unexp ectjously lengthen the APTT. iPs
Principle : t s pur by improper storage, Water j on
. aximum «. focks) can cd
a
activati ee » fe ary aishediby
by wiition
4 of ain oes. ma nee
1
tor
Reagent
iy 2 systems should
Hl be test, le d
deficiencies by performing the A
ritics, OF incor
the activator. Phospholipid en is supplied to subs tute for c platelet ;
factor 3 (PF3). From aie this point, ef the APTT is essentially the for sensitivityrons 0 f plasma. . :
‘ nh u
same . as a recalcification
ification titime of f plaplasma. on Aserial
failingdul
lightht sot
source, f uctuations
ee pitarad ; ; |o..
temperature,
. in ill
4 nation Will Cause Instrum,
Specimen Requirements “to calibration © 4 quality control program should reveal any j
Citrated platelet-poor plasma should be collected according error. A g0° : ent-related error. Refer to Chapter 62 for,
the guidelines in Chapter 51. strumental Of FEAT and quality control.
R eagents
discussion of instrume
Phospholipid with activators, (APTT reagent) and 0.025 or
CaCl; (or as recommended by reagent manufacturer) are used. HE EXTRINSIC

Controls TESTS FOR ON PATHWAYS

Commercial lyophilized controls are available ae normal, AND C
midrange, and extended ranges. It is recommended that a nor-
mal control and at least one abnormal control be used.”4 In- PROTHROMBIN TIME h physiologist de Bai
h Ouse preparations
i of pooled/frozen plasma may be used as Early exper iments by the French phy:
ainville! gi
controls. Each laboratory must specify when controls areto be showed that blood could be i de € to to c clot. pequickly ly b byadding;
tested, what the satisfactory control limits are, and, if duplicates tissue factors., The discovery that oo and Citrate inhibit
are run, how closely the values should agree. coagulation” led to the assumption that clotting was dependey
calcium. . .
Equipment
or The one-stage PT, as described by Dr. Armand Quickin
For the manual method, 12 x 75-mm glass tubes, a heat block, 1935, was believed to be an indirect measure of prothrombinin
and pipets are needed. For the fibrin strand method, an instru- lasma, dependent on the presence of fibrinogen. Subsequent
ment for electromechanical fibrin strand detection and appro-
viscoveries of factors V, VII, and X showed that the PT is
penis cope 7nd Piper neceamerced eh the apna ae reflection of the activities of several factors.*° Thus, the
instrument ‘the phoro-optical
ere zeqaizedand Forappropriate by spout
as listed Che
accessoriesmeshosl, the manu- PT used in today’s laboratory screens for deficiencies of factors
~. I, Il, V, VI, and X.
facturer, are needed. OI Although the PT is no longer considered a measure of pro-
Procedure ~~ thrombin itself, it is still the test of choice for monitoring
Platelet-poor plasma (0.1 mL) is added to 0.1 mL of APTT anticoagulant therapy by vitamin K antagonists (Chap. 57),
reagent and incubated at 37°C for the period oftime specified by Three of the five factors measured by the PT (II, VII, X) are
the reagent manufacturer (approximately 3 to 5 minutes). After sensitive to and depressed by these anticoagulants.” Only fac-
incubation, 0.1 mL of warmed CaCl is added, and the time for tor IX, the other factor de pressed by vitamin K antagonists, is
clotting to occur is recorded. not detected by the PT.

Reporting Principle
The APTT is reported in seconds, to the nearest tenth, along When tissue extract or thromboplastin is added to PPP along
with the reference range. yeueiium, it reacts with factor VII, to convert factor X to
Reference Range a Factorprotbromiie
converts X,, along witl factor Vay Phospholipid,a: and Ca #2
The ranges differ according to the reagent, method, and instru- converts fibrinogen to fibrin (see Fig 52 im Subsequently
ment used. The reference ranges may extend from a lower limit ddit; . 92-1). The time fr
of 20 seconds to an upper limit of 45 seconds. pO ete romboplastin/CaCl,
reported as the PT.9 to the formation of a clot i
Interpretation Specimen Requirem
ents
When properly performed, the APTT is an excellent screenin Citrated platelet. :
est for the intrinsic and common coagulation pathways, A Buidelines in a Is collected according to the
rolonged APTT in the absence of heparin use indicates a factor Rea gents and Equi
eficiency, an acquired ee anticoagulant such as the Th ‘pment
tpus inhibitor, or an antibo yto sata factor such as factor Sige mboplasti /CaCl, (PT rea oe
III. Most commercial reagents will demonstrate 3 prolonged ‘a in pers APTT Controls) ar Bent) yep (see ant
© as that used fo ¢ needed. The equipment 1s the
tthe APTT.

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 Routine aboratory Evaluation of Coagulati
on 609
pr ced i
ntrol and d p patient plasma a Te warm
ique of being used, The i thr Omboplasti ed accorg; time was abando
n tors Vy]25 ned as unr
cliable. The discovery of
Al metho incubating it at 37°C for 3 ¢ 05 and X™* fac-
memes
war t)
Minute
DY 5s added to 0.1 mL of pla Sma (patie Stypven ¢i me and explained the dis crepancy between the
. nt the PT,2! The Stypven time
of i.ing (1 me is recorded, > and until factor was rein-
c Jotting Sed ty help assa ys became more routine,
ghe nce Range di ifferenti ate betw it was
ere iffer een factor VII and factor Xx
with , Today
ref! al values diffe 10 to the
12 secreapen
BY t r sionally, » the Stypven time is ordered
of ay range from and Method, , se
The occa-
; values m
to 12 to 14 seconds wi : ones
th manual inMeso me ph Oto-opej In 1951, Drs, Owren and Aas?’
sys tems
thods, Ptical Mand proc
on vertin (P
developed the prothrom-
7 oe tions that & P) test based on earlier
times may be ming r defici observa-
prothrom reported in Se test plasma encies can be more Pronou
© ids) with the veral Ways: is di luted. Plasma nced when
ime (2 control refere (1) is tested at 1:10 dilution,
time (in second nce range; (2y PaticntPatitient
a Teagent
9 and
OVine brain, taining dilute thromboplastin extract
con
wi s) ; (3) Prothrombi me from
prthot <e atient divided by th
by 100—rare e Mean of the refe
n ratio (the CaCh, a nd
an excess of bovine fac
ly used in rence range and rino BEN)
is used." The tor s V and I
multip et activi the Unit addiri
r Se enetouil ty (outdated and not reco ed States). and common Pat
sensitive to those factor
s in the extrinsi
normalized rati mm
o (INR) has enbeded), The use hways th at are affected by vitamin K ant
of sts andard method of reportin 8. It is popular enj fy oPosed as
the
nists (i.e., fact
freeze-dried, co
ors Il, Vil,
an d AK). O wren later dev
ago-
i ; Spular in Euro pe but
not W:videly used in the United States (see Chap, 62). mm er cially availab le eloped a
test) based on reagent (Thrombo-
the ori ginal P @& p Pp rocedu
Int erpgat
prolon retion
ationof th WwW aS j adjust
ed 3f°
¢ re. Its sensitivity
ind icates an
e PT ind peutic tange.°? Bive reliable dete Tminations in the the
abnormalit:y of ra-
mon or extrinsic co
ag ul at io
one or more N etherlands andThrombotest is most Commonly used in
con be hereditary or Sca f; OT monitoring vitami the
n fa ctors. This abnorm ndi nav ia
an factor inhibitor acquired. Prolongation ma ality antagonist th
s. Using most co y also occur erap Y- In this cou n K
PT is sensitive
¥
gm mm rrc ial rea ts, the Particularly fa ntry, the P
of Jegg than 40% gen
to factor deficiencies v Ored at the time
when sodium oxalat
to 50% of used as an anti e was
normal. c Oagulant, as thi
factor V stable s Material failed
Comments . Today, the test to hold
Screen for thee has little value as an
Reporting of percent act Xtri nsic system bec overall
ivity is no Jon Ser factor V and fi ause of its insensitivit
recommended b y to
cause the dilution curve used to det be- sodium citrate » with the current
dilutes all factors, not just those e rm un e , percen as the anti
coagulant of cho use of
affecte dby antico t activity of factor V is no lon
ger a great problem ice, the instability
apy, and is therefore an ina
ccurate re Presenta
agulant ther:
The preferred method tion of therapy.
of Teporting in the
along with the refere
nce range. The Britis
US is patient PT
established standardized h and Dutch have CHAPTER SUMM
national systems for ARY
standardized reference PT. They use a
reagent (e.g., the British Comparat In this chapter, the de
Thromboplastin) and ive ve
that have proved to be
a national system of
reporting PT results routinely in Coagulat lopment of some of the procedures us
highly successfiy],26,28. ion testing was dis ed
dures were described cus
’ error in this test are
similar to those dis
37 The sources of , along with refere s ed. General Proce-
cance of abnormal nc
cussed for the APTT.
resu] ts, and potential € ranges, the signifi-
lizing these procedur so
es, general Screening urces of error, Une
abnormalities may
Other Tests be performed. In add for basic coagulation
ition, Many of the
Oring anticoagulant se
Two other laboratory therapy.
determinati ons related to
sic and common path the
wa ys are the Stypven time extrin- controlled, they can ct test results. [f
thrombin—proconvertin
time. Neither test
and pro- have adverse conseque nor
tients being monitore nces for those pa-
widespread Popularity, has enjoyed d on anticoagulant
therapy. Chapter 62
discusses in detail qua
day. Therefore, they willandbe they are rarely performed to- lity control for the
Coagulation labo_
only, discussed in a historical sens ratory.
e Two other procedures that
The Stypven have become r outine
tests in recent years were
Properties of Ru time u tilizes the powerful coagulant not discussed in t his chap coagulation
gen measurement and qua ter: fibrino-
ss
nake Viperg russ ell’s y iper venom, obtained from the dation products. These two
ntitation of fib rinogen/
fibrin degra-
elli. This venom is capa procedures a Te covered
yon of factor ble of bypa 53 and 54, Tespectively. in Chapters
VII and directly activating factor ssing the
X to X,.
“ It is combined with dilute thromboplast
in, a fibrin
‘lot wil form through the reaction of factors
X, and V,, REFERENCES
Mospholipid, factor II, and fibrinogen. In one of many
been ations of Quick’s original PT, Wites and Babee 1. de Bainville HMD: Injection
de matiare cérébrale dans
and the venom to be a convenient substitute for the veines. Gaz Med Paris 2:524, 1834 les
ta wtbop astins used in the PT system. This substitution 2. Brandt J: Principles of Coagulation
1982. Symposium spon-
sored by General Diagnostics Corporation,
nee Problems in managing patients on antioea ast 3. Davie EW, Ratnoff OD: Waterfall
Lexington, KY,1982
tines Ys as the Stypven time produced shorter ¢ orth ng sequence for intrinsic blood
clotti
ng. Science 145:1310, 1964
thanbleedj
"esy) tant di the PT and led to serious overdosi g, 4. Giddings JC: Hereditary coagulation
ng.. Because of the discrepancy, the Stypven disorders: Lab Oratory
techniques. In Thomson
JM (ed): Blood Coagulation a nd Hae-

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 Qualitative Disorders of Platelets 41
and Vasculature
Larry D. Brace

OUTLINE OBJECTIVES
Qualitative Platelet After completion of this chapter, the reader will be able to:
Disorders
Disorders of Platelet 1. Describe the effect of aspirin on the cyclooxygenase 7. Distinguish among the following types of inherited
Aggregation Affecting pathway. platelet disorders: membrane receptor abnormality,
GP IIb/IIIa Function 2. Describe the defect in each of the following heredi- secretion disorder, and storage pool deficiency.
Disorders of Platelet tary disorders: storage pool disease, gray platelet 8. For each of the inherited platelet disorders listed,
Adhesion Affecting syndrome, Glanzmann thrombasthenia, and Bernard- name useful laboratory tests and recognize diagnos-
GP Ib/IX/V Complexes Soulier syndrome (BSS). tic results.
Inherited Giant Platelet 3. Discuss the mechanisms of action of antiplatelet 9. Identify the most common type of hereditary platelet
Syndromes drugs. defect.
Disorders of Platelet Secretion 4. Explain the effects of paraproteins on platelet function. 10. Discuss the mechanism of the platelet defects
(Release Reactions)
5. Compare and contrast Glanzmann thrombasthenia associated with myeloproliferative diseases, uremia,
Inherited Disorders of Other
and BSS. and liver disease.
Receptors and Signaling
Pathways 6. Recognize the clinical presentation of patients with 11. Recognize conditions associated with acquired vas-
Acquired Defects of Platelet dysfunctional platelets. cular disorders.
Function
Vascular Disorders
Hereditary Vascular Disorders
Acquired Vascular Disorders

CASE STUDY
After studying the material in this chapter, the reader should be able to respond to the following case study:

A 19-year-old woman with a chief complaint of easy and collagen-induced aggregation was decreased, although
bruising, occasional mild nosebleeds, and heavy men- a near-normal aggregation response could be obtained
strual periods was examined by her physician. At the time with a high collagen concentration. Spontaneous aggrega-
of her examination, she had a few small bruises on her tion was not observed.
arms and legs, but no other problems. Initial laboratory 1. What are three possible explanations for the test results
data revealed a normal prothrombin time, and a normal so far?
partial thromboplastin time. A CBC, including platelet 2. Given the bleeding history in her family, which of the
count and morphology, yielded normal results. three explanations seems most likely?
A detailed history revealed that the patient’s bleeding A quantitative test for adenosine triphosphate (ATP)
problems occurred most frequently after aspirin ingestion. release was performed using the firefly luciferin-luciferase
Her mother and one of her brothers also had some of the bioluminescence assay. The result of this test showed a
same symptoms. Blood was drawn for platelet function marked decrease in the amount of ATP released when
studies, and platelet aggregation tests were performed. Ag- platelets were stimulated with thrombin.
gregation induced by ristocetin and adenosine diphosphate 3. Based on the ATP test results, what is the likely cause of
was near normal, arachidonic acid–induced aggregation the patient’s bleeding symptoms?
was absent, epinephrine induced only primary aggregation,

739
740 PART VI  Hemostasis and Thrombosis

C
linical manifestations of bleeding disorders can be
divided into two broad, rather poorly defined groups: BOX 41-1  ​Qualitative Abnormalities: Changes
(1) superficial bleeding (e.g., petechiae, epistaxis, or in Platelet Function (Thrombocytopathy)3,4
gingival bleeding), usually associated with a platelet defect or
vascular disorder; and (2) deep tissue bleeding (e.g., hemato- Disorders of Platelet Aggregation
mas or hemarthrosis), usually associated with plasma clotting Glanzmann thrombasthenia
factor deficiencies.1 This chapter describes platelet and vascular Hereditary afibrinogenemia
disorders. Bleeding problems resulting from defects in the co- Acquired defects of platelet aggregation:
agulation mechanism are described in Chapter 38. Acquired von Willebrand disease
Acquired uremia

QUALITATIVE PLATELET DISORDERS Disorders of Platelet Adhesion

Excessive bruising, superficial (mucocutaneous) bleeding, and Bernard-Soulier syndrome
a prolonged bleeding time in a patient whose platelet count is Von Willebrand disease
normal suggest an acquired or a congenital disorder of platelet Acquired defects of platelet adhesion:
function. Congenital disorders have been described that result Myeloproliferative, lymphoproliferative disorders, dysproteinemias
from abnormalities of each of the major phases of platelet Antiplatelet antibodies
function: adhesion, aggregation, and secretion. Rapid progress Cardiopulmonary bypass surgery
in this field began in the 1960s, mostly as a result of the Chronic liver disease
development of instruments and test methods for measuring Drug-induced membrane modification
platelet function.2
Qualitative platelet disorders are summarized in Box 41-1.3,4 Disorders of Platelet Secretion
This chapter discusses the individual qualitative platelet disor- (release reactions)
ders grouped by the mechanism that causes the defect. A sum- Storage pool diseases
mary of the primary disorders is illustrated in Figure 41-1 for Thromboxane pathway disorders
defects associated with surface components and Figure 41-2 for Hereditary aspirin-like defects:
defects associated with intracellular components.5 Cyclooxygenase or thromboxane synthetase deficiency
Drug inhibition of the prostaglandin pathways
Disorders of Platelet Aggregation Affecting Drug inhibition of platelet phosphodiesterase activity
GP IIb/IIIa Function
Glanzmann Thrombasthenia Changes in Membrane Phospholipid
Glanzmann thrombasthenia originally was described as a bleed- Distribution
ing disorder associated with abnormal in vitro clot retraction Scott syndrome
and a normal platelet count. Clot retraction is the process of Stormorken syndrome
the compaction of a formed clot (reducing its volume), medi-
ated by contraction of the intracellular actin-myosin cytoskel- Hyperactive Prothrombotic Platelets
eton of the activated platelets incorporated in the clot. It is in-
herited as an autosomal recessive disorder and is seen most
frequently in populations with a high degree of consanguinity. heterozygotes has been found to be 50% to 60% of normal.8,9
Heterozygotes are clinically normal, whereas homozygotes Binding of fibrinogen to the GP IIb/IIIa complex mediates
have serious bleeding problems. This rare disorder manifests normal platelet aggregation responses. Failure of such binding
itself clinically in the neonatal period or infancy, occasionally results in a profound defect in hemostatic plug formation and
with bleeding after circumcision and frequently with epistaxis the serious bleeding characteristic of thrombasthenia.2,8,10-13
and gingival bleeding. Hemorrhagic manifestations include More than 70 mutations are known to give rise to Glanzmann
petechiae, purpura (Figure 40-1), menorrhagia, gastrointesti- thrombasthenia.14-16 GP IIb/IIIa is coded by the ITGA2B and
nal bleeding, and hematuria. There are wide variations in the ITGB3 genes present on chromosome 17, and genetic defects
clinical symptoms. Some patients may have minimal symp- are distributed widely over the two genes.17 Rarely, thrombocy-
toms, whereas others may have frequent and serious hemor- topenia and large platelets may be seen with some mutations
rhagic complications. The severity of the bleeding episodes in these genes (Table 40-1). aIIb is synthesized in megakaryo-
seems to decrease with age.6,7 cytes as pro-aIIb, which complexes b3 in the endoplasmic re-
The biochemical lesion responsible for the disorder is a ticulum. The complex is transported to the Golgi body, where
deficiency or abnormality of the platelet membrane glycopro- aIIb is cleaved to heavy and light chains to form the complete
tein (GP) IIb/IIIa (integrin aIIb/b3) complex, a membrane re- complex. Uncomplexed aIIb and b3 are not processed in the Golgi
ceptor capable of binding fibrinogen, von Willebrand factor body. As with the GP Ib/IX/V complex, it is necessary for both
(VWF), fibronectin, and other adhesive ligands. Typically, the proteins of the GP IIb/IIIa complex to be produced and
platelets of homozygous individuals lack surface-expressed GP assembled into a complex for the complex to be expressed on
IIb/IIIa, whereas the GP IIb/IIIa content of the platelets from the platelet surface. Gene defects that lead to the absence of
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 741

Reduced response to collagen

Glanzmann thrombasthenia:
no aggregation with IV VI CD9 Scott syndrome:
natural agonists
decreased thrombin
generation

llb3
61

51
Reduced response
to collagen

21
IX Bernard-Soulier
syndrome:
V lb lack of adhesion to
Altered response to: VWF and abnormal
ADP, TXA2, adrenaline lb response to thrombin
Platelet-type VWD:
spontaneous binding
of VWF to GP lb
Figure 41-1  ​An illustration of the primary disorders associated with defects of the surface components of platelets.  (From Nurden P and Nurder AT.
Congenital disorders associated with platelet dysfunction. Thromb Haemost 2008;99:253-263.)

-Granules:
Gray platelet syndrome
Quebec platelet disorder
-Dense granule-storage pool deficiency

IV VI CD9

llb3
61

Signaling pathways:
G-proteins 51
Phospholipases
Enzymes:
Cyclooxygenase
TXA2 synthetase
21 Lipoxygenase
IX

V lb
Dense granules:
Hermansky-Pudlak syndrome lb
Chediak-Higashi syndrome Cytoskeleton:
Dense granule-storage pool deficiency Giant platelet syndromes
Wiskott-Aldrich syndrome
Figure 41-2  ​An illustration of the primary disorders associated with defects of the intracellular components of platelets. (From Nurden P and Nurder AT.
Congenital disorders associated with platelet dysfunction. Thromb Haemost 2008;99:253-263.)

production of either protein lead to absence of the complex on complex (e.g., fibrinogen binding or signal transduction) are
the platelet surface. Defects that interfere with or prevent com- abnormal, however. Bleeding in these patients ranges from
plex formation or affect complex stability have the same effect. mild to severe.
Numerous variants of Glanzmann thrombasthenia have One component of the aIIb/b3 integrin, b3, is a compo-
been described in which the GP IIb/IIIa complex is qualita- nent of the vitronectin receptor, aV/b3, found on endothelial
tively abnormal. aIIb and b3 are produced, form a complex, cells, osteoclasts, fibroblasts, monocytes, and activated B
and are processed normally. One or more functions of the lymphocytes, where it acts as a receptor for a variety of
742 PART VI  Hemostasis and Thrombosis

adhesive protein ligands. Patients who have b3 gene defects platelets may interfere with the normal transfused platelets,
that result in the absence of aIIb/b3 integrin also lack the and it may be necessary to infuse more donor platelets
vitronectin receptor. These patients do not seem to have a than expected to control bleeding. As in Bernard-Soulier
more severe form of Glanzmann thrombasthenia.18,19 The syndrome or any situation in which repeated transfusions
vitronectin receptor is thought to play a role in vasculariza- are required, patients with Glanzmann thrombasthenia may
tion, but so far, no evidence for abnormal blood vessel devel- become alloimmunized. Strategies to reduce alloimmuni­
opment has been documented in individuals lacking the zation include use of single-donor platelet apheresis prod-
vitronectin receptor. It also is unclear whether platelet vitro- ucts, HLA-matched donor platelets, or ABO-matched donor
nectin receptors play any significant role in platelet functional platelets.27
processes.14 A variety of treatments have been used successfully to
Rarely, a thrombasthenia-like state can be acquired. Such control or prevent bleeding, alone or in combination
conditions include development of autoantibodies against GP with platelet transfusion. To a large extent, the site of hem-
IIb/IIIa, multiple myeloma in which the paraprotein is directed orrhage determines the therapeutic approach used. Hor-
against GP IIIa, and afibrinogenemia. A thrombasthenia-like monal therapy (norethindrone acetate) has been used to
state also can be induced in individuals with otherwise normal control menorrhagia. If the patient is treated with oral
platelet function by the therapeutic antiplatelet drugs ticlopi- contraceptives, excessive bleeding should be reduced. Men-
dine and clopidogrel.7,12 orrhagia at the onset of menses is uniformly severe and
can be life-threatening, which has led some to suggest that
Laboratory Features.  The typical laboratory features birth control pills be started before menarche. Also, antifi-
of Glanzmann thrombasthenia are a normal platelet count, brinolytic therapy (aminocaproic acid or tranexamic acid)
normal platelet morphology, and a lack of platelet aggrega- can be used to control gingival hemorrhage or excessive
tion in response to all platelet activating agents (including bleeding after tooth extraction.26 Recombinant factor VIIa
ADP, collagen, thrombin, and epinephrine).2,8,10,12 If the has proved useful to treat severe bleeding in patients with
stimulating agent is strong enough (e.g., thrombin), the isoantibodies to aIIb/b3 and in patients undergoing invasive
platelets undergo the release/secretion reaction, even in the procedures.28 Recombinant factor VIIa (rVIIa, Novo­Seven,
absence of aggregation. Ristocetin-induced binding of VWF Novo Nordisk Inc, Princeton, NJ) is thought to enhance
to platelets and the resulting platelet agglutination are nor- thrombus formation at the site of a lesion by stimulating
mal. The results of the complete blood count (CBC) are usu- tissue factor-independent thrombus generation and fibrin
ally normal, unless there is another underlying disorder or formation.29
the patient has had a recent hemorrhagic episode. Tests for
platelet procoagulant activity, previously called the platelet Disorders of Platelet Adhesion Affecting
factor 3 test, usually show diminished activity.2,9,20 There GP Ib/IX/V Complexes
seem to be several reasons for this. When normal platelets Bernard-Soulier (Giant Platelet) Syndrome
are activated, procoagulant microvesicles are shed from the Bernard-Soulier syndrome (BSS) is a rare disorder of platelet ad-
platelet surface, and coagulation factors assemble on the hesion that is usually manifested in infancy or childhood with
microvesicle surfaces during activation of the coagulation hemorrhage characteristic of defective platelet function: ecchy-
cascade. In Glanzmann thrombasthenia, markedly fewer mi- moses, epistaxis, and gingival bleeding. Hemarthroses and ex-
crovesicles are produced. Second, prothrombin binds directly panding hematomas are only rarely seen. BSS is inherited as an
to GP IIb/IIIa. Because this complex is missing in Glanzmann autosomal recessive disorder in which the GP Ib/IX/V complex
thrombasthenia, significantly less thrombin is generated in re- is missing from the platelet surface or exhibits abnormal func-
sponse to tissue factor. Finally, Glanzmann thrombasthenia plate- tion. Heterozygotes who have about 50% of normal levels of
lets are not as activated by thrombin as are normal platelets.21-24 GP Ib, GP V, and GP IX have normal or near-normal platelet
A subdivision of Glanzmann thrombasthenia into type 1 function. Homozygotes have a moderate to severe bleeding
and type 2 has been proposed. In general, individuals with disorder characterized by enlarged platelets, thrombocytope-
type 2 disease have more residual GP IIb/IIIa complexes (10% nia, and usually decreased platelet survival. Platelet counts
to 20% of normal) than those with type 1 disease (0% to 5% generally range from 40,000/mL to near normal.30 On periph-
of normal), although there is considerable variability within eral blood films, platelets typically are 5 to 8 mm in diameter,
each subdivision.25,26 but they can be 20 mm (Figure 41-3). Viewed by electron mi-
croscopy, BSS platelets contain a larger number of cytoplasmic
Treatment.  Thrombasthenia is one of the few forms of vacuoles and membrane complexes, and these observations ex-
platelet dysfunction in which hemorrhage is severe and dis- tend to megakaryocytes, in which the appearance of the demar-
abling. Bleeding of all types, including epistaxis, ecchymosis, cation membrane system is irregular.2,3,8,10,31
hemarthrosis, subcutaneous hematoma, menorrhagia, and Four glycoproteins are required to form the GP Ib/IX/V
gastrointestinal and urinary tract hemorrhage, has been re- complex: GP Iba, GP Ibb, GP IX, and GP V. In the complex,
ported. Treatment of bleeding episodes in patients with these proteins are present in the ratio of 2:2:2:1. The gene for
Glanzmann thrombasthenia requires the transfusion of nor- GP Iba is located on chromosome 17, the gene for GP Ibb is
mal platelets. In Glanzmann thrombasthenia, the defective located on chromosome 22, and the genes for GP IX and GP V
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 743

ristocetin and have diminished response to thrombin.2,8,10 The

lack of response to ristocetin is due to the lack of GP Ib/IX/V
complexes and the inability of BSS platelets to bind VWF. Lack
of binding to VWF also accounts for the inability of platelets to
adhere to exposed subendothelium and the resultant bleeding
characteristic of this disorder. This defect in adhesion shows
the importance of initial platelet attachment in primary hemo-
stasis. In many respects, this disorder resembles the defect
seen in von Willebrand disease (VWD). In contrast to VWD,
however, this abnormality cannot be corrected by the addition
of normal plasma or cryoprecipitate, which is consistent with a
defect that resides in the platelets.

Treatment.  There is no specific treatment for BSS. Plate-

let transfusions are the therapy of choice, but patients invari-
ably develop alloantibodies, so further platelet transfusion is
not possible. BSS patients tend to do better if apheresis plate-
Figure 41-3  ​Giant platelets in Bernard-Soulier syndrome (peripheral lets are used for transfusion because this tends to limit the
blood, 31000).  (Modified from Carr JH, Rodak BF: Clinical hematology atlas, number of donors to which the patient is exposed, and the rate
ed 3, Philadelphia, 2009, Saunders.) of alloimmunization tends to be lower.8,10 Other treatments
that have been used include desmopressin acetate (DDAVP)
and, more recently, recombinant factor VIIa (rVIIa, NovoSeven,
are located on chromosome 3. For surface expression of the Novo Nordisk Inc, Princeton, NJ).
GP Ib/IX complex, it seems that synthesis of three proteins,
GP Iba, GP Ibb, and GP IX, is required. Only GP V can be Inherited Giant Platelet Syndromes
expressed alone in significant quantities on the surface of In addition to BSS, there are several other inherited giant plate-
platelets, but the expression seems to be enhanced if the rest of let syndromes. See Table 40-1 and Mhawech and Saleem40 for
the complex is present. The most frequent forms of BSS involve a more complete discussion of these syndromes. Box 41-2
defects in GP Iba synthesis or expression (Table 40-1). The provides a listing of the inherited giant platelet syndromes.
presence of GP Iba is essential to normal function because it
contains binding sites for VWF and thrombin. Defects in the Giant Platelets with Velocardiofacial Syndrome
GP Ibb and GP IX genes also are known to result in BSS.32-34 This disorder is considered an autosomal recessive heterozy-
Diseases causing mutations include missense, frameshift, and gous variant of BSS. It is characterized by mild thrombocyto-
nonsense mutations.35,36 penia (with platelet counts ranging from 100 to 220 3 109/L),
giant platelets, and association with velocardiofacial syn-
Variants.  Several unusual variants of BSS have been de- drome (VCFS). The clinical picture of VCFS includes velopha-
scribed in which all or most of the GP Ib/IX/V complex is pres- ryngeal insufficiency, conotruncal heart disease, and learning
ent, but mutations that affect binding domains affect interactions disabilities. VCFS is considered to be a milder form of Di-
between elements of the complex, or result in truncation of a George syndrome (thymic hypoplasia, conotruncal cardiac
specific protein in the complex. In these cases, the complex fails defects, and cardiac abnormalities). No bleeding symptoms
to bind VWF or does so poorly.33 In rare circumstances, an anti-
body to GP Ib/V can cause a Bernard-Soulier–like syndrome
(pseudo-BSS), in which the GP Ib/IX/V complex is nonfunctional.
BOX 41-2  ​Inherited Giant Platelet Disorders
Mutations in the GP Ib/IX/V complex can also give rise to
Bernard-Soulier syndrome
gain of function and result in platelet-type VWD (pseudo-VWD).
Giant platelets with velocardiofacial syndrome
This gain of function results in spontaneous binding of plasma
Giant platelets with abnormal surface glycoproteins and mitral valve
VWF to the mutated GP Ib/IX/V complex. As a consequence,
insufficiency
platelets and large VWF multimers with their associated factor
Familial macrothrombocytopenia with GP IV abnormality
VIII are removed from the circulation, resulting in thrombocy-
Montreal platelet syndrome
topenia and reduced factor VIII clotting activity. GP Iba muta-
Gray platelet syndrome
tions that give rise to platelet-type VWD are 233 Gly n Val or
May-Hegglin anomaly
Ser and 239 Met n Val.37,38 Loss of residues 421 to 429 in GP
Fechtner syndrome
Iba also has been reported to result in platelet-type VWD.39
Sebastian syndrome
Hereditary macrothrombocytopenia
Laboratory Features.  BSS platelets have normal aggre-
Epstein syndrome
gation responses to adenosine diphosphate (ADP), epineph-
Mediterranean macrothrombocytopenia
rine, collagen, and arachidonic acid but do not respond to
744 PART VI  Hemostasis and Thrombosis

have been reported. Other than increased platelet size, no demonstrated and may result in defective regulation of bind-
ultrastructural abnormalities have been observed. GP Ibb is ing sites of platelets for adhesive proteins. As a result, platelet
mapped on chromosome 22q11.2,14, which is located in the binding sites may be abnormally exposed, leading to an abnor-
same region that is deleted in most patients with VCFS. mal binding of adhesive proteins to the exposed platelet sur-
Therefore, patients with a 22q11 deletion are obligate hetero- faces and spontaneous aggregation. The pathogenesis of giant
zygous carriers for the GP Ibb deletion and are heterozygotes platelets, severe thrombocytopenia, spontaneous aggregation,
for BSS. and the role of calpain remain to be clarified.

Giant Platelets with Abnormal Surface Glycoproteins Gray Platelet Syndrome

and Mitral Valve Insufficiency See a-Granule Deficiency: Gray Platelet Syndrome later in this
This extremely rare autosomal recessive disorder is charac- chapter for a discussion of this syndrome.
terized by thrombocytopenia with platelet counts ranging
from 50 to 60 3 109/L, giant platelets, and mild bleeding May-Hegglin Anomaly
symptoms in association with mitral valve insufficiency. May-Hegglin anomaly (MHA) is characterized by the presence
Platelets generally are larger than 20 mm. Other than in- of giant platelets, thrombocytopenia with platelet counts
creased platelet size, ultrastructural characteristics are nor- 60 to 100 3 109/L, Döhle body-like neutrophil inclusions, and
mal. These patients usually have a mild bleeding disorder mild bleeding symptoms (Figure 29-5). MHA is the most com-
expressed primarily as ecchymosis and epistaxis occurring mon of the inherited giant platelet disorders and is inherited
early in childhood. This disorder is distinguished from as an autosomal dominant trait. There appear to be two popu-
other inherited giant platelet syndromes by mitral valve in- lations of platelets in MHA. Normal-sized platelets have nor-
sufficiency. Platelet surface glycoproteins GP Ia, GP Ic, and mal ultrastructure and function. The large platelets show clear
GP IIa are absent, while GP Ib, GP IIb, and GP IIIa are nor- evidence of abnormal distribution of the platelet microtubule
mal. The surface glycoproteins absent in this disorder do not system, and this may be responsible for the large size and de-
seem to have a clear role. However, the abnormal bleeding fective platelet function that could lead to bleeding symptoms.
tendency clearly reflects a platelet dysfunction that could The cause of the thrombocytopenia is unknown. Chapter 40
result from a defect in glycoprotein composition, the ab- provides a detailed discussion of this disorder.
sence or defective function of an anchor protein necessary to
attach the glycoproteins to the cytoskeleton, or both. The Fechtner Syndrome
cause of large platelets is unclear. Fechtner syndrome is characterized by thrombocytopenia with
platelet counts ranging from 30 to 90 3 109/L, giant platelets,
Familial Macrothrombocytopenia and the development of deafness, cataracts, and nephritis. Its
with GP IV Abnormality mode of inheritance appears to be autosomal dominant. Glau-
Familial macrothrombocytopenia with GP IV abnormality is an coma and cataracts occur at an early age. Nephritis progresses
extremely rare, autosomal dominant disorder characterized by to end-stage renal failure by the age of 20 to 40 years, necessi-
thrombocytopenia, giant platelets, and variable degrees of tating hemodialysis and renal transplant. By the third decade
bleeding tendencies. The thrombocytopenia varies, and the of life, there is usually high-frequency hearing loss. Neutrophils
platelet count ranges from 45 3 109/L to normal. GP IV is and occasionally eosinophils contain one or more 1- to 2-mm
present in normal concentration, but there seems to be a defect irregularly shaped cytoplasmic inclusions that appear pale
in its glycosylation. GP IV seems to be involved in the early blue with Wright-Giemsa stain. They are smaller and less in-
stages of adhesion, and abnormal function may be related to tensely stained than the inclusions found in May-Hegglin
the mild bleeding tendency. Peripheral blood smears show anomaly. The thrombocytopenia in this disorder may be due
large platelets and neutrophils without inclusions. At this to ineffective megakaryocytopoiesis and thrombopoiesis as
point, the cause of large platelets remains to be clarified. reflected by large numbers of megakaryocytes, abnormal mor-
phology, and low platelet count.
Montreal Platelet Syndrome
Montreal platelet syndrome (MPS) is characterized by severe Sebastian Syndrome
thrombocytopenia, giant platelets, spontaneous platelet aggre- Sebastian syndrome is an extremely rare autosomal dominant
gation, and bleeding symptoms, including significant bruising thrombocytopenia characterized by giant platelets, neutro-
and episodes of hemorrhage. Of the inherited giant platelet phil inclusions, a mild bleeding disorder, and no other clini-
syndromes, MPS patients have the most severe thrombocytope- cal manifestations. The thrombocytopenia is mild with plate-
nia with platelet counts ranging from 5 to 40 3 109/L. Ultra- let counts ranging from 40 to 120 3 109/L. Mild bleeding
structurally, the platelets are large with no other abnormalities. such as epistaxis may occur in early childhood, but severe
Glycoprotein analysis is normal. It has been suggested that postoperative hemorrhage has been observed. Peripheral
calpain, which is known to be involved in the cleavage of cyto- blood smears show large platelets and faintly blue cytoplas-
skeleton proteins—in particular, actin-binding protein and mic inclusions in the neutrophils, similar in appearance to
talin—may have a role in spontaneous platelet aggregation. Döhle bodies. At the ultrastructural level, the platelets are en-
Low calpain proteolytic activity in MPS platelets has been larged but have normal structural elements. The pathogenesis
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 745

of the thrombocytopenia and the neutrophil inclusions re- spontaneous bruising. Petechiae are less common than in
mains to be clarified. other qualitative platelet disorders. Hemorrhage is rarely severe
but may be exacerbated by ingestion of aspirin or other anti-
Hereditary Macrothrombocytopenia platelet agents. In most of these disorders, the platelet count is
Hereditary macrothrombocytopenia is a rare disorder character- normal, and the bleeding time is usually, although not always,
ized by mild thrombocytopenia with platelet counts ranging prolonged. Platelet aggregation abnormalities are usually seen
from 50 to 123 3 109/L, giant platelets, bleeding tendency, and but vary, depending on the disorder.2,8,41,42
high-frequency hearing loss. It appears to be inherited in an
autosomal dominant fashion. These patients have a mild Storage Pool Diseases
bleeding tendency, the most frequent symptoms being gingival Platelet disorders of the storage pool type can be related
bleeding after brushing teeth, epistaxis, easy bruising, and to defects of the dense granules or defects of the a-granules.
menorrhagia. Bleeding symptoms appear early in childhood, Box 41-3 lists these disorders.
but hearing loss appears later. Ultrastructurally, the platelets
are large but have no additional abnormalities. Interestingly, Dense Granule Deficiencies.  The inheritance of dense
flow cytometry studies have shown a distinct population of granule deficiency does not follow a single mode, and it is likely
platelets in these patients that have glycophorin A on their that a variety of genetic abnormalities lead to the development
surface. Glycophorin A is generally considered an erythroid- of this disorder. Dense granule deficiencies can be subdivided
specific protein and is not expressed on normal platelets. How- into deficiency states associated with albinism and those in
ever, the pathogenesis of thrombocytopenia and giant platelets otherwise normal individuals (nonalbinos).
remains unclear, as does the presence of glycophorin A on the In the platelets of nonalbinos, there is evidence for the pres-
surface of the giant platelets in this disorder. ence of dense granule membranes in normal to near-normal
numbers, which suggests that the disorder arises from an in-
Epstein Syndrome ability to package the dense granule contents.43,44 Serotonin
Epstein syndrome is a very rare, autosomal dominant disorder accumulates in normal dense granules by an active uptake
characterized by thrombocytopenia, large platelets, and mild mechanism in which plasma serotonin is transported by a
bleeding diathesis, in association with nephritis and high- specific carrier-mediated system across the plasma membrane
frequency hearing loss. These patients also have persistent into the cytoplasm, and a second carrier-mediated system in
proteinuria, microscopic hematuria, and moderate hyperten- the dense granule membrane transports serotonin from the
sion. The mild bleeding tendency includes epistaxis, gastroin- cytoplasm to the interior of the dense granules.45 These trans-
testinal bleeding, and female genital tract bleeding. Bleeding port mechanisms are used in the serotonin release assay em-
symptoms occur early in life but may disappear later. The ployed to detect heparin-dependent antiplatelet antibodies
thrombocytopenia is often severe, with platelet counts ranging (Chapter 40). In addition to serotonin transport mechanisms,
from 30 to 60 3 109/L. The peripheral blood smear shows in- a nucleotide transporter MRP4 (ABCC4) that is highly ex-
creased platelet size in approximately 50% of the platelets. pressed in platelets and dense granules has been identified.
Ultrastructurally, the giant platelets are spherical and have a It would be expected that mutations in the gene for this
prominent surface-connected open canalicular system leading transporter could affect nucleotide accumulation in dense
to a sponge-like cytoplasm. The pathogenesis of the thrombo- granules.46
cytopenia and large platelets has not been determined. As an isolated abnormality, dense granule deficiency does
not typically result in a serious hemorrhagic problem. Bleeding
Mediterranean Macrothrombocytopenia is usually mild and most often is limited to easy bruisability.
Mediterranean macrothrombocytopenia is characterized by throm- Dense granule deficiency affects the results of platelet ag-
bocytopenia and large platelets. There is no bleeding or others gregation tests. Dense granules are intracellular storage sites for
symptom. This disorder has a high prevalence among persons ADP, adenosine triphosphate, calcium, pyrophosphate, and
originating from Greece, Italy, and the Balkan Peninsula. The serotonin. The contents of these granules are extruded when
incidence is unknown. The thrombocytopenia is mild, with platelet secretion is induced, and secreted ADP plays a major
platelet counts ranging from 89 to 290 3 109/L. The peripheral
blood smear shows large platelets, and electron microscopy
reveals no other abnormalities. Of unknown significance, sto-
matocytes are also present on peripheral blood smears. The
BOX 41-3  ​Platelet Storage Pool Diseases
pathogenesis of this disorder remains to be elucidated.
Dense granule deficiencies
Hermansky-Pudlak syndrome
Disorders of Platelet Secretion Chédiak-Higashi syndrome
(Release Reactions) Wiskott-Aldrich syndrome
Of the hereditary platelet function defects, disorders involving
Thrombocytopenia–absent radius (TAR) syndrome
storage pool defects and the release reaction are the most com-
a-Granule deficiencies
mon. The clinical features of this group of disorders are muco-
Gray platelet syndrome
cutaneous hemorrhage and hematuria, epistaxis, and easy and
746 PART VI  Hemostasis and Thrombosis

role in the propagation of platelet activation, recruitment, and episodes vary from mild to moderate but worsen as the platelet
aggregation and growth of the hemostatic plug. count decreases.10,20
In patients with dense granule deficiency, addition of Wiskott-Aldrich Syndrome.  This is a rare X-linked dis-
arachidonic acid to platelet-rich plasma fails to induce an ag- ease caused by mutations in the gene that encodes for a
gregation response (Chapter 42). Epinephrine and low-dose 502-amino acid protein—the Wiskott-Aldrich syndrome protein
ADP induce a primary wave of aggregation, but a secondary (WASp)—that is found exclusively in hematopoietic cells, in-
wave is missing. Responses to low concentrations of collagen cluding lymphocytes. WASp plays a crucial role in actin cyto-
are decreased to absent, but a high concentration of collagen skeleton remodeling. T cell function is defective due to abnor-
may induce a near-normal aggregation response.42,45 This ag- mal cytoskeletal reorganization, leading to impaired migration,
gregation pattern is caused by the lack of ADP secretion and is adhesion, and insufficient interaction with other cells. There is
almost identical to the pattern observed in patients taking a wide range of disease severity associated with Wiskott-
aspirin. Aldrich syndrome (WAS) gene mutations that range from the
Hermansky-Pudlak Syndrome.  In addition to occur- classic form of WAS with autoimmunity and/or malignancy to
ring as an isolated problem, dense granule deficiency is found a milder form with isolated microthrombocytopenia (X-linked
in association with several disorders. Hermansky-Pudlak syndrome thrombocytopenia; XLT) (Chapter 40) to X-linked neutropenia
is an autosomal recessive disorder characterized by tyrosinase- (XLN). Approximately 50% of patients with WAS gene muta-
positive oculocutaneous albinism, defective lysosomal func- tions have the WAS phenotype, and the other half have the XLT
tion in a variety of cell types, ceroid-like deposition in the cells phenotype. WAS gene mutations causing XLN are very rare.50
of the reticuloendothelial system, and a profound platelet Homozygous mutations of the WIPF1 gene on chromosome
dense granule deficiency.47 Several of the mutations responsi- 2 that encodes WASp-interacting protein (WIP)—a cytoplasmic
ble for Hermansky-Pudlak syndrome have been mapped to protein required to stabilize WASp—can also cause a WAS
chromosome 19. Mutations in at least seven genes individually phenotype.51
can give rise to Hermansky-Pudlak syndrome. These genes en- The classic form of WAS is characterized by susceptibility to
code for proteins that are involved in intracellular vesicular infections associated with immune dysfunction with recurrent
trafficking and are active in the biogenesis of organelles.48 bacterial, viral, and fungal infections, microthrombocytopenia,
Whereas the bleeding associated with most dense granule defi- and severe eczema. Thrombocytopenia is present at birth,
ciencies is rarely severe, Hermansky-Pudlak syndrome seems to but the full expression of WAS develops over the first 2 years
be an exception. Most bleeding episodes in Hermansky-Pudlak of life. Individuals with this disorder lack the ability to make
syndrome are not severe; however, lethal hemorrhage has been antipolysaccharide antibodies, which results in a propensity for
reported, and in one series hemorrhage accounted for 16% of pneumococcal sepsis. Patients with classic WAS tend to develop
deaths in patients with Hermansky-Pudlak syndrome. A unique autoimmune disorders, lymphoma, or other malignancies,
morphologic abnormality has been described in the platelets often leading to early death. Bleeding episodes are typically
of four families with Hermansky-Pudlak syndrome. This ab- moderate to severe. In WAS, a combination of ineffective
normality consists of marked dilation and tortuosity of the thrombocytopoiesis and increased platelet sequestration
surface-connecting tubular system (the so-called Swiss cheese and destruction accounts for the thrombocytopenia. As with
platelet).2,10,20,49 all X-linked recessive disorders, it is found primarily in
Chédiak-Higashi Syndrome.  This is a rare autosomal males.9,10,20,33,52
recessive disorder characterized by partial oculocutaneous Wiskott-Aldrich platelets are also structurally abnormal.
albinism, frequent pyogenic bacterial infections, giant lyso- The number of dense granules is decreased, and the platelets
somal granules in cells of hematologic (Figure 29-4) and are small, a feature of diagnostic importance. Other than in
nonhematologic origin, platelet dense granule deficiency, and WAS, such small platelets are seen only in TORCH (Toxoplasma,
hemorrhage. other agents, rubella virus, cytomegalovirus, herpesvirus) infec-
The Chédiak-Higashi syndrome protein gene is located on tions. Diminished levels of stored adenine nucleotides are re-
chromosome 13, and a series of nonsense and frameshift flected in the lack of dense granules observed on transmission
mutations all result in a truncated Chédiak-Higashi syndrome electron micrographs. The platelet aggregation pattern in WAS
protein that gives rise to a disorder of generalized cellular dys- is typical of a storage pool deficiency. The platelets show a
function involving fusion of cytoplasmic granules. decreased aggregation response to ADP, collagen, and epineph-
The disorder progresses to an accelerated phase in 85% of rine and lack a secondary wave of aggregation in response to
patients with Chédiak-Higashi syndrome and is marked by these agonists (Chapter 42). The response to thrombin is
lymphocytic proliferation in the liver, spleen, and marrow and normal, however.10,20 The most effective treatment for the
macrophage accumulation in tissues. During this stage, the thrombocytopenia seems to be splenectomy, which would be
pancytopenia worsens, which leads to hemorrhage and ever- consistent with peripheral destruction of platelets. Platelet
increasing susceptibility to infection; the result is death at transfusions may be needed to treat hemorrhagic episodes.
an early age. Initially, bleeding is increased because of dense Bone marrow transplantation also has been attempted with
granule deficiency and consequent defective platelet function. some success.20,53
During the accelerated phase, however, the thrombocytopenia Thrombocytopenia with absent radii syndrome.  TAR
also contributes to a prolonged bleeding tendency. Bleeding (Chapter 40) is a rare autosomal recessive disorder characterized
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 747

by the congenital absence of the radial bones (the most pro- autosomal dominant characteristic. In these patients, other
nounced skeletal abnormality), numerous cardiac and other membrane abnormalities also have been described.33
skeletal abnormalities, and thrombocytopenia (90% of cases). It Quebec platelet disorder is an autosomal dominant bleeding
is mentioned here because the platelets have structural defects in disorder that results from a deficiency of multimerin (a multi-
dense granules with corresponding abnormal aggregation re- meric protein that is stored complexed with factor V in
sponses. Marrow megakaryocytes may be decreased in number, a-granules) and shows protease-related degradation of many
immature, or normal.10,54 a-granule proteins, even though a-granule structure is main-
tained. Thrombocytopenia may be present, although it is not a
a-Granule Deficiency: Gray Platelet Syndrome.  The consistent feature.33
a-granules are the storage site for proteins (Chapter 13) pro-
duced by the megakaryocyte (e.g., platelet-derived growth factor, Thromboxane Pathway Disorders:
thrombospondin, and platelet factor 4) or present in plasma Aspirin-Like Effects
and taken up by platelets and transported to a-granules for stor- Platelet secretion requires the activation of several biochemical
age (e.g., albumin, immunoglobulin G [IgG], and fibrinogen). pathways. One such pathway is the one leading to thrombox-
There are 50 to 80 a-granules per platelet, which are primarily ane formation. A series of phospholipases catalyze the release
responsible for the granular appearance of platelets on stained of arachidonic acid and several other compounds from mem-
blood films. brane phospholipids. Arachidonic acid is converted to interme-
Gray platelet syndrome, a rare disorder first described in 1971, diate prostaglandins by cyclooxygenase and to thromboxane A2
is characterized by the specific absence of morphologically by thromboxane synthase (Figure 13-20). Thromboxane A2 and
recognizable a-granules in platelets. The disorder is inherited other substances generated during platelet activation cause
in an autosomal recessive fashion. Clinically, gray platelet syn- mobilization of ionic calcium from internal stores into the
drome is characterized by lifelong mild bleeding tendencies, cytoplasm, occupancy of several activation receptors, and initia-
moderate thrombocytopenia, fibrosis of the marrow, and large tion of a cascade of events resulting in secretion and aggregation
platelets whose gray appearance on a Wright-stained blood of platelets (Figure 13-21).60
film is the source of the name of this disorder.2,8,10,55 More re- Several acquired or congenital disorders of platelet secre-
cently, a mutation in region 3p21 involving the gene NBEAL2 tion can be traced to structural and functional modifications
has been identified. This gene is crucial in the development of of arachidonic acid pathway enzymes. Inhibition of cyclooxy-
a-granules.56,57 genase occurs on ingestion of drugs such as aspirin and ibupro-
In electron photomicrographs of platelets and megakaryocytes, fen. As a result, the amount of thromboxane A2 produced from
the platelets appear to have virtually no a-granules, although they arachidonic acid depends on the degree of inhibition. Throm-
do contain vacuoles and small a-granule precursors that stain boxane A2 is required for storage granule secretion and maximal
positive for VWF and fibrinogen. Other types of granules are platelet aggregation in response to epinephrine, ADP, and low
present in normal numbers. The membranes of the vacuoles concentrations of collagen.6,10,20,61
and the a-granule precursors have P-selectin (CD62) and GP Hereditary absence or abnormalities of the components of
IIb/IIIa, and these proteins can be translocated to the cell the thromboxane pathway are usually termed aspirin-like defects
membrane on stimulation with thrombin. This indicates that because the clinical and laboratory manifestations resemble
these structures are a-granules that cannot store the typical those that follow aspirin ingestion. Platelet aggregation re-
a-granule proteins. This may provide an explanation for the sponses are similar to those in dense granule storage pool dis-
observation that, in gray platelet syndrome, the plasma levels orders (see earlier). Unlike in storage pool disorders, however,
of platelet factor 4 and b-thromboglobulin are increased be- ultrastructure and granular contents are normal. Deficiencies of
cause while the proteins normally contained in a-granules are the enzymes cyclooxygenase and thromboxane synthase are
produced, storage in those granules is not possible. As a result, well documented, and dysfunction or deficiency of thrombox-
they are released into the circulation. Most patients develop ane receptors is known.44
early-onset myelofibrosis, which can be attributed to the in-
ability of megakaryocytes to store newly synthesized platelet- Inherited Disorders of Other Receptors
derived growth factors.44 and Signaling Pathways
Treatment of severe bleeding episodes may require platelet Collagen Receptors
transfusions. Few other treatments are available for these pa- The a2b1 (GP Ia/IIa) integrin is one of the collagen receptors
tients. Cryoprecipitate has been used to control bleeding. Des- in the platelet membrane. A deficiency of this receptor has
mopressin acetate was found to shorten the bleeding time and been reported in a patient who lacked an aggregation response
has been used as successful prophylaxis in a dental extraction to collagen, whose platelets did not adhere to collagen, and
procedure. Some authors believe that desmopressin acetate who had a lifelong mild bleeding disorder.62 A deficiency in
should be the initial therapy of choice.2,8,44,58,59 another collagen receptor, GP VI, also has been reported in
patients with mild bleeding. The platelets of these patients
Other Storage Pool Diseases.  A rare disorder in which failed to aggregate in response to collagen, and adhesion to
both a-granules and dense granules are deficient is known as collagen also was impaired.63 A family with gray platelet syn-
a-dense storage pool deficiency. It seems to be inherited as an drome and defective collagen adhesion has been described.
748 PART VI  Hemostasis and Thrombosis

Affected members of the family have a severe deficiency of lost, and phosphatidylserine and phosphatidylethanolamine
GP VI.64 flip to the outer leaflet and facilitate the assembly of clotting
factor complexes. The phospholipid flip is mediated by a calcium-
ADP Receptors dependent enzyme, scramblase.72 In Scott syndrome, platelet
Platelets seem to contain at least three receptors for ADP. plug formation (including adhesion, aggregation, and secre-
P2X1 is linked to an ion channel that facilitates calcium ion tion) occurs normally, but clotting factor complexes do not
influx. P2Y1 and P2Y12 (P2TAC) are members of the seven- assemble on the activated platelet surface, and thrombin gen-
transmembrane domain (STD) family of G protein–linked eration is absent or much reduced. Because lack of thrombin
receptors (Chapter 13). P2Y1 is thought to mediate calcium generation leads to inadequate fibrin, the platelet plug is not
mobilization and shape change in response to ADP. Pathol- stabilized, and a bleeding diathesis results.73,74
ogy of the P2Y1 receptor has not yet been reported. P2Y12
is thought to be responsible for macroscopic platelet aggrega- Stormorken Syndrome
tion and is coupled to adenylate cyclase through a G-inhibitory Lastly, Stormorken syndrome is a condition in which platelets are
(Gi) protein complex.65 Some patients have been reported to always in an “activated” state and express phosphatidylserine
have decreased platelet aggregation in response to ADP but nor- on the outer leaflet of the membrane without prior activation.
mal platelet shape change and calcium mobilization. These pa- It has been postulated that patients with this syndrome have a
tients have an inherited deficiency of the P2Y12 receptor.66-68 defective aminophospholipid translocase.75
Bleeding problems seem to be relatively mild in these patients,
but the only treatment for severe bleeding is platelet transfusion. Acquired Defects of Platelet Function
Therapeutic drugs have been developed with the target to in-
Epinephrine Receptors hibit platelet function. Other drugs and certain agents have
Congenital defects of the a2-adrenergic (epinephrine) receptor been identified that also affect platelet function. The agents
associated with decreased platelet activation and aggregation in that will be discussed are summarized in Table 41-1.
response to epinephrine are known. The receptors that mediate
aggregation in response to epinephrine, ADP, and collagen are Drug-Induced Defects
STD receptors, as are the protease-activated receptors (PARs) for Drugs That Inhibit the Prostaglandin Pathway.  Un-
thrombin. So far, defects in the PAR receptors have not been like inherited disorders of platelet function, which are rare,
described.33 acquired disorders of platelet function are commonly encoun-
tered. The most frequent cause of acquired platelet dysfunction
Calcium Mobilization Defects is drug ingestion, with aspirin and other drugs that inhibit
A group of intracellular defects that affect platelet function the platelet prostaglandin synthesis pathways being the most
includes defects in which all elements of the thromboxane common culprits.
pathway are normal, but insufficient calcium is released
from the dense tubular system, and the cytoplasmic concen-
tration of ionic calcium in the cytoplasm never reaches TABLE 41-1  ​Antiplatelet Agents
levels high enough to support secretion. This group of dis-
orders is often referred to as calcium mobilization defects. Drug Mechanism of Action
These represent a heterogeneous group of disorders in Therapeutic Antiplatelet Agents
which the defects reside in the various intracellular signal- Aspirin Irreversible inhibition of COX1
ing pathways, including defects in G protein subunits and Naproxen Reversible inhibition of COX1
phospholipase C isoenzymes.20,69,70 Sulfinpyrazone Reversible inhibition of COX1
Ibuprofen (and related) Reversible inhibition of COX1
Scott Syndrome Clopidogrel Irreversible inhibition of P2Y12 receptors
Scott syndrome is a rare autosomal recessive disorder of calcium- Prasugrel receptors Irreversible inhibition of P2Y12 receptors
induced membrane phospholipid scrambling (necessary for Ticagrelor receptors Reversible inhibition of P2Y12 receptors
coagulation factor assembly) and thrombin generation on Abciximab Inhibition of GP IIb/IIIa (aIIb/b3)
platelets. Platelets secrete and aggregate normally but do Eptifibatide Inhibition of GP IIb/IIIa (aIIb/b3)
not transport phosphatidylserine and phosphatidylethanol- Tirofiban Inhibition of GP IIb/IIIa (aIIb/b3)
amine from the inner leaflet to the outer leaflet of the plasma Dipyridamole Inhibition of PDE (and cAMP breakdown)
Aggrenox Inhibition of COX1 and PDE
membrane. This phospholipid “flip” normally occurs during
platelet activation and is essential for the binding of vitamin
K–dependent clotting factors. In the membrane of resting
Drugs with Antiplatelet Effects
Alcohol Inhibition of thromboxane synthesis (?)
platelets, phosphatidylserine and phosphatidylethanolamine
Nitrofurantoin Unknown
are restricted to the inner leaflet of the plasma membrane, and Dextrans Interference with membrane function
phosphatidylcholine is expressed on the outer leaflet. This Hydroxyethyl (HETA) Interference with membrane function
asymmetry is maintained by the enzyme aminophospholipid starch
translocase.71 When platelets are activated, the asymmetry is
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 749

A single 200-mg dose of acetylsalicylic acid (aspirin) can irrevers- the inhibition is reversible. These drugs are said to be competitive
ibly acetylate 90% of the platelet cyclooxygenase (Figure 13-20). inhibitors of cyclooxygenase, and as the blood concentration of the
In platelets the acetylated cyclooxygenase (cyclooxygenase-1, drug decreases, platelet function is recovered. This group of
or COX-1) enzyme is completely inactive. Platelets lack a drugs includes ibuprofen and related compounds, such as ketopro-
nucleus and cannot synthesize new enzymes. The inhibitory fen and fenoprofen, naproxen, and sulfinpyrazone. In contrast
effect is permanent for the circulatory life span of the platelet to aspirin, most of these agents have little effect on the platelet
(7 to 10 days). function tests (Chapters 42 and 44). Except for their potential
Endothelial cells synthesize new cyclooxygenase, and endo- to irritate the gastric mucosa, these drugs have not been re-
thelial cell cyclooxygenase seems to be less sensitive to aspirin ported to cause clinically important bleeding.6,11,20,80 Interest-
than the platelet enzyme, at least at low dosages. This has led ingly, ibuprofen appears to have a prothrombotic effect when
to the view that low dosages of aspirin may be better than ingested within 2 hours of aspirin because it blocks the acetyla-
higher dosages for cardiovascular protection, because platelet tion site for aspirin on COX-1. Patients taking aspirin should be
thromboxane production is inhibited, whereas endothelial cautioned to avoid ibuprofen and related drugs near the time of
cells recover prostacyclin production with its accompanying aspirin ingestion.
antiplatelet effects. Others argue that inhibition of platelet The association of chronic alcohol consumption with throm-
function is the more important effect and that higher dosages bocytopenia is well known. Chronic, periodic, and even acute
of aspirin are better for this purpose. For these reasons, there alcohol consumption may result in a transient decrease in
are wide-ranging opinions as to the optimal dosage of aspirin. platelet function, however, and the inhibitory effect seems to
What is lost in these arguments is that endothelial cells also be more pronounced when alcohol is consumed in excess.
produce another potent platelet inhibitor, nitric oxide (NO), Most patients who are scheduled to undergo a medical proce-
and its production is not affected by aspirin. Although aspirin dure in which there may be hemostatic challenge are advised
may inhibit a proaggregatory mechanism (thromboxane pro- to abstain from alcohol consumption for about 3 days before
duction) and an antiaggregatory mechanism (prostacyclin pro- the procedure. The impaired platelet function seems to be re-
duction) in endothelial cells, the NO platelet inhibitory mech- lated at least in part to inhibition of thromboxane synthesis. A
anism is not affected. reduced platelet count and impaired platelet function may
It may be necessary to define a test system to determine the contribute to the increased incidence of gastrointestinal hem-
optimal dosage of aspirin for cardiovascular protection on an orrhage associated with chronic excessive alcohol intake.6,20,81,82
individual basis because some patients have, or develop, aspirin
resistance, and a dosage that previously was sufficient to inhibit Drugs That Inhibit Membrane Function.  Many drugs
platelet function effectively may no longer be able to produce interact with the platelet membrane and cause a clinically sig-
that effect. In addition, unlike the practice with almost all nificant platelet function defect that may lead to hemorrhage.
other therapeutic agents, a single dose of aspirin is usually Some of these drugs are useful antiplatelet agents, whereas for
prescribed in a “one dose fits all” fashion (e.g., 325 mg) with- many other drugs, their effects on the platelet membrane are
out regard to the patient’s weight, age, health status, or other an adverse side effect.60
measurable parameters. This practice is based on the assump- P2Y12 (ADP) Receptor Inhibitors.  The thienopyridine de-
tion that the biologic effect will be the same in all patients. rivatives clopidogrel, prasugrel, their predecessor ticlopidine,
Evidence is emerging, however, that there are considerable in- and the nucleoside ticagrelor are antiplatelet agents that bind to
terindividual differences in the response to a single dose of the P2Y12 platelet receptor thus inhibiting platelet function.
aspirin.3,20,76-78 One study has shown that patients who do not These drugs are for treatment of patients with arterial occlusive
respond well to aspirin have worse cardiovascular outcomes disease for prevention of myocardial infarction, for patients
than patients who respond well.79 The VerifyNow (Accumet- with cerebrovascular disease for reduction of the risk of throm-
rics, San Diego, CA) is one system that provides measurement botic stroke, for stroke and myocardial infarction prophylaxis,
of a patient’s response to antiplatelet medication (Chapters 42 and for patients who are intolerant of aspirin.
and 44). Individual tests for aspirin, P2Y12 receptor inhibitors In contrast to the effects of aspirin, the effects of these
(clopidogrel, prasugrel, ticagrelor), or GP IIb/IIIa receptor in- agents do not reach a steady state for 3 to 5 days, although a
hibitors (abciximab, eptifibatide, tirofiban) are available on steady state can be reached sooner with a loading dose. As
the VerifyNow. prophylactic agents, they have been shown to be as efficacious
Individuals known to have a defect in their hemostatic as aspirin. P2Y12 inhibitors and aspirin are often used in com-
mechanism, such as a storage pool deficiency, thrombocytope- bination to prevent arterial thrombosis, primarily based on the
nia, a vascular disorder, or VWD, may experience a marked synergistic action of these two drugs, which inhibit platelet
increase in bleeding tendency after aspirin ingestion, and such function by different mechanisms.
individuals should be advised to avoid the use of aspirin and The mechanism of action for P2Y12 inhibitors is binding to
related agents.20 the platelet membrane STD receptors for ADP and the prevention
The list of drugs affecting the prostaglandin pathway that of ADP binding to those receptors.66 As with aspirin inhibition of
converts arachidonic acid to thromboxane is long and beyond cyclooxygenase, the effect of the irreversible thienopyridines on
the scope of this chapter. Many of these drugs inhibit cyclooxy- platelet recovery of function following drug cessation is 50% of
genase, but, unlike with aspirin and closely related compounds, normal at 3 days, and complete at 7 to 10 days.83
750 PART VI  Hemostasis and Thrombosis

The major effect of the binding of these drugs to P2Y12 re- enzymes, mutations that result in decreased function of one
ceptors appears to be inhibition of stimulus-response coupling or more of these enzyme have less impact, and the response
between those receptors and fibrinogen binding to GP IIb/IIIa. is much more uniform than clopidogrel. Pharmacogenetic
As a consequence, platelet activation and aggregation induced testing for mutations affecting prasugrel activation is not
by ADP are markedly inhibited, and responses to other aggre- recommended.
gating agents, such as collagen, are reduced. Ticagrelor (Brilinta) is a nuceloside and the newest of the
Clopidogrel (Plavix), a second-generation thienopyridine de- P2Y12 inhibitors. While its antiplatelet effect is similar, it has
rivative, is an effective antiplatelet agent for a variety of clinical two important differences from prasugrel and clopidogrel.
applications, though its clinical effectiveness varies from pa- First, it is not a pro-drug and therefore does not require bioac-
tient to patient based on metabolism to the active drug. Clo­ tivation. Because it is rapidly absorbed, its antiplatelet effect is
pidogrel is a pro-drug and requires conversion to the active predictable and achieved in a short period of time. In addition,
drug by the P450 enzyme systems of the liver. For clopidogrel, ticagrelor binds to a slightly different site on the P2Y12 (ADP)
the isoform of P450 involved in clopidogrel metabolism is receptor than clopidogrel or prasugrel. This difference results
2C19 (a.k.a. CYP2C19). There are numerous mutations in in reversible binding. Therefore, unlike clopidogrel and prasu-
CYP2C19 that result in decreased activity of the enzyme and grel whose effects are irreversible, platelet function returns
therefore inhibit the conversion of clopidogrel to the active quite rapidly with cessation of ticagrelor. However, this short
drug. CYP2C19*1/*1 (wild type) represents two normal func- half-life (7 to 9 hours) requires twice-daily dosing and may be
tioning alleles and normal metabolism of clopidogrel to the a compliance issue for some patients.
active drug. Hypofunctional alleles are CYP2C19*2 to *10, while GP IIb/IIIa (aIIb/b3) receptor inhibitors.  Another target
CYP2C19*17 is a hyperfunctional allele. Patients with one for antiplatelet agents to reduce cardiovascular thrombotic risk
of the mutations resulting in decreased activity (e.g., is the platelet membrane GP IIb/IIIa (aIIb/b3) receptor. Interfer-
CYP2C19*1/*2) are considered intermediate metabolizers, ence with the ability of this receptor to bind fibrinogen inhibits
and the usual dose of clopidogrel does not achieve the degree platelet aggregation stimulated in response to all of the usual
of platelet inhibition desired. These individuals remain at in- platelet aggregating agents. Results of platelet function studies
creased risk for thromboembolic events. Increasing the dose of on platelets from patients receiving therapeutic doses of these
clopidogrel may increase the degree of platelet inhibition, but drugs essentially mimic those of a mild form of Glanzmann
it does not decrease the thromboembolic risk. Those who have thrombasthenia.
two hypofunctional alleles (CYP2C19*2/*2, *2/*3, or any Two different types of agents are included in this group. The
combination of two hypofunctional alleles) are poor metabo- first such agent approved for clinical use in the United States
lizers and do not derive significant benefit from clopidogrel was the Fab fragment of the mouse/human chimeric monoclo-
therapy. Approximately 25% of individuals have one or two nal antibody 7E3 (c7E3 Fab; abciximab [ReoPro]), which binds
hypofuctional alleles of CYP2C19 and are considered to be to GP IIb/IIIa, prevents the binding of fibrinogen, and prevents
clopidogrel resistant. In contrast, those with a *17 allele are rapid platelet aggregation. Numerous studies have shown the efficacy
metabolizers and convert clopidogrel to the active drug at a of this drug as an antiplatelet and antithrombotic agent.
faster rate. This results in increased blood levels of the active The second type of agent in this group targets a GP IIb/IIIa
drug following a dose of clopidogrel and an increased risk of recognition site for an arginine–glycine–aspartic acid (RGD)
bleeding. Those with one normal allele and one *17 allele sequence found in fibrinogen and several adhesive proteins.
(*1/*17) are considered to be rapid metabolizers and those These agents bind to the RGD recognition site, prevent the
who are *17/*17 are ultra-rapid metabolizers. Finally, indi- binding of fibrinogen, and consequently prevent platelet ag-
viduals with one hypofunctional and one hyperfunctional allele gregation. These compounds are relatively easily synthesized.
(e.g., *2/*17) have normal to intermediate clopidogrel metabo- Tirofiban (Aggrastat) is a nonpeptide RGD mimetic, and eptifiba-
lism. There are a variety of molecular methods available to test tide (Integrilin) is a cyclic heptapeptide that is also an RGD
for the most common alleles of CYP2C19. Although the FDA mimetic. When the receptor site is occupied by the drug, GP
has recommended pharmacogenetic testing for these alleles, it is IIb/IIIa is no longer able to bind fibrinogen or other adhesive
not a common practice. proteins and is no longer functional as the aggregation receptor.
Clopidogrel has more effect on the platelet function tests The goal of therapy with these drugs is to induce a con-
than aspirin, although there is little difference in the risk of trolled thrombasthenia-like state. At present, these agents are
clinical bleeding.84 Clopidogrel can occasionally produce ma- primarily used in patients undergoing percutaneous coronary
jor side effects in some patients, including long-lasting neutro- intervention and are administered concurrently with heparin
penia, aplastic anemia, thrombocytopenia, gastrointestinal and other antiplatelet agents. The use of these agents is limited
distress, and diarrhea. by the need to administer them by constant intravenous infu-
Prasugrel (Effient) is a third-generation thienopyridine de- sion and by their short half-lives. There have been attempts to
rivative. It has the same mechanism of action as clopidogrel. It make an orally active agent of this type, but none has been
is also a pro-drug, but its metabolism to the active form does approved for use.85-87
not require CYP2C19. Instead, it is metabolized to the active
drug by several enzymes of the cytochrome P450 system, in- Other Therapeutic Drugs That Inhibit Platelet Function. 
cluding CYP3A4 and CYP2B6. Because it is activated by several Dipyridamole is an inhibitor of platelet phosphodiesterase, the
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 751

enzyme responsible for converting cyclic adenosine mono- The dextrans, another class of commonly used drugs, can
phosphate (cAMP) to AMP. Elevation of cytoplasmic cAMP inhibit platelet aggregation, and impair platelet procoagulant
is inhibitory to platelet function, and inhibition of phosphodi- activity when given as an intravenous infusion. These drugs
esterase allows the accumulation of cAMP in the cytoplasm have no effect on platelet function, however, when added di-
(Figure 13-21). Dipyridamole alone does not inhibit platelet rectly to platelet-rich plasma. Dextrans are partially hydro-
aggregation in response to the usual platelet agonists, but it lyzed, branched-chain polysaccharides of glucose. The two
promotes inhibition of agents that stimulate cAMP formation, most commonly used are dextran 70 (molecular mass of
such as prostacyclin, stable analogues of prostacyclin, and NO. 70,000 to 75,000 Daltons) and dextran 40 (molecular mass of
At one time, dipyridamole, alone or in combination with aspi- 40,000 Daltons), also known as low-molecular-weight dex-
rin, was widely used. By the 1990s, interest in dipyridamole tran. Both drugs are effective plasma expanders and are com-
had waned. There has been a resurgence of interest in dipy­ monly used for this purpose. Because of their effects on plate-
ridamole, however, as a combination agent compounded with lets, they have been extensively used as antithrombotic agents.
aspirin (Aggrenox). There does not seem to be any increased risk of hemorrhage
associated with the use of these agents, but their efficacy in
Miscellaneous Agents That Inhibit Platelet Function.  preventing postoperative pulmonary embolism is equal to that
Well known for their ability to interfere with platelet function of low-dose subcutaneous heparin.6,52,81,88
are antibiotics. Most of the drugs with this effect contain the Hydroxyethyl starch, or hetastarch, is a synthetic glucose poly-
b-lactam ring and are either a penicillin or a cephalosporin mer with a mean molecular mass of 450,000 Daltons that also
(Figure 41-4). These drugs can inhibit platelet function tests, is used as a plasma expander. It has effects similar to those of
but this effect is seen only in patients receiving large parenteral the dextrans. The mechanism of action of these drugs has not
doses and is thus only a problem for hospitalized patients. been clearly elucidated but is presumed to involve interaction
One postulated mechanism for the antiplatelet effect of these with the platelet membrane.6,52,81
drugs is that they associate with the membrane via a lipophilic Several other agents of diverse chemical structure and func-
reaction and block receptor-agonist interactions or stimulus- tion are known to inhibit platelet function. The mechanisms by
response coupling between receptors and fibrinogen binding which they induce platelet dysfunction are largely unknown.
to GP IIb/IIIa. They also may inhibit calcium influx in response Nitroglycerin, nitroprusside, propranolol, and isosorbide dini-
to thrombin stimulation, reducing the ability of thrombin to trate are drugs used to regulate cardiovascular function that
activate platelets. Although these drugs may prolong the bleed- seem to be able to cause a decrease in platelet secretion and
ing time test and in vitro aggregation responses to certain ago- aggregation. Patients taking phenothiazine or tricyclic antide-
nists, their association with a hemostatic defect severe enough pressants may have decreased secretion and aggregation re-
to cause clinical hemorrhage is uncertain and is not predicted sponses, but these effects are not associated with an increased
by the bleeding time test results.3,11,61,88 risk for hemorrhage. Local and general anesthetics may impair
Nitrofurantoin is an antibiotic that is not related to the in vitro aggregation responses. The same is true of antihista-
b-lactam drugs but may inhibit platelet aggregation when high mines. Finally, some radiographic contrast agents are known to
concentrations are present in the blood. This drug is not inhibit platelet function.88
known to cause clinical bleeding, however.88
Disorders That Affect Platelet Function
Myeloproliferative Neoplasms.  Chronic myeloprolif-
erative neoplasms (MPNs) include polycythemia vera, chronic
myelogenous leukemia, essential thrombocythemia, and myelofibro-
sis with myeloid metaplasia (Chapter 33). Platelet dysfunction is
a common finding in patients with these disorders. Hemor-
rhagic complications occur in about one third, thrombosis
occurs in another third, and, although it is uncommon, both
develop in some patients. These complications are serious
causes of morbidity and mortality.
Although the occurrence of hemorrhage or thrombosis in
MPN patients is largely unpredictable, certain patterns have
emerged. Hemorrhage and thrombosis are less common in
chronic myelogenous leukemia than in the other MPNs.
Bleeding seems to be more common in myelofibrosis with
myeloid metaplasia, but thrombosis is more common in the
other MPNs.
Abnormal platelet function has been postulated as a con-
Figure 41-4  ​Chemical structure of major classes of b-lactam antibiotics. tributing cause. This hypothesis is supported by the observa-
R, Nonspecified side chain.  (From Mahon CR, Lehman DE, Manuselis G: tion that bleeding is usually mucocutaneous in nature, and
Textbook of diagnostic microbiology, ed 4, St. Louis, 2011, Saunders.) thrombosis may be arterial or venous.
752 PART VI  Hemostasis and Thrombosis

In patients with these disorders, thrombosis may occur in Causes of platelet activation include adherence and aggrega-
unusual sites, including the mesenteric, hepatic, and portal tion of platelets to fibrinogen (adsorbed onto the surfaces of
circulations. Patients with essential thrombocythemia may the bypass circuit material), mechanical trauma and shear
develop digital artery thrombosis and ischemia of the fingers stresses, use of blood conservation devices and bypass pump-
and toes, occlusions of the microvasculature of the heart, priming solutions during surgery, hypothermia, complement
and cerebrovascular occlusions that result in neurologic activation, and exposure of platelets to the blood-air interface
symptoms.88 in bubble oxygenators.
In MPNs, a variety of platelet function defects have been Some degree of platelet degranulation typically is found
described, but their clinical importance is uncertain. Platelets after cardiac surgery using the cardiopulmonary bypass ma-
have been reported to have abnormal shapes, decreased proco- chine, which indicates that platelet activation and secretion
agulant activity, and a decreased number of secretory granules. have occurred during the operation. Platelet membrane frag-
In essential thrombocythemia, platelet survival may be short- ments, or “microparticles,” are found consistently in the blood
ened. The bleeding time is prolonged in only a few patients, of these surgical patients, providing additional evidence of the
and hemorrhage can occur in patients with a normal bleeding severe mechanical stress encountered by platelets during these
time. The risk of thrombosis or hemorrhage correlates poorly procedures.
with the elevation of the platelet count. The severity of the platelet function defect closely correlates
The most common abnormalities are decreased aggregation with the length of time on the bypass machine. After an un-
and secretion in response to epinephrine, ADP, and collagen.89 complicated surgical procedure, normal platelet function re-
Possible causes of the platelet dysfunction include loss of turns in about 1 hour, although the platelet count does not
platelet surface membrane a-adrenergic (epinephrine) recep- return to normal for several days. Thrombocytopenia is caused
tors, impaired release of arachidonic acid from membrane by hemodilution, accumulation of platelets on the surfaces of
phospholipids in response to stimulation by agonists, im- the bypass materials, sequestration or removal of damaged
paired oxidation of arachidonic acid by the cyclooxygenase platelets by the liver and reticuloendothelial system, and con-
and lipoxygenase pathways, a decrease in the contents of dense sumption associated with normal hemostatic processes after
granules and a-granules, and loss of a variety of platelet mem- surgery.81,88
brane receptors for adhesion and activation. There seems to be
no correlation between a given MPN and the type of platelet Liver Disease.  Moderate to severe liver disease is reported
dysfunction observed, with the exception that most patients to be associated with a variety of hemostatic abnormalities,
with essential thrombocythemia lack an in vitro platelet ag- including reduction in clotting proteins, reduction of proteins
gregation response to epinephrine. This observation may be in the natural anticoagulant pathways, dysfibrinogenemia, and
helpful in the differential diagnosis.6,11,88,90 excessive fibrinolysis (Chapter 38). Mild to moderate throm-
bocytopenia is seen in approximately one third of patients
Multiple Myeloma and Waldenström Macroglobulin- with chronic liver disease in association with hypersplenism or
emia.  Platelet dysfunction is observed in approximately one as a result of alcohol toxicity.8,88
third of patients with IgA myeloma or Waldenström macroglobu- Abnormal platelet function test results seen in patients with
linemia, a much smaller percentage of patients with IgG multi- chronic liver disease include reduced platelet adhesion, abnor-
ple myeloma, and only occasionally in patients with monoclonal mal platelet aggregation (in response to ADP, epinephrine, and
gammopathy of undetermined significance. thrombin), abnormal platelet factor 3 (phospholipids) avail-
Platelet dysfunction results from coating of the platelet ability, and reduced procoagulant activity. An acquired storage
membranes by paraprotein and does not depend on the type pool deficiency also has been suggested. The abnormal platelet
of paraprotein present. In addition to interacting with plate- function in these patients may respond to infusion of desmo-
lets, the paraprotein may interfere with fibrin polymerization pressin acetate. It is unclear, however, whether desmopressin
and the function of other coagulation proteins. acetate provides a benefit in preventing bleeding in these
Almost all patients with malignant paraprotein disorders patients or is simply correcting an abnormal laboratory test
have clinically significant bleeding, but thrombocytopenia is result.
still the most likely cause of bleeding in these patients. Other In chronic alcoholic cirrhosis, the thrombocytopenia and
causes of bleeding include hyperviscosity syndrome, complica- platelet abnormalities may result from the direct toxic effects of
tions of amyloidosis (e.g., acquired factor X deficiency), and, in alcohol on bone marrow megakaryocytes. The severe bleeding
rare instances, presence of a circulating heparin-like anticoagu- diathesis associated with end-stage liver disease has many
lant or fibrinolysis.9,81,88 causes, such as markedly decreased or negligible coagulation
factor production, excessive fibrinolysis, dysfibrinogenemia,
Cardiopulmonary Bypass Surgery.  The use of the car- thrombocytopenia, and (occasionally) disseminated intravas-
diopulmonary bypass machine (CPB; heart-lung machine) during cular coagulation. Upper gastrointestinal tract bleeding is a
cardiac surgery induces thrombocytopenia and a severe platelet relatively common feature of cirrhosis, particularly alcoholic
function defect that assumes major importance in bleeding cirrhosis, and recombinant factor VIIa (rVIIa, NovoSeven,
after surgery. The function defect most likely results from plate- Novo Nordisk Inc, Princeton, NJ) has been shown to be effec-
let activation and fragmentation in the extracorporeal circuit. tive treatment in some patients.91
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 753

Uremia.  This is commonly accompanied by bleeding antibodies to fibrinogen, and this treatment then becomes
caused by platelet dysfunction. In uremia, guanidinosuccinic ineffective.93
acid (GSA) is present in the circulation in higher than normal
levels as a result of inhibition of the urea cycle. GSA is dialyz- Hyperaggregable Platelets
able, and dialysis (peritoneal dialysis or hemodialysis) is usu- Patients with a variety of disorders associated with thrombosis
ally effective in correcting the prolonged bleeding time test and or increased risk for thrombosis, including hyperlipidemia,
the abnormal platelet function characteristic of uremia. NO diabetes mellitus, peripheral arterial occlusive disease, acute
diffuses into platelets; activates soluble guanylate cyclase; arterial occlusion, myocardial infarction, and stroke, have been
and inhibits platelet adhesion, activation, and aggregation.92 reported to have increased platelet reactivity. Platelets from
Because GSA is an NO donor, NO is present in the circulation these patients tend to aggregate at lower concentrations of ag-
at higher than normal levels in uremia. Abnormal platelet gregating agents than do platelets from individuals without
function is far more common than clinically significant bleed- these conditions.
ing in uremic patients.2,60,62 Sticky platelet syndrome is an inherited disorder with autoso-
Platelet aggregation pattern abnormalities are not uniform, mal dominant characteristics and is associated with venous and
and any combination of defects may be seen. There is evidence arterial thromboembolic events. The disorder is characterized
of a deficient release reaction, such as lack of primary ADP- by hyperaggregable platelets in response to ADP, epinephrine,
induced aggregation, and subnormal platelet procoagulant or both. In these patients, venous and/or arterial thrombotic
activity. The bleeding time test is characteristically prolonged events are often associated with emotional stress. Prophylactic
in uremia and seems to correlate with the severity of renal fail- treatment of these patients with low-dose aspirin reverses clini-
ure in these patients. There does not seem to be any significant cal symptoms and normalizes hyperaggregable responses to
correlation, however, between the bleeding time test and the aggregating agents in the laboratory.
risk of clinically significant bleeding. Anemia is an indepen- Spontaneous aggregation (aggregation in response to in vitro
dent cause of prolonged bleeding time, and the severity of stirring only) is also an indicator of abnormally increased
anemia in uremic patients correlates with the severity of renal platelet reactivity and often accompanies increased sensitivity
failure. Many uremic patients are treated with recombinant to platelet agonists. The presence of spontaneous aggregation
erythropoietin to increase their hematocrit. Maintenance of by itself is considered to be consistent with the presence of
the hematocrit at greater than 30% also may help to normalize a hyperaggregable state. Because participation of platelets
the bleeding times.2,81,84 is necessary for the development of arterial thrombosis, the
Bleeding is uncommon in uremic patients and is seen more presence of hyperaggregable platelets is often an indication
often with concurrent use of drugs that interfere with platelet that an antiplatelet agent should be used as part of a therapeu-
function or in association with heparin use in hemodialysis. tic or prophylactic regimen for arterial thrombosis.81,94,95
Platelet concentrates often are used to treat severe hemor- Acquired platelet function defects are seen occasionally in
rhagic episodes in patients with uremia but usually do not patients with autoimmune disorders, including systemic lupus
correct the bleeding. Other therapies that are sometimes erythematosus, rheumatoid arthritis, scleroderma, and the im-
effective include cryoprecipitate, desmopressin acetate, and mune thrombocytopenias, such as immune thrombocytopenic
conjugated estrogen.2,81,84 purpura.88
Purified fibrin degradation products can induce platelet
Hereditary Afibrinogenemia.  Hereditary afibrinogen- dysfunction in vitro. The pathophysiologic relevance of this
emia has been documented in more than 150 families. Al- observation is uncertain because the concentrations of fibrin
though it is not truly a platelet function disorder, platelets do degradation products required are unlikely to be reached in
not exhibit normal function in the absence or near-absence vivo. Patients with disseminated intravascular coagulation may
of fibrinogen. In most patients, the bleeding time is pro- have reduced platelet function, however, as a result of in vivo
longed, and because fibrinogen is essential for normal plate- stimulation by thrombin and other agonists resulting in in
let aggregation, platelet aggregation test results are abnormal. vivo release of granule contents. This has been called acquired
Abnormal results on platelet retention and adhesion studies storage pool disease; the term exhausted platelets may be more
involving the use of glass beads also have been documented. appropriate.96
In addition, results of all clot-based tests (including partial
thromboplastin time, prothrombin time, reptilase time,
VASCULAR DISORDERS
thrombin time, and whole-blood clotting time) are abnor-
mal. Addition of fibrinogen to samples or infusion of fibrino- The pathophysiology of disorders of vessels and their support-
gen into the patient results in correction of the abnormal test ing tissues is obscure. Laboratory studies of platelets and blood
results.2,93 coagulation usually yield normal results. The diagnosis is often
A high incidence of hemorrhagic manifestations is found based on medical history and is made by ruling out other
in patients with afibrinogenemia (or severe hypofibrinogen- sources of bleeding disorders. The usual clinical sign is the
emia). Bleeding is the cause of death in about one third of tendency to bruise easily or to bleed spontaneously, especially
such patients. Cryoprecipitate or fibrinogen concentrates can from mucosal surfaces. Vascular disorders are summarized in
be used to treat bleeding episodes. Some patients develop Box 41-4.
754 PART VI  Hemostasis and Thrombosis

and a bleeding diathesis. The hemangiomas are visceral or

BOX 41-4  ​Vascular Disorders3 subcutaneous, but rarely both. External hemangiomas may
become engorged with blood and resemble hematomas. Other
Hereditary Vascular Disorders well-recognized features of Kasabach-Merritt syndrome include
Hereditary hemorrhagic telangiectasia (Rendu-Osler-Weber acute or chronic disseminated intravascular coagulation
syndrome) (Chapters 38 and 39) and microangiopathic hemolytic anemia.
Hemangioma-thrombocytopenia syndrome (Kasabach-Merritt A hereditary basis for this syndrome has not been established,
syndrome) but the condition is present at birth. Several treatment modalities
Ehlers-Danlos syndrome and other genetic disorders are available for the angiomas and the associated coagulopathy
and range from corticosteroid therapy to surgery.10,98
Acquired Vascular Disorders
Allergic purpura (Henoch-Schönlein purpura) Ehlers-Danlos Syndrome and Other Genetic
Paraproteinemia and amyloidosis Disorders
Senile purpura Ehlers-Danlos syndrome may be transmitted as an autosomal
Drug-induced vascular purpuras dominant, recessive, or X-linked trait. It is manifested by
Vitamin C deficiency (scurvy) hyperextensible skin, hypermobile joints, joint laxity, fragile
tissues, and a bleeding tendency, primarily subcutaneous
Purpuras of Unknown Origin hematoma formation. Eleven distinct varieties of the disorder
Purpura simplex (easy bruisability) are recognized. The severity of bleeding ranges from easy
Psychogenic purpura bruisability to arterial rupture. The disorder generally can be
ascribed to defects in collagen production, structure, or cross-
linking, with resulting inadequacy of the connective tissues.
Hereditary Vascular Disorders Platelet abnormalities have been reported in some patients.10
Hereditary Hemorrhagic Telangiectasia Other inherited vascular disorders include pseudoxanthoma
(Rendu-Osler-Weber Syndrome) elasticum and homocystinuria (autosomal recessive disorders),
The mode of inheritance of hereditary hemorrhagic telangiectasia is and Marfan syndrome and osteogenesis imperfecta (autosomal
autosomal dominant. The vascular defect of this disorder is char- dominant disorders). In addition to vascular defects, Marfan
acterized by thin-walled blood vessels with a discontinuous endo- syndrome is characterized by skeletal and ocular defects.10
thelium, inadequate smooth muscle, and inadequate or missing
elastin in the surrounding stroma. Telangiectasias (dilated super- Acquired Vascular Disorders
ficial blood vessels that create small, focal red lesions) occur Allergic Purpura (Henoch-Schönlein Purpura)
throughout the body but are most obvious on the face, lips, The term allergic purpura or anaphylactoid purpura generally is ap-
tongue, conjunctiva, nasal mucosa, fingers, toes, and trunk and plied to a group of nonthrombocytopenic purpuras characterized
under the tongue. The lesions blanch when pressure is applied. by apparently allergic manifestations, including skin rash and
The disorder usually becomes manifest by puberty and progresses edema. Allergic purpura has been associated with certain foods
throughout life. Telangiectasias are fragile and prone to rupture. and drugs, cold, insect bites, and vaccinations. The term Henoch-
Epistaxis is an almost universal finding, and symptoms almost Schönlein purpura is applied when the condition is accompanied
always worsen with age. The age at which nosebleeds begin is by transient arthralgia, nephritis, abdominal pain, and purpuric
a good gauge of the severity of the disorder. Although the oral skin lesions, which are frequently confused with the hemorrhagic
cavity, gastrointestinal tract, and urogenital tract are common sites rash of immune thrombocytopenic purpura.2,10,52
of bleeding, bleeding can occur in virtually every organ.97 General evidence implicates autoimmune vascular injury,
The diagnosis of hereditary hemorrhagic telangiectasia is based but the pathophysiology of the disorder is unclear. Preliminary
on the characteristic skin or mucous membrane lesions, a history evidence indicates that the vasculitis is mediated by immune
of repeated hemorrhage, and a family history of a similar disorder. complexes containing IgA antibodies. It has been suggested
Patients with hereditary hemorrhagic telangiectasia do well that allergic purpura may represent autoimmunity to compo-
despite the lack of specific therapy and the seriousness of their nents of vessel walls.2,10
hemorrhagic manifestations.2,10,97 There are several other disor- Henoch-Schönlein purpura is primarily a disease of chil-
ders and conditions in which telangiectasias are present, includ- dren, occurring most commonly in children 3 to 7 years of age.
ing cherry-red hemangiomas (common in older men and It is relatively uncommon among individuals younger than age
women), ataxia-telangiectasia (Louis-Bar syndrome), and chronic 2 and older than age 20. Twice as many boys as girls are
actinic telangiectasia; they also are seen in association with affected. The onset of the disease is sudden, often following
chronic liver disease and pregnancy.97 an upper respiratory tract infection. The infectious organism
may damage the endothelial lining of blood vessels, which results
Hemangioma-Thrombocytopenia Syndrome in vasculitis. Attempts have been made to implicate a specific in-
(Kasabach-Merritt Syndrome) fectious agent, particularly b-hemolytic streptococcus.2,10
Kasabach and Merritt originally described the association of a giant Malaise, headache, fever, and rash may be the presenting
cavernous hemangioma (vascular tumor), thrombocytopenia, symptoms. The delay in the appearance of the skin rash often
 CHAPTER 41  Qualitative Disorders of Platelets and Vasculature 755

poses a difficult problem in differential diagnosis. The skin le- Amyloid is a fibrous protein consisting of rigid, linear, non-
sions are urticarial and gradually become pinkish, then red, branching, aggregated fibrils approximately 7.5 to 10 nm wide
and finally hemorrhagic. The appearance of the lesions may be and of indefinite length. Amyloid is deposited extracellularly
very rapid and accompanied by itching. The lesions have been and may lead to damage of normal tissues. Various proteins
described as “palpable purpura,” in contrast to the perfectly flat can serve as subunits of the fibril, including monoclonal light
lesions of thrombocytopenia and most other forms of vascular chains (l more frequently than k). Amyloidosis, the deposi-
purpura. These lesions are most commonly found on the feet, tion of abnormal quantities of amyloid in tissues, may be pri-
elbows, knees, buttocks, and chest. Ultimately, a brownish-red mary or secondary and localized or systemic. A discussion of
eruption is seen. Petechiae also may be present.2,10 the clinical spectrum of amyloidosis is beyond the scope of this
As the disease progresses, abdominal pain, polyarthralgia, chapter. Purpura, hemorrhage, and thrombosis may be a part
headaches, and renal disease may develop. Renal lesions are of the clinical presentation of patients with amyloidosis, how-
present in 60% of patients during the second to third week ever. Thrombosis and hemorrhage have been ascribed to amy-
of the disorder. Proteinuria and hematuria are commonly loid deposition in the vascular wall and surrounding tissues.
present.2,8,10 Platelet function has been shown to be abnormal in a few
The platelet count is normal. Tests of hemostasis, including cases, and in rare cases patients may have thrombocytopenia.
the bleeding time, and tests of blood coagulation, usually yield Current treatments for amyloidosis are not effective.100
normal results in patients with allergic purpura. Anemia gener-
ally is not present unless the hemorrhagic manifestations have Senile Purpura
been severe. The white blood cell count and the erythrocyte Senile purpura occurs more commonly in elderly men than in
sedimentation rate are usually elevated. The disease must be women and is due to a lack of collagen support for small blood
distinguished from other forms of nonthrombocytopenic pur- vessels and loss of subcutaneous fat and elastic fibers. The inci-
pura. Numerous infectious diseases that may be associated dence increases with advancing age. The dark blotches are flat-
with purpura also must be considered in the differential diag- tened, are about 1 to 10 mm in diameter, do not blanch with
nosis. Drugs or chemicals sometimes may be implicated.2,10 pressure, and resolve slowly, often leaving a brown stain in the
In the pediatric age group, the average duration of the initial skin (age spots). The lesions are limited mostly to the extensor
episode is about 4 weeks. Relapses are frequent, usually after a surfaces of the forearms and backs of the hands and occasion-
period of apparent well-being. Except for patients in whom ally occur on the face and neck. With the exception of increased
chronic renal disease develops, the prognosis is usually good. capillary fragility, results of laboratory tests are normal, and no
Occasionally, death from renal failure has occurred. Manage- other bleeding manifestations are present.2,10
ment is directed primarily at symptomatic relief, because there
currently is no effective treatment. Corticosteroids sometimes Drug-Induced Vascular Purpuras
have been helpful in alleviating symptoms. Most patients re- Purpura associated with drug-induced vasculitis occurs in the
cover without treatment.2,10 presence of functionally adequate platelets. A variety of drugs
are known to cause vascular purpura, including aspirin, warfarin,
Paraproteinemia and Amyloidosis barbiturates, diuretics, digoxin, methyldopa, and several antibi-
Platelet function can be inhibited by myeloma proteins. Abnor- otics. Sulfonamides and iodides have been implicated most
malities in platelet aggregation, secretion, and procoagulant activ- frequently. The lesions vary from a few petechiae to massive,
ity (Chapter 42) correlate with the concentration of the plasma generalized petechial eruptions. Mechanisms include develop-
paraprotein and are likely due to coating of the platelet mem- ment of antibodies to vessel wall components, development of
brane with the paraprotein. Under these conditions, platelet ad- immune complexes, and changes in vessel wall permeability. As
hesion and activation receptor functions are inhibited, and the soon as the disorder is recognized, the offending drug should
paraprotein coating also inhibits assembly of clotting factors on be discontinued. No other treatment is necessary.10
the platelet surface. High concentrations of paraprotein can cause
severe hemorrhagic manifestations as a result of a combination of Miscellaneous Causes of Vascular Purpura
hyperviscosity and platelet dysfunction. About one third of pa- Insufficient dietary intake of vitamin C (ascorbic acid) results
tients with IgA myeloma and Waldenström macroglobulinemia and in scurvy and decreased synthesis of collagen, with weakening
approximately 5% of patients with IgG myeloma (usually IgG3) of capillary walls and the appearance of purpuric lesions.10
exhibit platelet function abnormalities. Finally, the paraprotein A diagnosis of purpura simplex (simple vascular purpura) or
may contribute further to bleeding by inhibiting fibrin polymer- vascular fragility is made when a cause for purpura cannot be
ization. In these patients, there is poor correlation between abnor- found. The ecchymoses are superficial, bleeding is usually
mal results on laboratory tests (e.g., prothrombin time, activated mild, and laboratory test results are most often normal.10 Cu-
partial thromboplastin time, thrombin time, bleeding time) and taneous bleeding and bruising through intact skin has been
evidence of clinical bleeding. Treatment for the bleeding compli- observed in patients in whom no vascular or platelet dysfunc-
cations of these disorders is primarily reduction in the level of the tion can be detected. Most such cases involve women with
paraprotein. This can be accomplished quickly, albeit transiently, emotional problems, and the bruising is often accompanied by
by plasmapheresis. Longer-term treatment is usually chemother- nausea, vomiting, or fever. Evidence for a psychosomatic origin
apy for the underlying plasma cell malignancy.88,99 is equivocal. Laboratory test results are invariably normal.10
756 PART VI  Hemostasis and Thrombosis

SUMMARY

• Inherited qualitative platelet disorders can cause bleeding disor- • Drugs are the most common cause of acquired platelet dysfunc-
ders ranging from mild to severe. tion, and aspirin is the most frequent culprit. Several new classes
• Bernard-Soulier syndrome is caused by the lack of expression of of antiplatelet agents with effects different from aspirin are now
GP Ib/IX/V complexes on the platelet surface. This receptor com- available and gaining in popularity.
plex is responsible for platelet adhesion and its absence results in • A variety of pathologic conditions can result in platelet dysfunction
a severe bleeding disorder. and range from hematologic malignancies to kidney disease and
• Glanzmann thrombasthenia is caused by the lack of expression of liver disease.
GP IIb/IIIa complexes on the platelet surface. This complex is • Vascular disorders that result in bleeding are uncommon. There
known as the platelet aggregation receptor, and its absence is are a few well-recognized inherited disorders, however, such as
associated with a severe bleeding disorder. Ehlers-Danlos syndrome and hereditary hemorrhagic telangiecta-
• Storage pool disorders result from the absence of intraplatelet sia that can result in substantial blood loss.
a-granules, dense granules, or both. Platelet dysfunction associ- • Vascular disorders can be acquired, and these are much more
ated with these disorders is generally mild; bleeding symptoms common than inherited disorders. Causes range from the effects
also are usually mild. of aging to drug effects to allergic reaction.
• Aspirin-like effects result from defects in elements of the arachi-
donic acid metabolic pathway. Platelet dysfunction mimics that Now that you have completed this chapter, go back and
seen after aspirin ingestion. read again the case study at the beginning and respond
• Deficiencies of several of the receptors for platelet-activating to the questions presented.
substances have been documented, and bleeding symptoms of
varying severity are associated with these deficiencies.

R E V I E W Q UESTIONS

Answers can be found in the Appendix. 5. Which of the following is the most common of the heredi-
tary platelet function defects?
1. The clinical presentation of platelet-related bleeding may a. Glanzmann thrombasthenia
include all of the following except: b. Bernard-Soulier syndrome
a. Bruising c. Storage pool defects
b. Nosebleeds d. Multiple myeloma
c. Gastrointestinal bleeding
d. Bleeding into the joints (hemarthroses) 6. A mechanism of antiplatelet drugs targeting GP IIb/IIIa
function is:
2. A defect in GP IIb/IIIa causes: a. Interference with platelet adhesion to the subendothe-
a. Glanzmann thrombasthenia lium by blocking of the collagen binding site
b. Bernard-Soulier syndrome b. Inhibition of transcription of the GP IIb/IIIa gene
c. Gray platelet syndrome c. Direct binding to GP IIb/IIIa
d. Storage pool disease d. Interference with platelet secretion

3. Aspirin ingestion blocks the synthesis of: 7. The impaired platelet function in myeloproliferative neo-
a. Thromboxane A2 plasms results from:
b. Ionized calcium a. Abnormally shaped platelets
c. Collagen b. Extended platelet life span
d. ADP c. Increased procoagulant activity
d. Decreased numbers of a- and dense granules
4. Patients with Bernard-Soulier syndrome have which of the
following laboratory test findings? 8. Which is a congenital qualitative platelet disorder?
a. Abnormal platelet response to arachidonic acid a. Senile purpura
b. Abnormal platelet response to ristocetin b. Ehlers-Danlos syndrome
c. Abnormal platelet response to collagen c. Henoch-Schönlein purpura
d. Thrombocytosis d. Waldenström macroglobulinemia
 Thrombocytopenia 40
and Thrombocytosis
Larry D. Brace

OUTLINE OBJECTIVES
Thrombocytopenia: De- After completion of this chapter, the reader will be able to:
crease in Circulating
Platelets 1. Define thrombocytopenia and thrombocytosis, and 7. Differentiate between neonatal isoimmune thrombocy-
Impaired or Decreased state their associated platelet counts. topenia and neonatal autoimmune thrombocytopenia.
Platelet Production 2. Compare and contrast the clinical symptoms of platelet 8. Explain the laboratory findings and pathophysiology
Increased Platelet Destruc- disorders and clotting factor deficiencies. associated with thrombotic thrombocytopenic purpura
tion 3. Explain the primary pathophysiologic processes of and hemolytic uremic syndrome.
Abnormalities in Distribution thrombocytopenia. 9. Summarize the pathophysiology of thrombotic complica-
or Dilution 4. Name and list the unique diagnostic features of at least tions in heparin-induced thrombocytopenia and describe
Thrombocytosis: Increase four disorders included in congenital hypoplasia of the the sequence of treatment options.
in Circulating Platelets bone marrow and describe their inheritance patterns. 10. Given clinical history and laboratory test results for
Reactive (Secondary)
5. Differentiate between acute and chronic immune patients with thrombocytopenia or thrombocytosis,
Thrombocytosis
thrombocytopenia. suggest a diagnosis that is consistent with the infor-
Thrombocytosis Associated
with Myeloproliferative 6. Describe the immunologic and nonimmunologic mech- mation provided.
Disorders anisms by which drugs may induce thrombocytopenia.

CASE STUDY
After studying the material in this chapter, the reader should be able to respond to the following case study:

An 18-month-old African-American girl sustained severe

Patient Results Reference Range
burns over 40% to 50% of her body, including both lower
extremities. Within 1 month, she underwent a below-knee Protein C antigen 78% 70% to 137%
amputation of the left lower extremity. Over the next sev- Protein S antigen 120% 63% to 156%
eral years, she underwent skin-grafting surgeries, central Antithrombin activity 111% 76% to 136%
venous line placement, and other burn-related surgeries.
During these procedures, the patient was exposed to hepa- The patient’s results were normal on tests for the factor V
rinized saline irrigation. Four years after the burn injury, Leiden and prothrombin G20210A mutations, and she was
thrombosis was noted in the right femoral artery during a found not to have antiphospholipid antibody syndrome.
grafting surgery. Unfractionated heparin was used during Her platelet count had been decreasing steadily for 7 days
the surgery. Surgeons were unable to save the leg, and before surgery but was still within the reference range.
an above-knee amputation was necessary. At this time, 1. Is the heparin used during the grafting surgeries sig-
hypercoagulability studies were ordered. Results were as nificant in this patient’s case?
follows: 2. What test should be ordered next?

713
714 PART VI  Hemostasis and Thrombosis

B
leeding disorders resulting from platelet abnormali- Small-vessel bleeding in the skin attributed to thrombocytope-
ties, whether quantitative or qualitative, usually are nia is manifested by hemorrhages of different sizes (Figure 40-1).
manifested by bleeding into the skin or mucous Petechiae are small pinpoint hemorrhages about 1 mm in diame-
membranes or both (mucocutaneous bleeding). Common ter, purpura are about 3 mm in diameter and generally round,
presenting symptoms include petechiae, purpura, ecchymo- and ecchymoses are 1 cm or larger and usually irregular in shape.
ses, epistaxis, and gingival bleeding. Similar findings also are Ecchymosis corresponds with the lay term bruise. Other condi-
seen in vascular disorders, but vascular disorders (e.g., Ehlers- tions such as defective platelet function, vascular fragility, and
Danlos syndrome, hereditary hemorrhagic telangiectasia) are trauma contribute to the hemorrhagic state.
relatively rare. In contrast, deep tissue bleeding, such as he- Clinical bleeding varies and often is not closely correlated
matoma and hemarthrosis, is associated with clotting factor with the platelet count. It is unusual for clinical bleeding to
deficiencies. occur when the platelet count is greater than 50,000/mL, but
the risk of clinical bleeding increases progressively as the plate-
let count decreases from 50,000/mL. Patients with platelet
THROMBOCYTOPENIA: DECREASE IN
counts of 20,000/mL or sometimes lower may have little or no
CIRCULATING PLATELETS
bleeding symptoms. In general, patients with platelet counts of
Although the reference range for the platelet count varies fewer than 10,000/mL are considered to be at high risk for a
among laboratories, it is generally considered to be approxi- serious hemorrhagic episode.
mately 150,000 to 450,000/mL (150,000 to 450,000/mm3 or
150 to 450 3 109/L). Thrombocytopenia (platelet count of Impaired or Decreased Platelet Production
fewer than 100,000/mL) is the most common cause of clini- Abnormalities in platelet production may be divided into two
cally important bleeding. True thrombocytopenia has to be categories: One type is associated with megakaryocyte hypo-
differentiated from the thrombocytopenia artifact that can re- plasia in the bone marrow, and the other type is associated
sult from poorly prepared blood smears or automated cell with ineffective thrombopoiesis, as may be seen in disordered
counts when platelet clumping or platelet satellitosis are pres- proliferation of megakaryocytes.
ent (Chapters 15 and 16). The primary pathophysiologic pro-
cesses that result in thrombocytopenia are decreased platelet Inherited Thrombocytopenia/Congenital Hypoplasia
production, accelerated platelet destruction, and abnormal It is increasingly apparent that most inherited thrombocytope-
platelet distribution (sequestration) (Box 40-1). nias can be linked to fairly specific chromosomal abnormalities

BOX 40-1  ​Classification of Thrombocytopenia

Impaired or Decreased Production of Platelets Drug induced: immunologic

Congenital Heparin-induced thrombocytopenia
May-Hegglin anomaly Neonatal alloimmune (isoimmune neonatal) thrombocytopenia
Bernard-Soulier syndrome Neonatal autoimmune thrombocytopenia
Fechtner syndrome Posttransfusion isoimmune thrombocytopenia
Sebastian syndrome Secondary autoimmune thrombocytopenia
Epstein syndrome Nonimmune
Montreal platelet syndrome Thrombocytopenia in pregnancy and preeclampsia
Fanconi anemia Human immunodeficiency virus infection
Wiskott-Aldrich syndrome Hemolytic disease of the newborn
Thrombocytopenia with absent radii (TAR) syndrome Thrombotic thrombocytopenia purpura
Congenital amegakaryocytic thrombocytopenia Disseminated intravascular coagulation
Autosomal dominant and X-linked thrombocytopenia Hemolytic uremic syndrome
Neonatal Drug induced
Acquired
Viral Abnormalities of Distribution or Dilution
Drug induced Splenic sequestration
Kasabach-Merritt syndrome
Increased Platelet Destruction Hypothermia
Immune Loss of platelets: massive blood transfusions, extracorporeal
Acute and chronic immune thrombocytopenic purpura circulation

From Colvin BT: Thrombocytopenia, Clin Haematol 14:661-681, 1985; and Thompson AR, Harker LA: Manual of hemostasis and thrombosis, ed 3, Philadelphia, 1983,
FA Davis.
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 715

or specific genetic defects. Table 40-1 provides a list of inher-

ited thrombocytopenias associated with specific gene and
chromosomal abnormalities, mode of inheritance, and associ-
ated features.
Lack of adequate bone marrow megakaryocytes (mega-
karyocytic hypoplasia) is seen in a wide variety of congenital
disorders, including Fanconi anemia (pancytopenia), throm-
bocytopenia with absent radius (TAR) syndrome, Wiskott-
Aldrich syndrome, Bernard-Soulier syndrome, May-Hegglin
anomaly, and several other less common disorders. Although
thrombocytopenia is a feature of Bernard-Soulier syndrome
and Wiskott-Aldrich syndrome, the primary abnormality in
these disorders is a qualitative defect, and these disorders are
discussed in Chapter 41.

May-Hegglin Anomaly.  May-Hegglin anomaly is a rare

autosomal dominant disorder; the exact frequency is un-
known. Large platelets (20 mm in diameter) are present on the
peripheral blood film, and Döhle-like bodies are present in
A neutrophils (Figure 40-2) and occasionally in monocytes.
Other than the increase in size, platelet morphology is normal.
Thrombocytopenia is present in about one third to one half
of affected patients. Platelet function in response to platelet-
activating agents is usually normal. In some patients, mega-
karyocytes are increased in number and have abnormal ultra-
structure. Mutations in the MYH9 gene that encodes for
nonmuscle myosin heavy chain (a cytoskeletal protein in plate-
lets) have been reported.1 This mutation may be responsible for
the abnormal size of platelets in this disorder. Most patients are
asymptomatic unless severe thrombocytopenia is present, but
bleeding times may be prolonged in some patients in the
absence of bleeding complications (Table 40-1).
Three other disorders involving mutations of the MYH9
gene have been reported: Sebastian syndrome, Fechtner syn-
B drome, and Epstein syndrome.2 Sebastian syndrome is inher-
ited as an autosomal dominant disorder characterized by large
platelets, thrombocytopenia, and granulocytic inclusions. Sim-
ilar abnormalities are observed in Fechtner syndrome and are
accompanied by deafness, cataracts, and nephritis. In Epstein
syndrome, large platelets are associated with deafness, ocular
problems, and glomerular nephritis.3 These disorders are dis-
cussed in more detail in Chapter 41.

TAR Syndrome.  TAR syndrome is a rare autosomal re-

cessive disorder characterized by severe neonatal thrombocyto-
penia and congenital absence or extreme hypoplasia of the
radial bones of the forearms with absent, short, or malformed
ulnae and other orthopedic abnormalities. TAR syndrome is
associated with a mutation in the RBM8A gene located on the
C long arm of chromosome 1 or a 200 Kb deletion involving the
RBM8A gene (1q21.1). TAR can result from two deletions of
Figure 40-1  ​A, Petechiae, B, purpura, and C, ecchymoses indicate the the RBM8A gene, two mutations of the RBM8A gene, or, most
various patterns of systemic (mucocutaneous) hemorrhage. (Fig. 40-1A commonly, a combination of the two (Table 40-1). In addition
and 40-1C from Gary P. Williams, MD, University of Wisconsin Clinical
to bony abnormalities, patients tend to have cardiac lesions
Science Center, Madison, WI.) (Fig. 40-1B from Kitchens CS, Alving BM,
and a high incidence of transient leukemoid reactions with
Kessler CM: Consultative hemostasis and thrombosis, Philadelphia, 2002,
Saunders.) elevated white blood cell (WBC) counts (sometimes with
counts above 100,000/mL) in 90% of patients.4 Platelet counts
716 PART VI  Hemostasis and Thrombosis

TABLE 40-1  List of Inherited Thrombocytopenias

Frequency*/
Disease (abbreviation, Spontaneous Gene (chromosome
OMIM entry) Bleeding Inheritance localization) Other Features
SYNDROMIC FORMS
Wiskott-Aldrich syndrome 1111/yes XL WAS (Xp11) Severe immunodeficiency leading to death in
(WAS, 301000) infancy; small platelets
X-linked thrombocytopenia Mild immunodeficiency; small platelets
(XLT, 313900)
MYH9-related disease (MYH9-RD, nd) 1111/no AD MYH9 (22q12-13) Cataracts, nephropathy and/or deafness; giant
platelets; also non-syndromic
Paris-Trousseau thrombocytopenia 1111/yes AD Large deletion Cardiac and facial defects, developmental delay
(TCPT, 188025/600588), Jacobsen (11q23-ter) and/or other defects; large platelets
syndrome (JBS, 147791)
Thrombocytopenia with absent radii 1111/yes AR RBM8A (1q21.1) Platelet count tends to rise and often normalizes
(TAR, 274000) in adulthood; reduced megakaryocytes;
normal-sized platelets. Bilateral radial aplasia
and/or other malformations
GATA1-related disease (GATA1-RD) 11yes XL GATA1 (Xp11) Hemolytic anemia, possible unbalanced globin
(Dyserythropoietic anemia with chain synthesis, possible congenital erythro-
thrombocytopenia-nd, 300367- poietic porphyria; large platelets
X-linked thrombocytopenia with
thalassemia-XLTT, 314050)
Congenital thrombocytopenia with 1/yes AD HOXA11 (7p15-14) Radio-ulnar synostosis and/or other defects;
radio-ulnar synostosis possible evolution into aplastic anemia; normal
(CTRUS, 605432) sized platelets
Thrombocytopenia associated with 1/no AR ABCG5, ABCG8 (2p21) Anemia, tendon xanthomas, atherosclerosis;
sitosterolemia (STSL, 210250) large platelets; also non-syndromic
FLNA-related thrombocytopenia 1/yes XL FLNA (Xq28) Periventricular nodular heterotopia (MIM
(FLNA-RT, nd) 300049); large platelets; also non-syndromic
NON-SYNDROMIC FORMS
Bernard-Soulier syndrome
(BSS, 231200)
Biallelic 1111/yes AR GP1BA (17p13), Giant platelets
Monoallelic 111/no AD GP1BB (22q11), Large platelets
GP9 (3q21)
Congenital amegakaryocytic thrombo- 111/yes AR MPL (1p34) Always evolves into bone marrow aplasia in
cytopenia (CAMT, 604498) infancy; normal-sized platelets
Familial platelet disorder and predis- 11/no AD RUNX1 (21q22) High risk of developing leukemia or MDS; normal-
position to acute myelogenous sized platelets
leukemia (FPD/AML, 601399)
Gray platelet syndrome 11/yes AR NBEAL2 (3p21.1) High risk of developing evolutive myelofibrosis
(GPS, 139090) and splenomegaly; giant platelets
ANKRD26-related thrombocytopenia 11/no AD ANKRD26 (10p11-12) May be at risk of leukemia; normal-sized platelets
(THC2, 313900)
ITGA2B/ITGB3-related thrombocyto- 1/no AD ITGA2B (17q21.31), Large platelets
penia (ITGA2B/ITGB3-RT, nd) ITGB3 (17q21.32)
TUBB1-related thrombocytopenia 1/no AD TUBB1 (6p21.3) Giant platelets
(TUBB1-RT, nd)
CYCS-related thrombocytopenia 1/no AD CYCS (7p15.3) Normal-sized platelets
(THC4, 612004)

AD, Autosomal dominant; AR, autosomal recessive; MDS, myelodysplastic syndrome; nd, not defined; OMIM, Online Mendelian Inheritance in Man; XL, X-linked. Some forms are
categorized as both syndromic and non-syndromic. *1111, >100 families; 111, >50 families; 11 >10 families; 1, <10 families.
From: Balduini CL, Savoia A, Seri M. Inherited thrombocytopenias frequently diagnosed in adults. J Thromb Haemost 2013;11:1006-1019.
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 717

thrombocytopenias and is a relatively frequent form of

autosomal dominant thrombocytopenia.8 Bleeding in these
patients is usually absent or mild, and platelet function is
usually normal (Table 40-1).9,10

X-Linked Thrombocytopenia
X-linked thrombocytopenia can result from mutations in the
WAS (Wiskott-Aldrich syndrome) gene on the X chromosome
(Xp11) or mutations in the GATA1 gene, also on the X chromo-
some at Xp11.11-13 X-linked thrombocytopenias range from
mild thrombocytopenia and small platelets and absent or mild
bleeding to macrothrombocytopenia with severe bleeding
(Table 40-1).

Other Inherited Thrombocytopenias

In addition to the inherited thrombocytopenias discussed above,
there are several others that are due to gene mutations, including
Figure 40-2  ​Döhle body in segmented neutrophil and giant platelets HOXA11, ABCG5 and ABCG8, FLNA, RUNX1, ITGA2B, ITGB3,
associated with May-Hegglin anomaly (peripheral blood, 31000). (From Carr TUBB1, and CYCS (Table 40-1).
JH, Rodak BF: Clinical hematology atlas, ed 4, Philadelphia, 2013, Saunders.)
Neonatal Thrombocytopenia
are usually 10,000 to 30,000/mL in infancy. Interestingly, plate- Neonatal thrombocytopenia (platelet count ,150,000/mL) is
let counts usually increase over time, with normal levels often present in 1% to 5% of infants at birth. The causes of neonatal
achieved within 1 year of birth. thrombocytopenia are numerous as illustrated in Table 40-2.
In 75% of cases, the thrombocytopenia is present at or within
Fanconi Anemia.  Fanconi anemia is also associated with 72 hours of birth. Only a minority of these patients have immu-
thrombocytopenia, although other abnormalities are extensive, nologic disorders or coagulopathy causing thrombocytopenia.
including bony abnormalities, abnormalities of visceral organs, Causes of neonatal thrombocytopenia include infection with
and pancytopenia. Chapter 22 contains a more detailed Toxoplasma, rubella, cytomegalovirus (CMV), herpes (TORCH),
description. and human immunodeficiency virus (HIV), and in utero expo-
sure to certain drugs, particularly chlorothiazide diuretics and
Congenital Amegakaryocytic Thrombocytopenia the oral hypoglycemic tolbutamide and other agents. TORCH
Congenital amegakaryocytic thrombocytopenia is an autoso- infections cause thrombocytopenia with characteristically small
mal recessive disorder reflecting bone marrow failure.5 Affected platelets. CMV is the most common infectious agent causing
infants usually have platelet counts of fewer than 20,000/mL at congenital thrombocytopenia, with an overall incidence of 0.5%
birth, petechiae and evidence of bleeding at or shortly after to 1% of all births,14 but only 10% to 15% of infected infants
birth, and frequent physical anomalies. About half of the in- have symptomatic disease,15 which suggests that the incidence of
fants develop aplastic anemia in the first year of life, and there significant neonatal thrombocytopenia caused by CMV is about
are reports of myelodysplasia and leukemia later in childhood. 1 in 1000 infants. Although the mechanism of thrombocytope-
Allogeneic stem cell transplantation is considered curative for nia is not well understood, reports suggest that CMV inhibits
infants with clinically severe disease or aplasia.6 This disorder megakaryocytes and their precursors, which results in impaired
is caused by mutations in the MPL gene on chromosome 1 platelet production.16 About 1 in 1000 to 1 in 3000 infants
(1p34), resulting in complete loss of thrombopoietin receptor are affected by congenital toxoplasmosis. About 40% of such
function (Table 40-1). This loss of function results in reduced infants develop thrombocytopenia.17 While persistent thrombo-
megakaryocyte progenitors and high thrombopoietin levels.7 cytopenia is a prominent feature in infants with congenital
rubella syndrome, it is now rare in countries with organized
Autosomal Dominant Thrombocytopenia immunization programs.18,19 Thrombocytopenia also is a fea-
Autosomal dominant thrombocytopenia has been mapped to ture of maternal transmission of HIV to the neonate and is a sign
a mutation(s) in the ANKRD26 gene on the short arm of chro- of intermediate to severe disease.20
mosome 10 (10p11-12). Mutations in this gene appear to lead Maternal ingestion of chlorothiazide diuretics or tolbuta-
to incomplete megakaryocyte differentiation and the resultant mide can have a direct cytotoxic effect on the fetal marrow
thrombocytopenia. Platelet morphology and size are usually megakaryocytes. Thrombocytopenia may be severe, with plate-
normal. Until recently, autosomal dominant thrombocytope- let counts of 70,000/mL and sometimes lower. Bone marrow
nia was considered a very rare disorder. However, ANKRD26 examination reveals a marked decrease or absence of mega-
mutations have recently been found in 21 of 210 thrombocy- karyocytes. The thrombocytopenia develops gradually and is
topenic pedigrees. This indicates that ANKRD26 mutations slow to regress when the drug is stopped. Recovery usually
may be responsible for approximately 10% of inherited occurs within a few weeks after birth.10,21,22
718 PART VI  Hemostasis and Thrombosis

TABLE 40-2  Classification of Fetal and Neonatal thrombocytopenia later in this chapter). Another 10% to 15%
Thrombocytopenias of cases are due to disseminated intravascular coagulation
(DIC), almost always in very ill infants, particularly those
Fetal Alloimmune
with perinatal asphyxia or infections. Other examples include
Congenital infection (e.g., CMV, toxoplasma,
thrombosis, platelet activation, or immobilization at sites of
rubella, HIV)
inflammation (e.g., necrotizing enterocolitis). In very sick
Aneuploidy (e.g., trisomies 18, 13, 21, or triploidy)
infants, splenic sequestration may be a contributing factor to
Autoimmune (e.g., ITP, SLE)
thrombocytopenia.
Severe Rh hemolytic disease
Inherited thrombocytopenic syndromes are increasingly
Congenital/inherited (e.g., Wiskott-Aldrich syndrome)
being recognized as causes of neonatal thrombocytopenia
Early onset Placental insufficiency (e.g., preeclampsia, IUGR,
(Tables 40-1 and 40-2). Although considered to be rare, they
neonatal diabetes)
may be more common than once believed.
(,72 hours) Perinatal asphyxia
Perinatal infection (e.g., E. coli, group B streptococcus,
Acquired (Drug-Induced) Hypoplasia
Haemophilus influenzae)
A wide array of chemotherapeutic agents used for the treat-
DIC
ment of hematologic and nonhematologic malignancies
Alloimmune
suppress bone marrow megakaryocyte production and the
Autoimmune (e.g., ITP, SLE)
production of other hematopoietic cells. Examples include
Congenital infection (e.g., CMV, toxoplasma,
the commonly used agents methotrexate, busulfan, cytosine
rubella, HIV)
arabinoside, cyclophosphamide, and cisplatin. The resulting
Thrombosis (e.g., aortic, renal vein)
thrombocytopenia may lead to hemorrhage, and the platelet
Bone marrow replacement (e.g., congenital
count should be monitored closely. Drug-induced thrombo-
leukemia)
cytopenia is often the dose-limiting factor for many chemo-
Kasabach-Merritt syndrome
therapeutic agents. Recombinant interleukin-11 has been
Metabolic disease (e.g., propionic and
approved for treatment of chemotherapy-induced thrombo-
methylmalonic acidemia)
cytopenia, and thrombopoietin may prove to be useful for
Congenital/inherited (e.g., TAR, CAMT)
this purpose.23-25 Zidovudine (used for the treatment of HIV
Late onset Late onset sepsis
infection) is also known to cause myelotoxicity and severe
neonatal NEC
thrombocytopenia.26
(.72 hours) Congenital infection (e.g., CMV, toxoplasma,
Several drugs specifically affect megakaryocytopoiesis with-
rubella, HIV)
out significantly affecting other marrow elements. Anagrelide is
Autoimmune
one such agent, although its mechanism of action is unknown.
Kasabach-Merritt syndrome
This characteristic has made anagrelide useful for treating the
Metabolic disease (e.g., propionic and methylmalonic
thrombocytosis of patients with essential thrombocythemia
acidemia)
and other myeloproliferative disorders.27
Congenital/inherited (e.g., TAR, CAMT)
Ingestion of ethanol for long periods (months to years)
CAMT, Congenital amegakaryocytic thrombocytopenia; CMV, cytomegalovirus; may result in persistent severe thrombocytopenia. Although
DIC, disseminated intravascular coagulation; ITP, immune thrombocytopenic purpura; the mechanism is unknown, studies indicate that alcohol
IUGR, intrauterine growth restriction; NEC, necrotizing enterocolitis; SLE, systemic can inhibit megakaryocytopoiesis in some individuals. Mild
lupus erythematosus; TAR, thrombocytopenia with absent radii.
From: Roberts I, Murray NA. Neonatal thrombocytopenia: causes and management. thrombocytopenia is a common finding in alcoholic patients,
Arch Dis Child Fetal Neonatal Ed 2003;88:F359-F364. but other causes unrelated to ethanol use, such as portal hyper-
tension, splenomegaly, and folic acid deficiency, should be
excluded. The platelet count usually returns to normal within
While infectious agents and certain drugs are well-known a few weeks of alcohol withdrawal, but thrombocytopenia may
causes of neonatal thrombocytopenia, the overwhelming cause persist for longer periods. A transient rebound thrombocytosis
is impaired production. Most patients are preterm neonates may develop when alcohol ingestion is stopped.10
born after pregnancies complicated by placental insufficiency Interferon therapy commonly causes mild to moderate
and/or fetal hypoxia (preeclampsia and intrauterine growth thrombocytopenia, although under certain circumstances, the
restriction). These neonates have early-onset thrombocytope- thrombocytopenia can be severe and life-threatening. Inter-
nia and impaired megakaryopoiesis in spite of increased levels feron-a and interferon-g inhibit stem cell differentiation and
of thrombopoietin (Table 40-2). proliferation in the bone marrow, but the mechanism of action
Increased platelet consumption/sequestration is another is unclear.28
mechanism of neonatal thrombocytopenia accounting for Thrombocytopenia presumably caused by megakaryocyte
approximately 2% to 25% of neonatal thrombocytopenia. Of suppression also has been reported to follow the administration
these, 15% to 20% result from transplacental passage of of large doses of estrogen or estrogenic drugs such as diethylstil-
maternal alloantibodies and autoantibodies (see neonatal bestrol. Other drugs, such as certain antibacterial agents (e.g.,
alloimmune thrombocytopenia and neonatal autoimmune chloramphenicol), tranquilizers, and anticonvulsants, also have
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 719

been associated with thrombocytopenia caused by bone marrow thrombocytopenic only when production capacity is no longer
suppression.29-31 adequate to compensate for the increased rate of destruction.

Ineffective Thrombopoiesis Immune Mechanisms of Platelet Destruction

Thrombocytopenia is a usual feature of the megaloblastic ane- Immune (Idiopathic) Thrombocytopenic Purpura: Acute
mias (pernicious anemia, folic acid deficiency, and vitamin and Chronic.  The term idiopathic thrombocytopenic purpura
B12 deficiency). Quantitative studies indicate that, as with (ITP) was used previously to describe cases of thrombocytope-
erythrocyte production in these disorders, platelet production nia arising without apparent cause or underlying disease state.
is ineffective. Although the bone marrow generally contains Although the acronym for the disorder remains the same, the
an increase in the number of megakaryocytes, the total num- word idiopathic has been replaced by immune because of the
ber of platelets released into the circulation is decreased. realization that acute and chronic ITP are immunologically
Thrombocytopenia is caused by impaired DNA synthesis, mediated.
and the bone marrow may contain grossly abnormal mega- Acute ITP.  This is primarily a disorder of children,
karyocytes with deformed, dumbbell-shaped nuclei, some- although a similar condition is seen occasionally in adults. The
times in large numbers. Stained peripheral blood films reveal disorder is characterized by the abrupt onset of bruising, pete-
large platelets that may have a decreased survival time and chiae, and sometimes mucosal bleeding (e.g., epistaxis) in a
may have abnormal function. Thrombocytopenia is usually previously healthy child. The primary hematologic feature is
mild, and there is evidence of increased platelet destruction. thrombocytopenia, which frequently occurs 1 to 3 weeks after
Patients typically respond within 1 to 2 weeks to vitamin an infection.
replacement.22,32-34 The infection is most often a nonspecific upper respiratory
tract or gastrointestinal tract viral infection, but acute ITP also
Miscellaneous Conditions may occur after rubella, rubeola, chickenpox, or other viral ill-
Viruses are known to cause thrombocytopenia by acting on nesses and may follow live virus vaccination.37 The incidence
megakaryocytes or circulating platelets, either directly or in the of acute ITP is estimated to be 4 in 100,000 children, with a
form of viral antigen-antibody complexes. Live measles vaccine peak frequency in children between 2 and 5 years of age. There
can cause degenerative vacuolization of megakaryocytes 6 to is no sex predilection. In about 10% to 15% of the children
8 days after vaccination. Some viruses interact readily with initially thought to have acute ITP, the thrombocytopenia per-
platelets by means of specific platelet receptors. Other viruses sists for 6 months or longer, and these children are reclassified
associated with thrombocytopenia include CMV, varicella-zoster as having chronic ITP.38 The observation that acute ITP often
virus, rubella virus, Epstein-Barr virus (which causes infectious follows a viral illness suggests that some children produce an-
mononucleosis), and the virus that causes Thai hemorrhagic tibodies and immune complexes against viral antigens and
fever.10 that platelet destruction may result from the binding of these
Certain bacterial infections commonly are associated with antibodies or immune complexes to the platelet surface.
the development of thrombocytopenia. This may be the re- The diagnosis of acute ITP in a child with severe thrombo-
sult of toxins of bacterial origin, direct interactions between cytopenia almost always can be made without a bone marrow
bacteria and platelets in the circulation, or extensive damage examination. If the child has recent onset of bleeding signs and
to the endothelium, as in meningococcemia. Many cases of symptoms, otherwise normal results on complete blood count
thrombocytopenia in childhood result from infection. Pur- (for all red and white blood cell parameters and cell morphol-
pura may occur in many infectious diseases in the absence of ogy), and normal findings on physical examination (except for
thrombocytopenia, presumably because of vascular damage signs of bleeding), there is a high likelihood that the child has
(Chapter 41).10,35 ITP. In addition, if the bleeding symptoms develop suddenly
A common cause of unexplained thrombocytopenia is in- and there is no family history of hemorrhagic abnormalities or
filtration of the bone marrow by malignant cells with a pro- thrombocytopenia, the diagnosis of ITP is almost certain.
gressive decrease in marrow megakaryocytes as the abnormal There is, at present, no specific test that is diagnostic of acute
cells replace normal marrow elements. Inhibitors of thrombo- or chronic ITP.
poiesis may be produced by these abnormal cells and may In mild cases of acute ITP, patients may have only scattered
help to account for the thrombocytopenia associated with petechiae. In most cases of acute ITP, however, patients develop
conditions such as myeloma, lymphoma, metastatic cancer, fairly extensive petechiae and some ecchymoses and may have
and myelofibrosis.22,32,36 hematuria or epistaxis or both. About 3% to 4% of acute ITP
cases are considered severe, and typically generalized purpura
Increased Platelet Destruction is present, often accompanied by gastrointestinal bleeding,
Thrombocytopenia as a result of increased platelet destruction hematuria, mucous membrane bleeding, and retinal hemor-
can be separated into two categories: increased platelet destruc- rhage. Of patients with severe disease, 25% to 50% are consid-
tion caused by immunologic responses and increased destruc- ered to be at risk for intracranial hemorrhage, which is the
tion caused by mechanical damage or consumption or both. primary complication that contributes to the overall 1% to 2%
Regardless of the process, increased production is required to mortality rate for patients with acute ITP.38 Most patients with
maintain a normal platelet count, and the patient becomes life-threatening hemorrhage have a platelet count of less than
720 PART VI  Hemostasis and Thrombosis

4000/mL.39 Hemorrhage is rarely experienced by patients

whose platelet count exceeds 10,000/mL.
Most patients with acute ITP recover with or without treat-
ment in about 3 weeks, although for some, recovery may take
6 months. In a few children, recurrent episodes of acute ITP
are occasionally seen after complete recovery from the first
episode.40 Most patients with acute ITP have relatively mild
symptoms, and no treatment is needed. The most severe cases
may need to be treated, however, and intravenous immuno-
globulin (IVIG), platelet transfusions, and splenectomy (or
some combination of these) seem to offer the most immediate
benefit.37,38
Chronic ITP.  ​This disorder can be found in patients of
any age, although most cases occur in patients between the
ages of 20 and 50 years. Females with this disorder outnumber
males 2:1 to 3:1, with the highest incidence in women between Figure 40-3  ​Typical peripheral blood cell morphology in immune throm-
20 and 40 years of age. The incidence of chronic ITP ranges bocytopenic purpura. Note scarce platelets and increased platelet size but
normal red blood cell and leukocyte morphology (peripheral blood, 3500).
from 3.2 to 6.6 cases per 100,000 per year.41 Chronic ITP usu-
ally begins insidiously, with platelet counts that are variably
decreased and sometimes normal for periods of time. Present-
ing symptoms are those of mucocutaneous bleeding, with men-
orrhagia, recurrent epistaxis, and easy bruising (ecchymoses) smooth contour, and diminished cytoplasm are commonly
being most common. seen. In the absence of bleeding, infection, or other underlying
Platelet destruction in chronic ITP is the result of an immu- disorder, erythrocyte and leukocyte precursors are normal
nologic process. The offending antibodies attach to platelets, in number and morphology. Coagulation tests showing abnor-
and as a result, the antibody-labeled platelets are removed mal results include tests dependent on platelet function.
from the circulation by reticuloendothelial cells, primarily in the Although platelet-associated IgG levels are increased in most
spleen. Autoantibodies that recognize platelet surface glycopro- patients,10,21,45 it has not been shown conclusively that any
teins such as glycoprotein IIb (GP IIb) and GP IIIa (aIIb/b3), GP method of testing for platelet antibodies is sensitive or specific
Ia/IIa, and others can be demonstrated in 50% to 60% of ITP for ITP.
patients.42,43 Because megakaryocytes also express GP IIb/IIIa The initial treatment of chronic ITP depends on the urgency
and GP Ib/IX on their membranes, these cells are obvious tar- for increasing the platelet count. In ITP patients with platelet
gets of the antibodies. Platelet turnover studies have shown counts greater than 30,000/mL who receive no treatment, the
impaired platelet production in ITP. Overall, the life span of expected mortality rate is equal to that of the general popula-
the platelet is shortened from the normal 7 to 10 days to a few tion. Unless there are additional risk factors, ITP patients with
hours, and the rapidity with which platelets are removed from platelet counts greater than 30,000/mL should not be treated.
the circulation correlates with the degree of thrombocytopenia. If additional risk factors are present, such as old age, coagula-
If plasma from a patient with ITP is infused into the circulation tion defects, recent surgery, trauma, or uncontrolled hyperten-
of a normal recipient, the recipient develops thrombocytope- sion, the platelet count should be kept at 50,000/mL or higher,
nia. The thrombocytopenia-producing factor in the plasma of depending on the clinical situation. In patients in whom the
the ITP patient is an immunoglobulin G (IgG) antibody that need is considered urgent, IVIG remains the treatment of
can be removed from serum by adsorption with normal hu- choice. For most patients, however, the initial treatment of
man platelets. In addition, cytotoxic T cell-mediated lysis of chronic ITP consists principally of prednisone. About 70% to
platelets has been shown in vitro using CD31CD81 lympho- 90% of patients respond to this therapy, with an increase in
cytes from patients with active chronic ITP, although the in platelet count and a decrease in hemorrhagic episodes. Al-
vivo significance of this mechanism is not known.44 though reported response rates vary widely, about 50% of pa-
The only abnormalities in the peripheral blood of patients tients have a long-term beneficial effect from corticosteroid
with ITP are related to platelets. In most cases, platelets num- treatment.46 If the response to corticosteroids is inadequate or
ber between 30,000/mL and 80,000/mL. Patients with ITP no response is seen, steroid therapy can be supplemented with
undergo periods of remission and exacerbation, however, and IVIG or, in some cases, anti-D immunoglobulin.47 For patients
their platelet counts may range from near normal to fewer than in whom prednisone is ineffective, intravenous rituximab
20,000/mL during these periods (Figure 40-3). Morphologi- should be tried. Responses to rituximab are usually seen within
cally, platelets appear normal, although larger in diameter than 3 to 4 weeks. In some patients splenectomy may become neces-
usual. This is reflected in an increased mean platelet volume as sary. Splenectomy eliminates the primary site of platelet re-
measured by electronic cell counters. The marrow typically is moval and destruction, but it also removes an organ contain-
characterized by megakaryocytic hyperplasia. Megakaryocytes ing autoantibody-producing lymphocytes. Splenectomy is an
are increased in size, and young forms with a single nucleus, effective treatment for adult chronic ITP, with 88% of patients
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 721

showing improvement and 66% having a complete and lasting Acute ITP usually is self-limited, and spontaneous remis-
response.48 Vaccination with pneumococcal, meningococcal, sions occur in 80% to 90% of patients, although the duration
and Haemophilus influenzae vaccines should be performed at of the illness may range from days to months. In chronic ITP,
least 2 weeks prior to surgery. The use of laparoscopic surgery there is typically a fluctuating clinical course, with episodes of
speeds recovery and shortens hospitalization and is generally bleeding that last a few days or weeks, but spontaneous remis-
preferred to open splenectomy. In the most severe refractory sions are uncommon and usually incomplete.45
cases, immunosuppressive (chemotherapeutic) agents such as Symptoms of acute ITP vary, but petechial hemorrhages,
azathioprine given alone or with steroids may be necessary. In purpura, and often bleeding from the gums and gastrointesti-
such patients, platelet transfusions may be of transient benefit nal or urinary tract typically begin suddenly, sometimes over a
in treating severe hemorrhagic episodes but should not be few hours. Hemorrhagic bullae in the oral mucosa are often
given routinely.45 IVIG given alone or just before platelet trans- prominent in patients with severe thrombocytopenia of acute
fusion also may be beneficial.37,45 onset. Usually the severity of bleeding is correlated with the
Chronic ITP occurring in association with HIV infection, degree of thrombocytopenia.45 In contrast, presenting symp-
with hemophilia, or with pregnancy presents special problems toms of chronic ITP begin with a few scattered petechiae or
in diagnosis and therapy. Unexplained thrombocytopenia other minor bleeding manifestations. Occasionally, a bruising
in otherwise healthy members of high-risk populations may tendency, menorrhagia, or recurrent epistaxis is present for
be an early manifestation of acquired immune deficiency syn- months or years before diagnosis. Platelet counts range from
drome (AIDS).36,45 5000/mL to 75,000/mL and are generally higher than those in
Differentiation of Acute Versus Chronic Immune acute ITP. Giant platelets are commonly seen. Platelet-associated
Thrombocytopenic Purpura.  The differences between acute immunoglobulin levels are elevated in most patients, but the
and chronic ITP are summarized in Table 40-3. Acute ITP occurs test lacks sensitivity and specificity.45
most frequently in children 2 to 9 years of age and in young Treatment also varies for acute and chronic ITP. In chronic
adults, whereas chronic ITP occurs in patients of all ages, al- ITP, initial therapy often consists of glucocorticoids (corticoste-
though most frequently in adults aged 20 to 50 years, and more roids), which interfere with splenic and hepatic macrophages
commonly in women. Of patients with acute ITP, 60% to 80% to increase platelet survival time. If the ITP does not respond to
have a history of infection, usually viral (rubella, rubeola, chick- corticosteroids or the patient cannot tolerate them because of
enpox, and nonspecific respiratory tract infection), occurring 2 the resultant immunosuppression and toxicity, splenectomy
to 21 days before ITP onset. Acute ITP also may occur after im- may be necessary. In acute ITP, treatment for all but the most
munization with live vaccine for measles, chickenpox, mumps, severely thrombocytopenic and hemorrhagic patients is con-
and smallpox. traindicated. When treatment is necessary, a good response to
IVIG or corticosteroids or both usually can be obtained, and
splenectomy is rarely required.37,38
TABLE 40-3  ​Clinical Picture of Acute and Chronic
Immune Thrombocytopenic Purpura
Immunologic Drug-Induced Thrombocytopenia.  As
Characteristic Acute Chronic can be seen from Box 40-2, many drugs can induce acute
Age at onset 2–6 yr 20–50 yr thrombocytopenia. Drug-induced immune-mediated throm-
Sex predilection None Female over bocytopenia can be divided into several types based on the
male, 3:1 interaction of the antibody with the drug and platelets. Mecha-
Prior infection Common Unusual nisms of drug-antibody binding are shown in Figure 40-4.
Onset of bleeding Sudden Gradual Drug-dependent antibodies.  One mechanism of drug-
Platelet count ,20,000/mL 30,000–80,000/mL dependent antibodies is typified by quinidine- and quinine-
Duration 2–6 wk Months to years induced thrombocytopenia and has been recognized for more
Spontaneous remission 90% of patients Uncommon than 100 years. The antibody induced by drugs of this type
Seasonal pattern Higher incidence in None interacts with platelets only in the presence of the drug. Drug-
winter and spring dependent antibodies typically occur after 1 to 2 weeks of ex-
Therapy posure to a new drug. Many drugs can induce such antibodies,
Steroids 70% response rate 30% response rate but quinine, quinidine, and sulfonamide derivatives do so
Splenectomy Rarely required ,45 yr, 90% more often than other drugs. When antibody production has
response rate begun, the platelet count falls rapidly and often may be fewer
.45 yr, 40% than 10,000/mL. Patients may have abrupt onset of bleeding
response rate symptoms. If this type of drug-induced thrombocytopenia
develops in a pregnant woman, both she and her fetus may be
From Triplett DA, editor: Platelet function: laboratory evaluation and clinical application,
Chicago, 1978, American Society of Clinical Pathologists; Quick AJ: Hemorrhagic affected. Quinine previously was used to facilitate labor but is
diseases and thrombosis, ed 2, Philadelphia, 1966, Lea & Febiger; and Bussel J, no longer used for this purpose.
Cines D: Immune thrombocytopenia, neonatal alloimmune thrombocytopenia, and The initial studies of quinidine-induced thrombocytopenia
post-transfusion purpura. In Hoffman R, Benz EJ Jr, Shattil SJ, et al, editors: Hematology:
basic principles and practice, ed 3, New York, 2000, Churchill Livingstone, suggested that the drug first combines with the antibody and
pp 2096-2114. that the antigen-antibody (immune) complex then attaches to
722 PART VI  Hemostasis and Thrombosis

BOX 40-2  ​Common Drugs Causing Immune

Macrophage
Thrombocytopenia Fc receptor

Analgesics
Salicylates
Acetaminophen
Phenylbutazone
Fc receptor
Antibiotics
Cephalothin
Penicillin
Streptomycin
Platelet
Aminosalicylic acid
Rifampin
Novobiocin
Various sulfa drugs (chlorthalidone, furosemide)

Alkaloids
Quinidine Site on Fc segment of IgG binding to Fc receptors
Quinine
Structural antigenic determinant of the platelet membranes

Sedatives, Anticonvulsants Antigen of plasma immune complex

Methoin
Drug
Troxidone
Chlorpromazine Drug + platelet surface binding site
Diphenylhydantoin
Meprobamate Figure 40-4  ​Immunoglobulin binds a platelet membrane antigen or anti-
Phenobarbital gen and drug combination. Macrophage Fc receptors bind the Fc portion of
Carbamazepine the immunoglobulin. This may result in platelet removal and thrombocytope-
nia. IgG, immunoglobulin G. (From Rapaport SI: Introduction to hematology,
ed 2, Philadelphia, 1987, JB Lippincott, p. 489.)
Oral Hypoglycemics
Chlorpropamide
Tolbutamide usually the GP Ib/IX/V complex or the GP IIb/IIIa complex,
only in the presence of drug.49,50 The mechanism by which the
Heavy Metals drug promotes binding of a drug-dependent antibody to a
Gold specific target on the platelet membrane without covalently
Mercury linking to the target or the antibody remains to be deter-
Bismuth mined, however. Because the Fc portion of the immunoglobu-
Organic arsenicals lin is not involved in binding to platelets, it is still available
to the Fc receptors on phagocytic cells. This situation may
Miscellaneous contribute to the rapid onset and relatively severe nature of
Chloroquine the thrombocytopenia. Most drug-induced platelet antibodies
Chlorothiazide are of the IgG class, but in rare instances, IgM antibodies are
Insecticides involved.45
From Triplett DA, editor: Platelet function: laboratory evaluation and clinical ap- A similar pattern is seen with the antiplatelet/antithrom-
plication, Chicago, 1978, American Society of Clinical Pathologists; and Quick AJ: botic agents abciximab, tirofiban, and eptifibatide, although
Hemorrhagic diseases and thrombosis, ed 2, Philadelphia, 1966, Lea & Febiger. with these drugs thrombocytopenia tends to occur within
several hours of exposure. Such immediate reactions are due to
the platelet in an essentially nonspecific manner (the “inno- naturally occurring antibodies to the murine structural ele-
cent bystander” hypothesis). It now seems clear, however, that ments of abciximab (a mouse/human monoclonal antibody
the antibodies responsible for drug-induced thrombocytope- fragment) or to structural changes induced by binding of epti­
nia bind to the platelets by their Fab regions, rather than by fibatide or tirofiban to platelet GP IIb/IIIa.
attaching nonspecifically as immune complexes. The innocent Hapten-induced antibodies.  A second mechanism of
bystander/immune complex explanation for this type of drug- drug-induced thrombocytopenia is induction of hapten-
induced thrombocytopenia should be abandoned. The Fab por- dependent antibodies. Some drug molecules are too small by
tion of the antibody binds to a platelet membrane constituent, themselves to trigger an immune response, but they may act as
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 723

a hapten and combine with a larger carrier molecule (usually a IVIG may be used and is generally an effective treatment for
plasma protein or protein constituent of the platelet mem- most drug-induced immune thrombocytopenias. Laboratory
brane) to form a complex that can act as a complete antigen.45 testing to identify the specific drug involved is usually beyond
Penicillin and penicillin derivatives are the primary offending the capabilities of most laboratories. This type of testing is
agents causing drug-induced thrombocytopenia by this mecha- performed by many reference laboratories, however.
nism. Drug-induced thrombocytopenia of this type is often Immune complex–induced thrombocytopenia.  Heparin-
severe. The initial platelet count may be fewer than 10,000/mL induced thrombocytopenia (HIT) is a good example of another
and sometimes fewer than 1000/mL. The number of bone mar- type of drug-induced thrombocytopenia. Heparin binds to
row megakaryocytes is usually normal to elevated.45 Bleeding platelet factor 4 (PF4), a heparin-neutralizing protein made and
is often severe and rapid in onset, and hemorrhagic bullae in released by platelets (Figure 40-5). Binding of heparin by
the mouth may be prominent. plasma PF4 or platelet membrane–expressed PF4 causes a con-
Drug-induced autoantibodies.  Drug-induced autoanti- formational change in PF4, resulting in the exposure of neoepi-
bodies represent a third mechanism of drug-induced thrombocy- topes. Exposure of these neoepitopes (“new antigens”) stimu-
topenia. In this case, the drugs stimulate the formation of an lates the immune system of some individuals, which leads to
autoantibody that binds to a specific platelet membrane glycopro- the production of an antibody to one of the neoepitopes. In
tein with no requirement for the presence of free drug. Gold salts HIT, heparin and PF4 form a complex on the platelet surface or
and procainamide are two examples of such drugs. Levodopa circulating free complexes to which the antibody binds. The Fab
also may cause thrombocytopenia in the same way. The precise portion of the immunoglobulin molecule binds to an exposed
mechanism by which these drugs induce autoantibodies against neoepitope in the PF4 molecule; this leaves the Fc portion of
platelets is not known with certainty. the IgG free to bind with the platelet FcgIIa receptor, which
Treatment for any drug-induced thrombocytopenia is first causes platelet activation.51,52 Because the Fc portions of the IgG
to identify the offending drug, immediately discontinue its use, molecules bind to platelet FcgIIa receptor, they are not available
and substitute another suitable therapeutic agent. This is often to the Fc receptors of the cells of the reticuloendothelial system.
difficult to accomplish. Many patients are taking multiple This may explain the less severe decline in platelet count in this
drugs, and it is not always easy to determine which of the drugs thrombocytopenia. That does not mean, however, that the con-
is at fault for causing thrombocytopenia. Under these condi- sequences are less serious. The opposite may be true. Because
tions, identifying the causative agent may be a trial-and-error platelets are activated by occupancy of their FcgIIa receptor, in
procedure in which the most likely drugs are eliminated one at vivo platelet aggregation with thrombosis is possible. HIT
a time. In addition, even if the patient is taking only one agent, sometimes is referred to as heparin-induced thrombocytopenia
there may not be a suitable replacement, or a prolonged period and thrombosis (Chapter 39). Heparin binding to PF4 is required
may be required for the alternative drug to become effective. to expose the neoepitope to which the antibody binds. The
Drugs usually are cleared from the circulation rapidly, but treatment for HIT is to discontinue heparin administration and
dissociation of drug-antibody complexes may require longer replace it with another suitable anticoagulant. Low-molecular-
periods, perhaps 1 to 2 weeks.10 In some cases, such as those weight heparin should not be used as a heparin replacement
caused by gold salts, thrombocytopenia may persist for months. for this purpose, because the antibody cross-reacts with
Platelet transfusions may be necessary for patients with life- low-molecular-weight heparin and PF4 to result in platelet
threatening bleeds. Although it is true that the transfused platelets activation and aggregation.53
are destroyed rapidly, they may function to halt bleeding Heparin-induced thrombocytopenia.  ​HIT is a relatively
effectively before they are destroyed. In addition, high-dose common side effect of unfractionated heparin administration,

Circulating PF4- Circulating PF4-

heparin complexes heparin-Ab complexes

PF4 neoepitope + Immune Ab to PF4

and/or exposure response neoepitopes and/or

Platelet surface PF4- Platelet surface PF4-

heparin complexes heparin-Ab complexes

Ab Fc attaches to
Platelet agglutination, aggregation, and activation
platelet Fcγ receptor
occludes vessel with platelet thrombus

Figure 40-5  ​Heparin-induced thrombocytopenia with thrombosis. An antibody (Ab) binds the heparin–platelet factor 4 (PF4) complex in plasma or on the
platelet surface. The Fc portion binds platelet Fc receptors and activates the platelet. The activated platelets aggregate to form platelet thrombi in the arterial
circulation. Thrombi can also occur in the venous system.
724 PART VI  Hemostasis and Thrombosis

with about 1% to 5% of patients developing this complication. at a certain heparin dosage suddenly requires increasing amounts
Despite the thrombocytopenia, patients with HIT usually are of heparin to maintain the same level of anticoagulation. This
not at significant risk of bleeding, because the platelet count situation can result from in vivo activation of platelets and re-
typically does not fall below 40,000/mL. Ten percent to 30% of lease of PF4 and b-thromboglobulin from platelet a-granules.
patients with HIT develop thrombotic complications, however. Both of these substances neutralize heparin, which leads to a
In patients who develop HIT, heparin therapy should be normalization of results on the partial thromboplastin time test
stopped as soon as the diagnosis is made, because continued that is used to monitor heparin therapy. Heparin resistance often
heparin therapy can lead to significant morbidity and mortal- is seen before the development of thrombocytopenia.55
ity, including gangrene of the extremities, amputation, and A common benign form of HIT that occurs on heparin ad-
death. After discontinuation of heparin, the platelet count ministration is type I (non–immune mediated) HIT. It is im-
begins to increase and should return to normal within a few portant to distinguish benign type I from type II HIT. Type I HIT
days.54 is associated with a rapid decrease in the platelet count after
Because the immune system is involved in the development administration of heparin, but the thrombocytopenia is mild
of HIT, the clinical signs of HIT typically are not seen until 7 to (the platelet count rarely decreases to fewer than 100,000/mL)
14 days after the initiation of heparin therapy (the time neces- and transient, and the platelet count returns rapidly to the
sary to mount an immune response on first exposure to an preheparin level even if heparin therapy is continued. Careful
antigen). If the patient has been exposed to heparin previously, attention should be paid to the platelet count and other signs
however, symptoms of HIT may be seen in 1 to 3 days. Because of HIT so that this form of thrombocytopenia is not confused
the platelet count may fall sharply in 1 day, it is recommended with the clinically significant type II HIT. Although the mecha-
that platelet counts be measured daily in patients receiving nism of type I HIT has not been completely described, it may
unfractionated heparin therapy (Table 40-4). be related to the well-documented proaggregatory effects of
One other sign of impending HIT in some patients is the heparin.53,56 Because activated or aggregated platelets are
development of heparin resistance. This is the clinical situation in cleared from the circulation, these effects may explain the mild
which a patient who had experienced adequate anticoagulation decrease in the platelet count that occurs during the first few
days of heparin administration. This has not been clearly docu-
mented, however.
TABLE 40-4  ​Laboratory Tests for Heparin-Induced
The binding of heparin and related compounds depends on
Thrombocytopenia
polysaccharide chain length, composition, and degree of sul-
Laboratory Test Comment fation. Short-chain heparin polysaccharides (low-molecular-
Platelet count A .30% decrease from baseline weight heparin) have lower affinity to PF4 and are less prone
may signal HIT, even if still within to cause type II HIT. Pentasaccharide and its synthetic deriva-
the reference interval. tives (e.g., fondaparinux) do not seem to bind PF4 and are
unlikely to cause type II HIT.
Antigen Tests The detection of clinically significant HIT occurs by labora-
ELISAs: Use with a clinical scoring system tory testing using immunoassays and platelet function tests
GTI PVS:PF4* (Table 39-17). Can be used as a (Table 40-4 and Chapter 42). Laboratory testing, however, is
Stago H:PF4* first lab test to screen for the problematic because all tests lack sensitivity. Three methods
Hyphen BioMed presence of HIT antibodies; due to are commonly used, but all depend on the presence of free
Rapid tests (point-of-care): high frequency of false positive heparin–induced antiplatelet antibodies in the patient’s serum
Akers PIFA* results, functional tests should be or plasma in sufficient quantity to cause a positive test result.
DiaMed Pa-GIA performed to confirm a positive HIT can be detected by a platelet aggregation technique.57 In this
Coagulation instrument based tests: antigen test. False negative re- method, serum from the patient is added to platelet-rich
Milenia Biotec LFI-HIT sults can also occur. plasma from normal donors, heparin is added to the mixture,
HemosIL HIT-Ab and platelet aggregation is typically monitored for 20 minutes.
The specificity of the method is excellent (near 100%), but the
Functional Tests sensitivity is quite low (about 50%). The sensitivity of the test
Platelet Aggregation HIPA Important to perform functional test-
can be improved, but this requires the use of several heparin
Lumi-Aggregation HIPA ing that detects platelet activation
concentrations and platelet-rich plasma from two or more
Serotonin Release Assay by HIT antibodies to confirm a
blood donors, preferably of the same ABO blood type as the
diagnosis of HIT; sensitivities and
patient. In addition, the individuals donating blood for plate-
specificities differ among tests
let-rich plasma must not have taken aspirin for 10 to 14 days
with the SRA (washed platelet
before platelet donation because platelets from donors
assay) being the most sensitive.
who have ingested aspirin produce a false-negative result for
Require skill and experience of the
HIT.53 In the author’s experience, patients who develop HIT
operator to obtain quality results.
while receiving aspirin therapy rarely develop thrombotic
HIPA, Heparin-induced platelet aggregation. complications, and the drop in their platelet counts is less pre-
*FDA-cleared. cipitous, gradually decreasing over the course of several days
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 725

(unpublished observations). Given the efficacy of aspirin ther- heparin-dependent antiplatelet antibody (i.e., HIT). Under
apy in primary and secondary prevention of myocardial infarc- these same conditions, supratherapeutic concentrations of hep-
tion and thrombotic stroke, it is increasingly difficult to find arin do not activate platelets, however, and if platelets release
a sufficient pool of suitable (and willing) blood donors. the contents of dense granules at both therapeutic and suprath-
Although the platelet aggregation procedure is time consuming, erapeutic concentrations of heparin, the test result is not posi-
reasonable sensitivity can be obtained with sufficient attention tive for HIT. A similar phenomenon is observed in the test that
to the details of the technique.27 uses platelet aggregometry.53,59 The SRA is considered the gold
Dense granules of platelets contain serotonin, and platelets standard for detection of HIT. Its major drawback is the require-
have an active mechanism for rapid uptake and storage of ment for radiolabeled serotonin. Most clinical laboratories no
serotonin in dense granules. This property of platelets is used in longer use isotopic techniques. As a result, this test is performed
another test for HIT, the serotonin release assay (SRA), in which only in a few specialized centers. In addition, it has some of the
washed normal platelets from a donor are incubated with ra- same methodologic drawbacks as the platelet aggregation tech-
dioactive serotonin (Figure 40-6).58 Radioactive serotonin is nique, in particular the need for platelets from drug-free do-
taken up rapidly and stored in the dense granules of the donor nors. Nonetheless, when properly performed, this technique
platelets, which are washed and resuspended. In the presence of has similar specificity and superior sensitivity compared with
heparin-dependent antiplatelet antibody and heparin, the do- the platelet aggregation method.
nor platelets become activated and release the contents of their More recently, enzyme-linked immunosorbent assays (ELISAs)
dense granules when the concentration of heparin in the test have been developed based on the knowledge that the anti-
suspension is near the therapeutic range. The reappearance of genic target of the heparin-dependent antiplatelet antibody is
radioactive serotonin in the plasma indicates the presence of a a PF4-heparin complex (Figure 40-7). In this assay, PF4 and

No 3H-serotonin
release
Same ABO blood group (–)
as patient preferred

3H-serotonin
+ Therapeutic heparin
concentration release

3H-labeled +Patient
Donor PRP+ 3H-serotonin No 3H-serotonin HIT
donor platelets serum
release

+ Supertherapeutic
heparin concentration
3H-serotonin
release
(–)

Figure 40-6  ​The serotonin release assay (SRA) for heparin-induced thrombocytopenia (HIT). Donor platelets in platelet-rich plasma (PRP) are labeled with
tritiated (3H) serotonin, washed, and suspended in a buffer to which patient plasma is added. Heparin in therapeutic and saturating doses is added to two
aliquots. Release of radioactive serotonin in the therapeutic aliquot in combination with no release in the supratherapeutic system indicates HIT.

PF4-coated PF4-heparin PF4-heparin-Ab

microtiter Heparin complexes Patient complexes if Ab
plate wells on surfaces serum to PF4 is present

Enzyme-labeled
antihuman Ab
No color
(negative)

PF4-heparin-Ab-enzyme-labeled Substrate with

antihuman Ab complexes chromophore
Color development
if Ab to PF4 is present (positive)

Figure 40-7  ​Enzyme immunoassay for heparin-induced thrombocytopenia (HIT). The solid-phase target antigen is a complex of platelet factor 4 (PF4) and
heparin or a heparin surrogate. Anti–heparin-PF4 antibody in patient serum binds the antigen and is bound by enzyme-labeled anti–human antibody (Ab), a
“sandwich” assay. The enzyme catalyzes the release of a chromophore from its substrate.
726 PART VI  Hemostasis and Thrombosis

heparin (or a related compound) are coated to the surfaces of The most frequent cause of NAIT in whites is the HPA-1a
microplate wells. The serum or plasma from the patient sus- antigen expressed on GP IIIa of the surface membrane GP IIb/
pected to have HIT is added to wells of the microtiter plate. IIIa complex, followed by HPA-5b (Bra). The antigen HPA-3a
If the antibody is present, it adheres to the PF4-heparin (or (Baka) is present on GP IIb and is an important cause of neo-
heparin-like compound) complex. The plate wells are washed, natal thrombocytopenia in Asians. Platelet antigen HPA-4
and enzyme-labeled monoclonal antibodies against human (Penn and Yuk) accounts for the disorder in a few affected
IgG and IgM are added. After an appropriate incubation pe- neonates.
riod, the plate is washed, and a chromogenic substrate for the Clinically significant thrombocytopenia develops in an esti-
enzyme is added. Color development in the assay well indi- mated 1 in 1000 to 2000 newborns.62 With the first pregnancy,
cates the presence of the heparin-dependent antiplatelet anti- about 50% of neonates born to mothers lacking a specific
body in the patient specimen.60 This assay has greater sensitivity platelet antigen are affected, whereas with subsequent preg-
than the platelet aggregation method and similar sensitivity to nancies the risk is 75% to 97%.62 The incidence of intracranial
the SRA, but it has lower specificity than the serotonin release hemorrhage or death or both in affected offspring is about
assay or the platelet aggregation method. Unlike the functional 25%, and about half of the intracranial hemorrhages occur in
HIT assays (SRA and platelet aggregation), the ELISA can detect utero in the second trimester.
anti–PF4-heparin antibodies that are not pathologic; that is, the Affected infants may appear normal at birth but soon
test result is positive, but the patient does not have clinical HIT. manifest scattered petechiae and purpuric hemorrhages. In
Some ELISA assays can detect both IgG and IgM antibodies to many infants with NAIT, serious hemorrhage does not de-
heparin-PF4 complexes. More recently, ELISA kits that detect velop, however, and the infants recover over a 1- to 2-week
only IgG antibodies have been developed and have a better cor- period as the level of passively transferred antibody de-
relation with the SRA. The ELISA method is considerably less creases.10,38 In symptomatic cases, platelet levels are usually
labor intensive, does not require blood from healthy drug-free below 30,000/mL and may diminish even further in the first
donors, and can be performed in most laboratories. few hours after birth.
For patients who develop type II HIT, it is essential that The diagnosis of NAIT is one of exclusion of other causes
heparin therapy be withdrawn immediately. It is not prudent, of neonatal thrombocytopenia, including maternal ITP and
however, to discontinue administration of an anticoagulant/ maternal ingestion of drugs known to be associated with drug-
antithrombotic without substituting a suitable alternative. It is induced thrombocytopenia. The presence of thrombocytope-
clear from the literature that under these circumstances, with- nia in a neonate with a HPA-1a–negative mother or a history
drawal of heparin treatment without replacement anticoagulant of the disorder in a sibling is strong presumptive evidence in
therapy results in an unacceptably high rate of thrombotic events. favor of the diagnosis. Confirmation should include platelet
In the recent past, good alternatives for heparin were not avail- typing of both parents and testing for evidence of a maternal
able. Today, several alternative agents are suitable substitutes antibody directed at paternal platelets.37
(although considerably more expensive), including direct throm- In situations in which suspicion of NAIT is high or there is
bin inhibitors such as the intravenous use of argatroban and a history of NAIT in a first pregnancy, it may be necessary to
bivalirudin (Angiomax) (Chapter 43). Fondaparinux (Arixtra) is test or treat the fetus to prevent intracranial hemorrhage in
a synthetic heparin pentasaccharide, identical in chemical struc- utero. Fetal genotypes now can be determined at 10 to
ture to the antithrombin binding sequence in heparin and 18 weeks of gestation using polymerase chain reaction meth-
heparin-derived agents such as low-molecular-weight heparins. ods on cells obtained by chorionic villus sampling or amnio-
Fondaparinux is given subcutaneously, and while its use in pa- centesis.63 Periumbilical sampling to determine the fetal plate-
tients with HIT is “off-label” (not FDA approved), it is gaining let count can be performed at about 20 weeks of gestation.
favor in the clinical community as a second-line agent for When the fetus is thrombocytopenic, weekly maternal infu-
the management of suspected HIT to avoid progression into sions of IVIG have been shown to be effective in increasing the
acute HIT. fetal platelet count in most cases.64 In cases in which the fetal
platelet count does not increase with IVIG therapy, washed
Neonatal Alloimmune Thrombocytopenia.  Neonatal maternal platelets have been infused into the fetus with good
alloimmune thrombocytopenia (NAIT) develops when the results.65 Treatment of the mother with high-dose corticoste-
mother lacks a platelet-specific antigen (usually human platelet roids (to decrease maternal antibody production) is not rec-
antigen 1a, or HPA-1a [P1A1]) that the fetus has inherited from ommended because of potential fetal toxicity. In situations in
the father. HPA-1a is the most often involved (80% of cases), which the diagnosis of NAIT is known or highly suspected,
and HPA-5b accounts for another 10% to 15% of cases. Fetal delivery should be by cesarean section to avoid fetal trauma
platelet antigens may pass from the fetal to the maternal circula- associated with vaginal delivery. After delivery, the affected in-
tion as early as the fourteenth week of gestation.61 If the mother fant may be treated with transfusion of the appropriate
is exposed to a fetal antigen she lacks, she may make antibodies antigen-negative platelets (usually maternal). In addition, IVIG
to that fetal antigen. These antibodies cross the placenta, attach can be used alone or in combination with platelet transfusion.
to the antigen-bearing fetal platelets, and result in thrombocyto- IVIG should not be used as the sole treatment in a bleeding
penia in the fetus. In this regard, the pathophysiology of NAIT is infant, because response to this therapy usually takes 1 to
the same as that of hemolytic disease of the newborn. 3 days.62
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 727

Neonatal Autoimmune Thrombocytopenia.  The diag- never been transfused and in women who have never been
nosis of ITP or systemic lupus erythematosus in the mother is pregnant or never been transfused.68 PTP seems to require prior
a prerequisite for the diagnosis of neonatal autoimmune exposure to foreign platelet antigens and behaves in many re-
thrombocytopenia. Neonatal autoimmune thrombocytopenia spects like an anamnestic immune response.
is due to passive transplacental transfer of antibodies from a No clinical trials have been conducted on the treatment of
mother with ITP or, occasionally, systemic lupus erythemato- PTP, primarily because of the small number of cases. If PTP is
sus. The neonate does not have an ongoing autoimmune pro- untreated or treatment is ineffective, mortality rates may ap-
cess per se, but rather is an incidental target of the mother’s proach 10%.69 In addition, untreated or unresponsive patients
autoimmune process. During pregnancy, relapse is relatively have a protracted clinical course, with thrombocytopenia typi-
common for women with ITP in complete or partial remission; cally lasting 3 weeks but in some cases up to 4 months. Plas-
this has been attributed to the facilitation of reticuloendothe- mapheresis and exchange transfusion have been used with
lial phagocytosis by the high estrogen levels in pregnant some success in the past, but the treatment of choice is now
women. Women commonly develop chronic ITP during preg- IVIG. Many patients with PTP respond to a 2-day course of
nancy. ITP in the mother tends to remit after delivery. Cortico- IVIG, generally within the first 2 to 3 days, although a second
steroids are the primary treatment for pregnant women with course occasionally is necessary.69 IVIG also is much easier to
ITP, and at the dosages used, there is a relatively low incidence administer, and the response rates are higher than for plasma-
of adverse fetal side effects.66 Neonatal autoimmune thrombo- pheresis or exchange transfusion. Corticosteroid therapy is not
cytopenia develops in only about 10% of the infants of preg- particularly efficacious when used alone but may be beneficial
nant women with autoimmune thrombocytopenia, and intra- in combination with other, more effective treatments.37
cranial hemorrhage occurs in 1% or less. It is no longer
recommended that high-risk infants be delivered by cesarean Secondary Thrombocytopenia, Presumed to Be
section to avoid the trauma of vaginal delivery and accompany- Immune Mediated.  Severe thrombocytopenia has been ob-
ing risk of hemorrhage in the infant, regardless of maternal served in patients receiving biologic response modifiers such as
platelet count.67 interferons, colony-stimulating factors, and interleukin-2.70-72
Affected newborns may have normal to decreased platelet The thrombocytopenia associated with use of these substances
numbers at birth and have a progressive decrease in the platelet is reversible and, at least for interferon, may be immune medi-
count for about 1 week before the platelet count begins to in- ated, because increased levels of platelet-associated IgG have
crease. It has been speculated that the falling platelet count is been measured. Immune thrombocytopenia develops in about
associated with maturation of the infant’s reticuloendothelial 5% to 10% of patients with chronic lymphocytic leukemia and
system and accelerated removal of antibody-labeled platelets in a smaller percentage of patients with other lymphoprolifera-
by cells of the reticuloendothelial system. Neonatal thrombo- tive disorders.73,74 Thrombocytopenia also is noted in 14% to
cytopenia typically persists for about 1 to 2 weeks but some- 26% of patients with systemic lupus erythematosus.75 The
times lasts for several months. It usually does not require treat- clinical picture is similar to that of ITP: the bone marrow has a
ment. Severely thrombocytopenic infants generally respond larger than normal number of megakaryocytes, and increased
quickly to IVIG treatment. If an infant develops hemorrhagic levels of platelet-associated IgG frequently are found.36 Para-
symptoms, platelet transfusion, IVIG treatment, or corticoste- sitic infections also are known to cause thrombocytopenia.
roid therapy should be started immediately.37 Malaria is the most studied in this group and is regularly ac-
companied by thrombocytopenia, the onset of which corre-
Posttransfusion Purpura.  Posttransfusion purpura (PTP) sponds to the first appearance of antimalarial antibodies, a
is a relatively rare disorder that typically develops about 1 week decrease in serum complement, and control of parasitemia.
after transfusion of platelet-containing blood products, includ- There is evidence for the adsorption of microbial antigens to
ing fresh or frozen plasma, whole blood, and packed or washed the platelet surface and subsequent antibody binding via the
RBCs. PTP is manifested by the rapid onset of severe thrombo- Fab terminus.76 Immune destruction of platelets seems to be
cytopenia and moderate to severe hemorrhage that may be the most likely mechanism for the thrombocytopenia.
life-threatening. The recipient’s plasma is found to contain al-
loantibodies to antigens on the platelets or platelet membranes Nonimmune Mechanisms of Platelet Destruction
of the transfused blood product, directed against an antigen Nonimmune platelet destruction may result from exposure of
the recipient does not have. In more than 90% of cases, the platelets to nonendothelial surfaces, from activation of the co-
antibody is directed against the HPA-1a antigen; in most of the agulation process, or from platelet consumption by endovascu-
remaining cases, the antibodies are directed against PlA2 or lar injury without measurable depletion of coagulation factors.36
other epitopes on GP IIb/IIIa.37 Involvement of other alloanti-
gens, such as HPA-3a (Bak), HPA-4 (Penn), and HPA-5b (Br), Thrombocytopenia in Pregnancy and Preeclampsia
has been reported. The mechanism by which the recipient’s Incidental Thrombocytopenia of Pregnancy.  Incidental
own platelets are destroyed is unknown. Most patients with this thrombocytopenia of pregnancy also is known as pregnancy-
type of thrombocytopenia are multiparous middle-aged associated thrombocytopenia and gestational thrombocytopenia.
women. Almost all the other patients have a history of blood This disorder is the most common cause of thrombocytopenia in
transfusion. PTP seems to be exceedingly rare in men who have pregnancy. Random platelet counts in pregnant and postpartum
728 PART VI  Hemostasis and Thrombosis

women are slightly higher than normal, but about 5% of should be a part of the differential diagnosis of thrombocyto-
pregnant women develop a mild thrombocytopenia (100,000 penia in a pregnant woman. There is little or no correlation
to 150,000/mL), with 98% of such women having platelet between the level of maternal autoantibodies and the fetal
counts greater than 70,000/mL. These women are healthy platelet count. Other causes of thrombocytopenia during preg-
and have no prior history of thrombocytopenia. They do not nancy include HIV infection, systemic lupus erythematosus,
seem to be at increased risk for bleeding or for delivery antiphospholipid syndromes, TTP, and HUS. Of all women
of infants with neonatal thrombocytopenia. The cause of who develop TTP, 10% to 25% manifest the disease during
this type of thrombocytopenia is unknown. Maternal platelet pregnancy or in the postpartum period, and TTP tends to recur
counts return to normal within several weeks of delivery. in subsequent pregnancies.86,87 Plasmapheresis is the treatment
These women commonly experience recurrence in subse- of choice, and the maternal mortality is 90% or greater without
quent pregnancies. such treatment.
Preeclampsia and other hypertensive disorders of
pregnancy.  ​Approximately 20% of cases of thrombocytope- Hemolytic Disease of the Newborn.  Thrombocytope-
nia of pregnancy are associated with hypertensive disorders. nia, usually moderate in degree, occurs frequently in infants
These disorders include the classifications preeclampsia, pre- with hemolytic disease of the newborn. Although the RBC
eclampsia-eclampsia, preeclampsia with chronic hypertension, destruction characteristic of this disorder is antibody induced,
chronic hypertension, and gestational hypertension. Preeclamp- the antigens against which the antibodies are directed are not
sia complicates about 5% of all pregnancies and typically occurs expressed on platelets. Platelets may be destroyed as a result of
at about 20 weeks of gestation. The disorder is characterized by their interaction with products of RBC breakdown, rather than
the onset of hypertension and proteinuria and may include their direct participation in an immunologic reaction.36
abdominal pain, headache, blurred vision, or mental function
disturbances.77 Thrombocytopenia occurs in 15% to 20% of Thrombotic Thrombocytopenic Purpura.  TTP, some-
patients with preeclampsia, and about 40% to 50% of these times referred to as Moschcowitz syndrome, is characterized by
patients progress to eclampsia (hypertension, proteinuria, and the triad of microangiopathic hemolytic anemia, thrombocy-
seizures).78,79 topenia, and neurologic abnormalities.88 In addition, fever and
Some patients with preeclampsia have microangiopathic renal dysfunction (forming a pentad) are often present. Addi-
hemolysis, elevated liver enzymes, and a low platelet count, tional symptoms are present in most patients at the time of
termed HELLP syndrome. HELLP syndrome affects an estimated diagnosis and include diarrhea, anorexia, nausea, weakness,
4% to 12% of women with severe preeclampsia45,80,81 and and fatigue. TTP is uncommon but not rare, and its incidence
seems to be associated with higher rates of maternal and may be increasing. About twice as many women as men are
fetal complications. This disorder may be difficult to differenti- affected, and it is most common in women 30 to 40 years of
ate from thrombotic thrombocytopenic purpura (TTP), hemo- age.34,89 About half of the patients who develop TTP have a his-
lytic uremic syndrome (HUS), and disseminated intravascular tory of a viral-like illness several days before the onset of TTP.
coagulation (DIC). There seem to be at least four types of TTP. In most patients,
The development of thrombocytopenia in these patients TTP occurs as a single acute episode, although a small fraction
is thought to be due to increased platelet destruction. The of these patients may have recurrence at seemingly random
mechanism of platelet destruction is unclear, however. Some intervals. Recurrent TTP occurs in 11% to 28% of TTP pa-
evidence (elevated D-dimer) suggests that these patients have tients.90,91 A third type of TTP is drug induced. The primary
an underlying low-grade DIC.82 Elevated platelet-associated im- agents involved are the purinoreceptor (adenosine diphos-
munoglobulin is commonly found in these patients, however, phate) blocking agents ticlopidine (Ticlid) and clopidogrel
which indicates immune involvement.83 Early reports suggested (Plavix) used for inhibition of platelet function. Ticlopidine
that there may be a component of in vivo platelet activation seems to cause TTP in about 0.025% of patients, whereas the
because low-dose aspirin therapy has been shown to prevent incidence of clopidogrel-induced TTP is approximately four
preeclampsia in high-risk patients.84,85 When aspirin is used to times less frequent.89 The fourth type is chronic relapsing TTP,
prevent preeclampsia, however, reduction in risk is only 15%. a rare form of TTP, in which episodes occur at intervals of
The treatment of preeclampsia is delivery of the infant approximately 3 months starting in infancy.92,93
whenever possible. After delivery, the thrombocytopenia usu- Although it is unclear what triggers their deposition, hyaline
ally resolves in a few days. In cases in which delivery is not thrombi are found in the end arterioles and capillaries. These
possible (e.g., the infant would be too premature), bed rest and hyaline thrombi are composed of platelets and von Willebrand
aggressive treatment of the hypertension may help to increase factor (VWF) but contain very little fibrin or fibrinogen. As
the platelet count in some patients. Such treatments include these platelet-VWF thrombi are deposited, thrombocytopenia
magnesium sulfate and other antiepileptic therapies to inhibit develops. The degree of thrombocytopenia is directly related to
eclamptic seizures. the extent of microvascular platelet aggregation. RBCs flowing
Other causes of thrombocytopenia during pregnancy. ​ under arterial pressure are prone to fragmentation and he­
As has been discussed previously, ITP is a relatively common molysis when they encounter the strands of these thrombi.
disorder in women of childbearing age, and pregnancy does Hemolysis is usually quite severe, and most patients have less
nothing to ameliorate the symptoms of this disorder. ITP than 10 g/dL hemoglobin at the time of diagnosis. Examination
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 729

of the peripheral blood film reveals a marked decrease in plate- increased lactate dehydrogenase activity. Bone marrow examina-
lets, RBC polychromasia, and RBC fragmentation (microsphero- tion reveals erythroid hyperplasia and a normal to increased
cytes, schistocytes, keratocytes), a triad of features characteristic number of megakaryocytes. The partial thromboplastin time,
of microangiopathic hemolytic anemias (Figure 40-8). Nucle- prothrombin time, fibrinogen, fibrin degradation products, and
ated RBC precursors also may be present, depending on the de- D-dimer test results are usually normal and may be useful in
gree of hemolysis. Other laboratory evidence of intravascular differentiating this disorder from DIC.
hemolysis includes reduction of haptoglobin, hemoglobinuria, The thrombotic lesions also give rise to the other character-
hemosiderinuria, increased serum unconjugated bilirubin, and istic manifestations of TTP, because they are deposited in the
vasculature of all organs. The thrombi occlude blood flow and
lead to organ ischemia. Symptoms depend on the severity of
ischemia in each organ. Neurologic manifestations range from
headache to paresthesia and coma. Visual disturbances may be
of neurologic origin or may be due to thrombi in the choroid
capillaries of the retina or hemorrhage into the vitreous. Renal
dysfunction is common and present in more than half of
patients.90,91 Symptoms of renal dysfunction include protein-
uria and hematuria. Overwhelming renal damage with anuria
and fulminant uremia usually does not occur, however, which
helps to distinguish TTP from HUS.10 Gastrointestinal bleeding
occurs frequently in severely thrombocytopenic patients, and
abdominal pain is occasionally present due to occlusion of the
mesenteric microcirculation.
ADAMTS-13 and thrombotic thrombocytopenic purpura. 
​The development of TTP seems to be directly related to the ac-
A
cumulation of ultra-large von Willebrand factor (ULVWF) mul-
timers in the plasma of patients with TTP. VWF multimers are
made by megakaryocytes and endothelial cells. The primary
source of plasma VWF seems to be endothelial cells. Endothe-
lial cells secrete VWF into the subendothelium and plasma and
store it in Weibel-Palade bodies (storage granules). Endothelial
cells and megakaryocytes make even larger VWF multimers
(ULVWF multimers) than those found in plasma, and these are
even more effective than the normal plasma VWF multimers at
binding platelet GP Ib/IX or GP IIb/IIIa complexes under fluid
shear stresses (Figure 40-9).92 In the plasma, the ULVWF multi-
mers are rapidly cleaved into the smaller VWF multimers nor-
mally found in the plasma by a VWF-cleaving protease, called a
disintegrin and metalloprotease with a thrombospondin type 1
motif, member 13 (ADAMTS-13). This metalloprotease seems
B to be more effective when VWF multimers are partially un-
Figure 40-8  ​Microangiopathic hemolytic anemia. A,Thrombotic thrombocy- folded by high shear stress.94,95
topenic purpura (TTP); B, Hemolytic uremic syndrome (HUS). Abundant schis- Familial chronic relapsing TTP is a form characterized by
tocytes (arrows) reflect the platelet rich clots in the microvasculature that occur recurrent episodes of thrombocytopenia with or without is­
with TTP and HUS. TTP and HUS have similar blood morphologies. (From Carr chemic organ damage. In this type of TTP, the VWF-cleaving
JH, Rodak BF: Clinical hematology atlas, ed 4, Philadelphia, 2013, Saunders.) metalloprotease is completely deficient. The more common

+ADAMTS 13 Normal VWF

Normal VWF multimers
Spontaneous Unusually large VWF
multimer
synthesis multimers

–ADAMTS 13
Platelet agglutination
and aggregation

Figure 40-9  ​Mechanism for thrombotic thrombocytopenic purpura (TTP). Unusually large von Willebrand factor (ULVWF) multimers are normally digested
by the VWF–cleaving protease ADAMTS-13 (a disintegrin and metalloprotease with a thrombospondin type 1 motif, member 13). In TTP, the absence of
ADAMTS-13 allows the release of ULVWF, triggering platelet activation.
730 PART VI  Hemostasis and Thrombosis

form of TTP (usually not familial) does not tend to recur, but plasmapheresis, now 80% of patients who are treated early can
patients also are deficient in the metalloprotease. In this more be expected to survive. Because patients are known to experi-
common form of TTP, the metalloprotease deficiency occurs ence relapse, however, platelet counts should be monitored on
through removal of the enzyme (or blockade of its function) a regular basis until patients are in remission. The detection of
by a specific autoantibody that is present during TTP but ULVWF multimers in patient samples after complete remission
disappears during remission.96,97 An assay to measure the VWF- has predicted relapse accurately in 90% of the patients tested101;
cleaving enzyme has been introduced.98 However, the test this may prove to be useful in the long-term management of TTP.
may take several days, and treatment decisions must usually be
made before test results are available. In addition, the assay Hemolytic Uremic Syndrome.  Clinically, HUS resem-
lacks sensitivity and specificity for TTP. Additional tests for bles TTP except that it is found predominantly in children
ADAMTS-13 (and TTP) are in development and hold the 6 months to 4 years of age and is self-limiting. Approximately
promise for rapid diagnosis of TTP. 90% of cases are caused by Shigella dysenteriae serotypes or en-
In patients with TTP, ULVWF multimers tend to be present terohemorrhagic Escherichia coli OH serotypes, particularly
in the plasma at the beginning of the episode. These ULVWF O157:H7.102 These organisms sometimes can be cultured from
multimers, and usually the normal-sized plasma multimers, stool samples. The bloody diarrhea typical of childhood HUS
disappear as the TTP episode progresses and the thrombocyto- is caused by colonization of the large intestine with the offend-
penia worsens. Platelets and VWF are consumed during depo- ing organism, which causes erosive damage to the colon.
sition of the microvascular hyaline thrombi characteristic of S. dysenteriae produces Shiga toxin, and enterohemorrhagic
this disorder.99 If the patient survives an episode of TTP and E. coli produces either Shiga-like toxin-1 (SLT-1) or SLT-2,
does not experience relapse, the plasma VWF multimers are which can be detected in fecal samples from patients with
usually normal after recovery. If ULVWF multimers are found HUS. The toxins enter the bloodstream and attach to renal
in the plasma after recovery, however, it is likely that the glomerular capillary endothelial cells, which become damaged
patient will have recurrent episodes of TTP. The episodes may and swollen and release ULVWF multimers.102,103 This process
be infrequent and at irregular intervals (intermittent TTP) or leads to formation of hyaline thrombi in the renal vasculature
frequent and at regular intervals, as is often the case when TTP and the development of renal failure, thrombocytopenia, and
episodes occur in early childhood or infancy. microangiopathic hemolytic anemia, although the RBC frag-
The most effective treatment for TTP is plasma exchange mentation is usually not as severe as that seen in TTP (this is,
using fresh-frozen plasma or cryoprecipitate-poor plasma however, not a differentiating feature). The extent of renal in-
(plasma lacking most of the fibrinogen, fibronectin, and VWF).100 volvement correlates with the rate of recovery. In more severely
Either of these approaches may produce dramatic effects within affected children, renal dialysis may be needed. The mortality
a few hours. Because plasmapheresis is not available in all cen- rate associated with HUS in children is much lower than that
ters, the patient should be given corticosteroids and infusions for TTP, but there is often residual renal dysfunction that may
of fresh-frozen plasma immediately. Plasma exchange should lead to renal hypertension and severe renal failure. Because
be arranged as quickly as possible. Plasmapheresis and replace- HUS in children is essentially an infectious disorder, it affects
ment/infusion of plasma is effective on two fronts. First, some boys and girls equally and is often found in geographic clusters
of the ULVWF multimers will be removed by apheresis, and of cases rather than in random distribution.
plasma (fresh-frozen plasma or cryoprecipitate-poor plasma) The adult form of HUS is associated most often with expo-
supplies the deficient protease, which is able to degrade the sure to immunosuppressive agents or chemotherapeutic agents
ULVWF multimers in the blood of the patient. Because some or both, but it also may occur during the postpartum period.
patients with TTP have recovered while receiving immunosup- Usually the symptoms of HUS do not appear until weeks or
pressive treatment (corticosteroids) alone, it is recommended months after exposure to the offending agent.104 This disorder
that all patients with TTP be given high-dose corticosteroids in most likely results from direct renal arterial endothelial dam-
addition to undergoing plasma exchange. Plasma exchange age caused by the drug or one of its metabolic products. The
typically is continued over a 5-day period. If a response is not damage to endothelial cells results in release of VWF (includ-
seen within 5 days, or if the condition of the patient worsens ing ULVWF multimers), turbulent flow in the arterial system
during the first few days of plasma exchange, additional treat- with increased shear stresses on platelets, and VWF-mediated
ment is instituted. Such therapies include administering platelet aggregation in the renal arterial system. The renal im-
vincristine or azathioprine (or other immunosuppressive pairment in adults seems to be more severe than that in child-
agents), performing splenectomy, or passing the patient’s hood HUS, and dialysis is usually required. The cause of HUS
plasma over a staphylococcal A column to remove immune associated with pregnancy or oral contraceptive use is unclear,
complexes. The use of antiplatelet agents, prostacyclin, hepa- but it may be related to development of an autoantibody to
rin, or fibrinolytic agents is controversial and has not clearly endothelial cells. In outbreaks of HUS associated with con-
been shown to be helpful. Platelet transfusions should be sumption of E. coli–contaminated water, both children and
avoided unless intracranial bleeding or other serious hemor- adults have developed HUS.
rhagic problems arise.45 The cardinal signs of HUS are hemolytic anemia, renal fail-
Before 1990, TTP was fatal in more than 80% of patients. ure, and thrombocytopenia. The thrombocytopenia is usually
With the means for rapid diagnosis and the advent of exchange mild to moderate in severity. Renal failure is reflected in
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 731

elevated blood urea nitrogen and creatinine levels. The urine Abnormalities in Distribution or Dilution
nearly always contains RBCs, protein, and casts. The hemolytic An abnormal distribution of platelets also may cause thrombo-
process is shown by a hemoglobin level of less than 10 g/dL, cytopenia. The normal spleen sequesters approximately one
elevated reticulocyte count, and presence of schistocytes in the third of the total platelet mass. Mild thrombocytopenia may be
peripheral blood. present in any of the “big spleen” syndromes. The total body
Differentiating the adult form of HUS from TTP may be dif- platelet mass is often normal in these disorders, but numerous
ficult. The lack of neurologic symptoms, the presence of renal platelets are sequestered in the enlarged spleen, and conse-
dysfunction, and the absence of other organ involvement sug- quently the venous blood platelet count is low. Disorders such
gest HUS. Also, in HUS the thrombocytopenia tends to be as Gaucher disease, Hodgkin disease, sarcoidosis, lymphoma,
mild to moderate (platelet consumption occurs primarily in cirrhosis of the liver, and portal hypertension may result in
the kidneys), whereas in TTP the thrombocytopenia is usually splenomegaly and lead to thrombocytopenia.
severe. Similarly, fragmentation of RBCs and the resultant ane- Lowering the body temperature to less than 25° C, as is
mia tend to be milder than that observed in TTP, because RBCs routinely done in cardiovascular surgery, results in a transient
are being fragmented primarily in the kidneys. In some cases of but mild thrombocytopenia secondary to platelet sequestra-
HUS, other organs become involved, and the differentiation tion in the spleen and liver. An associated transient defect in
between HUS and TTP becomes less clear. In such cases, it is function also occurs with hypothermia. Platelet count and
prudent to treat as though the patient has TTP. function return to baseline values on return to normal body
temperature.32
Disseminated Intravascular Coagulation.  A common Thrombocytopenia often follows surgery involving extra-
cause of destructive thrombocytopenia is activation of the co- corporeal circulatory devices, as a consequence of damage and
agulation cascade (by a variety of agents or conditions), result- partial activation of platelets in the pump. In a few cases, severe
ing in a consumptive coagulopathy that entraps platelets in in- thrombocytopenia, marked impairment of platelet function,
travascular fibrin clots. This disorder is described in more detail and activation of fibrinolysis and intravascular coagulation
in Chapter 39 but is discussed here briefly for the sake of com- may develop.45
pleteness. DIC has many similarities to TTP, including microan- The administration of massive amounts of stored whole
giopathic hemolytic anemia and deposition of thrombi in the blood may produce a temporary thrombocytopenia. This phe-
arterial circulation of most organs. In DIC, however, the thrombi nomenon is explained by the fact that stored blood contains
are composed primarily of platelets and fibrinogen, whereas in platelets whose viability is severely impaired by the effects of
TTP the thrombi are composed primarily of platelets and VWF. storage and temperature. Under these conditions, the dead or
One form of DIC is acute with rapid platelet consumption damaged platelets are rapidly sequestered by the reticuloendo-
and results in severe thrombocytopenia. In addition, levels of thelial system of the patient. If this problem is encountered,
factor V, factor VIII, and fibrinogen are decreased as a result of it may be minimized by administering platelet concentrates
in vivo thrombin generation. The test for D-dimer (a break- or units of fresh whole blood along with the stored blood.
down product of stabilized fibrin) almost always yields posi- This situation is only rarely encountered, however, because
tive results. This form of DIC is life-threatening and must be the practice of transfusing whole blood has been replaced
treated immediately. almost completely by the use of specific components. Finally,
In chronic DIC, there is an ongoing, low-grade consumptive mild thrombocytopenia may be encountered in patients with
coagulopathy. Clotting factors may be slightly reduced or nor- chronic renal failure, severe iron deficiency, megaloblastic ane-
mal, and compensatory thrombocytopoiesis results in a mod- mia, postcompression sickness, and chronic hypoxia.
erately low to normal platelet count.34 D-dimer may not be
detectable or may be slightly to moderately increased. Chronic
THROMBOCYTOSIS: INCREASE
DIC is not generally life-threatening, and treatment usually is
IN CIRCULATING PLATELETS
not urgent. Chronic DIC is almost always due to some underly-
ing condition. If that condition can be corrected, the DIC usu- Thrombocytosis is defined as an abnormally high platelet
ally resolves without further treatment. Chronic DIC should count, typically more than 450,000/mL. The term reactive
be followed closely, however, because it can convert into the thrombocytosis is used to describe an elevation in the platelet
life-threatening acute form. count secondary to inflammation, trauma, or other underlying
and seemingly unrelated conditions. In reactive thrombocyto-
Drug-Induced Thrombocytopenia.  A few drugs directly sis, the platelet count is elevated for a limited period and usu-
interact with platelets to cause thrombocytopenia. Ristocetin, an ally does not exceed 800,000/mL, although platelet counts
antibiotic no longer in clinical use, facilitates the interaction of greater than 1 million/mL are occasionally seen (Figure 40-10).
VWF with platelet membrane GP Ib and leads to in vivo platelet A marked and persistent elevation in the platelet count is a
agglutination and thrombocytopenia. Hematin, used for the hallmark of myeloproliferative disorders such as polycythemia
treatment of acute intermittent porphyria, may give rise to a tran- vera, chronic myelogenous leukemia, and myelofibrosis with
sient thrombocytopenia that seems to be caused by stimulation myeloid metaplasia (or primary myelofibrosis). In these condi-
of platelet secretion and aggregation. Protamine sulfate and bleo- tions, the platelet count often exceeds 1 million/mL. Although
mycin may induce thrombocytopenia by a similar mechanism.45 the terms thrombocythemia and thrombocytosis are often used
732 PART VI  Hemostasis and Thrombosis

BOX 40-3  ​Processes Resulting in Thrombocytosis

Conditions Associated with Reactive

Thrombocytosis
Blood loss and surgery
Splenectomy
Iron deficiency anemia
Inflammation and disease
Stress or exercise

Myeloproliferative Disorders Associated

with Thrombocytosis
Polycythemia vera
Chronic myelogenous leukemia
Myelofibrosis with myeloid metaplasia
Thrombocythemia: essential or primary
Figure 40-10  ​Peripheral blood film showing cell morphology in reactive
thrombocytosis. Note the increased number of platelets but reasonably normal From Colvin BT: Thrombocytopenia, Clin Haematol 14:661-681, 1985; and
Thompson AR, Harker LA: Manual of hemostasis and thrombosis, ed 3, Philadelphia,
platelet morphology, characteristic of reactive thrombocytosis.
1983, FA Davis.

interchangeably, in this text the term thrombocythemia is used platelet production remains responsive to normal regulatory
only as part of the description of the myeloproliferative disorder stimuli (e.g., thrombopoietin, a humoral factor that is pro-
known as essential thrombocythemia (Figure 40-11). In essential duced in the kidney parenchyma), and morphologically nor-
thrombocythemia, platelet counts typically exceed 1 million/mL mal platelets are produced at a moderately increased rate. This
and may reach levels of several million.34,105,106 Processes result- is in contrast to essential thrombocythemia, which is character-
ing in thrombocytosis are summarized in Box 40-3. ized by unregulated or autonomous platelet production and
platelets of variable size.105,106
Reactive (Secondary) Thrombocytosis Examination of the bone marrow from patients with reac-
Platelet counts between 450,000/mL and 800,000/mL with no tive thrombocytosis reveals a normal to increased number of
change in platelet function can result from acute blood loss, megakaryocytes that are normal in morphology. Results of
splenectomy, childbirth, and tissue necrosis secondary to sur- platelet function tests, including aggregation induced by
gery, chronic inflammatory disease, infection, exercise, iron various agents, and bleeding time are usually normal in reac-
deficiency anemia, hemolytic anemia, renal disorders, and tive thrombocytosis but also may be normal in patients with
malignancy. Occasionally, patients manifest a platelet count of elevated platelet counts accompanying myeloproliferative
1 to 2 million/mL (Figure 40-10). In reactive thrombocytosis, disorders.
Reactive thrombocytosis is not associated with thrombosis,
hemorrhage, or abnormal thrombopoietin levels. It seldom
produces symptoms per se and disappears when the underly-
ing disorder is brought under control.105,106

Reactive Thrombocytosis Associated with

Hemorrhage or Surgery
After acute hemorrhage, the platelet count may be low for 2 to
6 days (unless platelets have been transfused) but typically re-
bounds to elevated levels for several days before returning to
the prehemorrhage level. A similar pattern of thrombocytope-
nia and thrombocytosis is seen after major surgical procedures
in which there is significant blood loss. In both cases, the plate-
let count typically returns to normal 10 to 16 days after the
episode of blood loss.

Figure 40-11  ​Peripheral blood film showing cell morphology in essential Postsplenectomy Thrombocytosis
thrombocythemia. Note the increased number of platelets and wide variation Removal of the spleen typically results in platelet counts that
in platelet size characteristic of essential thrombocythemia. Red and white can reach or exceed 1 million/mL regardless of the reason for
blood cell morphology is characteristically normal. (From Carr JH, Rodak BF: splenectomy. The spleen normally sequesters about one third
Clinical hematology atlas, ed 4, Philadelphia, 2013, Saunders.) of the circulating platelet mass. After splenectomy, one would
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 733

expect an initial increase in the platelet count of approximately left shift, and many patients develop a mild normochromic,
30% to 50%. The platelet count, however, far exceeds levels normocytic anemia. During this phase, cardiovascular compli-
that could result from rebalancing of the circulating platelet cations and aneurysms develop. The higher the platelet count,
pool to incorporate the splenic platelet pool. The cause of the the higher the risk of cardiovascular complication. After the
accelerated platelet production is unknown. Unlike after blood subacute phase comes the convalescent phase, during which all
loss from hemorrhage or other types of surgery, the platelet signs of illness disappear and the acute phase reactants subside
count reaches a maximum 1 to 3 weeks after splenectomy and to normal. The highest incidence of Kawasaki disease is found
remains elevated for 1 to 3 months. In some patients who un- in Japan and in individuals of Japanese descent, although the
dergo splenectomy for treatment of chronic anemia, the count disease seems to occur in most, if not all, ethnic groups. There
can remain elevated for several years. is no specific test for Kawasaki disease. Diagnosis is primarily by
excluding other diseases that cause similar signs and symptoms
Thrombocytosis Associated with Iron (e.g., scarlet fever, juvenile rheumatoid arthritis, Stevens-Johnson
Deficiency Anemia syndrome, and toxic shock syndrome). The treatment for
Mild iron deficiency anemia secondary to chronic blood loss is Kawasaki disease is administration of antiplatelet agents and
associated with thrombocytosis in about 50% of cases. Throm- immunoglobulin.
bocytosis can be seen in severe iron deficiency anemia, but An elevated platelet count also may be early evidence of a
thrombocytopenia also has been reported. In some cases of tumor (e.g., Hodgkin disease) and various carcinomas. Finally,
iron deficiency, the platelet count may be 2 million/mL. After hemophilic patients often have platelet counts above normal
iron therapy is started, the platelet count usually returns to limits, even in the absence of active bleeding.
normal within 7 to 10 days. It is believed that iron plays some
role in regulating thrombopoiesis, because treatment of the Exercise-Induced Thrombocytosis
iron deficiency with iron replacement has resulted in a normal- Strenuous exercise is a well-known cause of relative thrombo-
ization of the platelet count in thrombocytopenic patients and cytosis and is likely due to the release of platelets from the
has been reported to induce thrombocytopenia in patients splenic pool or hemoconcentration by transfer of plasma water
with normal platelet counts. Not enough research has been to the extravascular compartment or both. Normally, the plate-
done, however, to elucidate the role of iron in thrombopoiesis. let count returns to its preexercise baseline level 30 minutes
after completion of exercise.
Thrombocytosis Associated with Inflammation
and Disease Rebound Thrombocytosis
Similar to elevations in C-reactive protein, fibrinogen, VWF, Thrombocytosis often follows the thrombocytopenia caused
and other acute phase reactants, thrombocytosis may be an by marrow-suppressive therapy or other conditions. “Rebound”
indication of inflammation. Thrombocytosis may be found in thrombocytosis usually reaches a peak 10 to 17 days after with-
association with rheumatoid arthritis, rheumatic fever, osteo- drawal of the offending drug (e.g., alcohol or methotrexate)
myelitis, ulcerative colitis, acute infections, and malignancy. In or after institution of therapy for the underlying condition
rheumatoid arthritis, the presence of thrombocytosis can be with which thrombocytopenia is associated (e.g., vitamin B12
correlated with activation of the inflammatory process. deficiency).45
Kawasaki disease (Kawasaki syndrome) causes inflammation
of the walls of small and medium-sized arteries throughout the Thrombocytosis Associated with
body. It is also known as mucocutaneous lymph node syn- Myeloproliferative Disorders
drome because it affects lymph nodes, skin, and mucous mem- Primary or autonomous thrombocytosis is a typical finding in
branes in the mouth, nose, and throat. It is an acute febrile four chronic myeloproliferative disorders: polycythemia vera,
illness of infants and young children. Boys are more likely than chronic myelogenous leukemia, myelofibrosis with myeloid
girls to develop the disease, and children of Japanese and metaplasia (primary myelofibrosis), and essential thrombocy-
Korean descent have higher rates of Kawasaki disease. It is a self- themia. Depending on the duration and stage of the myelopro-
limited acute vasculitic syndrome of unknown origin, although liferative disorder at the time of diagnosis, it may be difficult to
an infectious etiology has been suspected. Although the disease differentiate among these diseases. Chapter 33 provides a more
is self-limiting, there can be lifelong sequelae, including coro- complete description of these disorders. In the other types of
nary artery thrombosis and aneurysms. The acute febrile stage myeloproliferative disorders, the platelet count seldom reaches
of the disease lasts 2 weeks or longer, with a fever of 40° C or the extreme values characteristic of essential thrombocythe-
higher, and is unresponsive to antibiotic therapy. The longer the mia. Diagnosis of essential thrombocythemia should not be
fever continues, the higher the risk of cardiovascular complica- based on the platelet count alone but should also take into
tions. The subacute phase lasts an additional week to 10 days. account the physical examination findings, history, and other
During this phase, the platelet count usually is elevated, and laboratory data.34,106
counts of 2 million/mL have been reported. In addition, acute
phase reactants such as C-reactive protein and sedimentation Essential (Primary) Thrombocythemia
rate are elevated, consistent with an inflammatory component. Essential thrombocythemia is a clonal disorder related to other
The WBC count can be moderately to markedly elevated with a chronic myeloproliferative diseases and is the most common
734 PART VI  Hemostasis and Thrombosis

cause of primary thrombocytosis. It is characterized by periph- A patient with essential thrombocythemia who has had a
eral blood platelet counts exceeding 1 million/mL and uncon- thrombotic event may have a hemorrhagic event later.109
trolled proliferation of marrow megakaryocytes. Although the The bleeding manifestations may be related to a variety of
platelet count may (or may not) be markedly elevated in other qualitative abnormalities in the platelets, including deficien-
myeloproliferative disorders, persistent marked elevation of the cies in epinephrine receptors and ultrastructural defects in
platelet count is an absolute requirement for the diagnosis of granules, mitochondria, and microfilaments. Platelets may be
essential thrombocythemia (Figure 40-11). There is evidence agranular or hypogranular and have a clear, light blue appear-
that essential thrombocythemia is caused by a clonal prolifera- ance on a routine Wright-stained film of the peripheral blood.
tion of a single abnormal pluripotential stem cell that eventu- Although platelet size is heterogeneous, giant and bizarrely
ally crowds out normal stem cells. As with most myeloprolifera- shaped platelets are characteristic of myeloproliferative dis-
tive disorders, essential thrombocythemia is neither congenital eases, and megakaryocyte fragments or nuclei are commonly
nor hereditary, is prevalent in middle-aged and older patients, encountered in the peripheral blood. Platelets may be notably
and affects equal numbers of men and women. In contrast to clumped on blood films, exhibiting marked variation in size
other myeloproliferative disorders, however, the other marrow and shape. The number and volume of megakaryocytes are
cell lines are not involved at the time of diagnosis. increased in the bone marrow, and they are predominantly
The clinical manifestations of essential thrombocythemia large, show some cellular atypia, and tend to form clusters.
are hemorrhage, platelet dysfunction, and thrombosis. Bleed- Often the platelets are functionally defective when tested in
ing times are usually normal. There is no specific clinical sign, vitro. Aggregation is usually absent in response to epinephrine
symptom, or laboratory test that establishes the diagnosis of and may be decreased with adenosine diphosphate but is usu-
essential thrombocythemia. The diagnosis must be made by ally normal with collagen. Lack of an epinephrine response
ruling out the other myeloproliferative disorders and systemic may help to differentiate essential thrombocythemia from re-
illnesses that produce reactive thrombocytosis. active thrombocytosis, because this response is usually normal
Thrombosis in the microvasculature is relatively common in in reactive thrombocytosis but absent in most cases of essential
essential thrombocythemia, and the incidence at the time of thrombocythemia. Platelet adhesion also may be decreased.106
diagnosis is 10% to 20%. This thrombosis can lead to digital The degree of thrombocytosis has not been found to predict
pain, digital gangrene, or erythromelalgia (throbbing, aching, hemorrhagic or thrombotic events reliably. The role of lowering
and burning sensation in the extremities, particularly in the platelet counts as a prophylactic treatment in this disease is not
palms and soles).106 The symptoms of erythromelalgia can be established. The risks from exposure to mutagenic alkylating
explained by arteriolar inflammation and occlusive thrombosis agents used to decrease the platelet count may be greater than the
mediated by platelets and can be relieved for several days by a risk of thrombosis or hemorrhage. When treatment is deemed
single dose of aspirin.107 Thrombosis of large veins and arteries necessary because of thrombotic tendencies or splenomegaly, a
also may occur in essential thrombocythemia. The arteries most variety of myelosuppressive agents (e.g., melphalan, busulfan,
commonly involved are those in the legs, the coronary arteries, hydroxyurea, or even radioactive phosphorus) can be used.106 In
and the renal arteries, but involvement of the mesenteric, subcla- patients with life-threatening hemorrhage or thrombosis and an
vian, and carotid arteries is not infrequent (in fact, neurologic extremely high platelet count, platelet apheresis may be used to
complications are relatively common). Venous thrombosis may reduce the platelet count rapidly. In these situations, a myelosup-
involve the large veins of the legs and pelvis, hepatic veins, or pressive agent is added for longer-term control of the platelet
splenic veins.108 The platelets of some patients who have experi- count.110 Interferon-a has been used to treat essential thrombocy-
enced thrombotic episodes have been shown to have increased themia and is associated with an approximately 60% rate of
binding affinity for fibrinogen and to generate more than the complete remission, but 28% of patients given the drug cannot
usual quantities of thromboxane B2, and these patients have tolerate the dosages required.111-113 A newer agent that has shown
elevated levels of thromboxane B2 and b-thromboglobulin in the great promise in the treatment of essential thrombocythemia is
blood. These findings suggest enhanced in vivo platelet activation anagrelide. The drug acts by inhibiting megakaryocyte matura-
and perhaps an explanation for the thrombotic tendencies of tion and platelet release.114 In one large study, anagrelide de-
patients with essential thrombocythemia. The primary cause of creased the platelet count in 93% of patients.115 Many patients
death of patients with essential thrombocythemia seems to be cannot tolerate anagrelide, however, and in these patients other,
thrombosis. Hemorrhagic episodes occur less frequently than more traditional chemotherapeutic agents seem to be more effec-
thrombotic episodes in patients with essential thrombocythemia. tive. Despite the availability of newer treatments for essential
As with bleeding secondary to platelet function disorders, thrombocythemia, whether a patient with essential thrombocy-
the hemorrhagic manifestations of essential thrombocythemia themia and an elevated platelet count who is asymptomatic
are mucocutaneous in nature, with gastrointestinal tract bleed- should or should not be treated remains controversial.
ing occurring most frequently. Other sites of bleeding include In patients with essential thrombocythemia there is a
the mucous membranes of the nose and mouth, the urinary low incidence of transformation to acute leukemia or fatal
tract, and the skin. Symptoms may be aggravated by aspirin use. thrombotic or hemorrhagic complications. Therapy to prevent
In an occasional patient with essential thrombocythemia, there thrombotic complications seems to be effective in preventing
is a paradoxical combination of thromboembolic (clotting) morbidity but does not seem to improve survival, at least in
and hemorrhagic episodes in association with this condition. high-risk patients.
 CHAPTER 40  Thrombocytopenia and Thrombocytosis 735

SUMMARY

• Thrombocytopenia is the most common cause of clinically signifi- • Treatment of drug-induced thrombocytopenia must begin with
cant bleeding. identification of the causative drug and discontinuation of its use.
• Thrombocytopenia may be a result of decreased platelet produc- • TTP presents with a triad of symptoms that includes microan-
tion, increased destruction, or abnormal distribution of platelets giopathic hemolytic anemia, thrombocytopenia, and neurologic
and manifests with small-vessel bleeding in the skin. abnormalities; these may be accompanied by fever and renal
• Decreased production of platelets can be attributed to mega- dysfunction.
karyocyte hypoplasia, ineffective thrombopoiesis, or replacement • Abnormal distribution of platelets can be caused by splenic
of marrow by abnormal cells. sequestration.
• Patients experiencing increased platelet destruction become • Reactive thrombocytosis is secondary to inflammation, trauma, or
thrombocytopenic only when the rate of platelet production can no a variety of underlying conditions. Platelet counts are increased for
longer increase enough to compensate. a limited time. The thrombocytosis seen in myeloproliferative dis-
• Pathologic destruction of platelets can be caused by both immu- orders is marked and persistent.
nologic and nonimmunologic mechanisms.
• Acute ITP commonly occurs in children after a viral illness, and Now that you have completed this chapter, go back and
there is usually spontaneous remission. Chronic ITP is more com- read again the case study at the beginning and respond
monly seen in women and requires treatment if the platelet count to the questions presented.
decreases to fewer than 30,000/mL.

R E V I E W Q UESTIONS

Answers can be found in the Appendix. 5. What is the first step in the treatment of HIT?
a. Start low-molecular-weight heparin therapy
1. The autosomal dominant disorder associated with decreased b. Stop heparin infusion immediately
platelet production is: c. Switch to warfarin (Coumadin) immediately
a. Fanconi anemia d. Initiate a platelet transfusion
b. TAR syndrome
c. May-Hegglin anomaly 6. A defect in primary hemostasis (platelet response to an
d. Wiskott-Aldrich anomaly injury) often results in:
a. Musculoskeletal bleeding
2. Which of the following is not a hallmark of ITP? b. Mucosal bleeding
a. Petechiae c. Hemarthroses
b. Thrombocytopenia d. None of the above
c. Large overactive platelets
d. Megakaryocyte hypoplasia 7. When a drug acts as a hapten to induce thrombocytopenia,
an antibody forms against which of the following?
3. The specific antigen most commonly responsible for the a. Typically unexposed, new platelet antigens
development of NAIT is: b. The combination of the drug and the platelet membrane
a. Bak protein to which it is bound
b. HPA-1a c. The drug alone in the plasma, but the immune complex
c. GP Ib then binds to the platelet membrane
d. Lewis antigen a d. The drug alone, but only when it is bound to the platelet
membrane
4. A 2-year-old child with an unexpected platelet count of
15,000/mL and a recent history of a viral infection most 8. TAR refers to:
likely has: a. Abnormal platelet morphology in which the radial stria-
a. HIT tions of the platelets are missing
b. NAIT b. Abnormal appearance of the iris of the eye in which
c. Acute ITP radial striations are absent
d. Chronic ITP c. Abnormal bone formation, including hypoplasia of the
forearms
d. Neurologic defects affecting the root (radix) of the spinal
nerves