1.0 INTRODUCTION Physiological changes in pregnancy affect the coagulation and fibrinolytic systems. Many of the clotting factors increase and… [601031]

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CHAPTER ONE
1.0 INTRODUCTION
Physiological changes in pregnancy affect the coagulation and fibrinolytic
systems. Many of the clotting factors increase and anticoagulation factors
decrease causing augmented coagulation and decreased fibrinolysis. Pre –
existing coagulopathies may affect the course of pregnancy and nature of
coagulopathy may also be modified by pregnancy (verdy et al., 1997) Changes
in coagulation affect the mode of delivery and the approach to management of
haematological cases in pregnancy. Pregnanc y involves the formation and the
development of an offspring, known as an embryo (foetus ), in the uterus of a
woma n in human reproduction. Most often , human pregnancy is medically
divided into three tri meste rs. A trimester corresponds to a three month period in
pregna ncy. These pregnancy periods serve as a me ans to simplify different
developmental stages of prenatal development. Implantation complications
such as miscarriage s are common mostly in the first trimester . Dur ing the
second trimester, the growth and devel opment of the foetus can b e easily
monitored and abnormalities can be diagnosed and corrected where possible .
Further growth and development of the foe tus which includes foetal fat stores
build up occurs during the third trimester (ACOG, 2002).
Pregnancy is a s tate characterized by many physiological haematological
changes, which may appear to be pathological in the non -pregnant state. Normal
pregnancy is characterized by profound changes in almost every organ and

2
system to accommodate the demands of faetoplacen tal unit (Harrison,
1966). The bleeding time remains normal throughout pregnancy. Screening tests
used for the investigation of bleeding; the Partial thromboplastin time with
Kaolin and the prothrombin time (PT) are within normal adult ranges during
pregnan cy, but in the third trimester the PT and the APTT are at the lower
(shorter) limits of normal or slightly shortened, and this must be taken into
account when assessing coagulation screen results from pregnant women
(Walker et al ., 1994). However, other re searchers have reported changes in
some coagulation parameters in normal pregnancy. As reported by Hellgren in
2003, prothrombin complex was increased in the pregnant women studied,
which was expressed as INR of less than 0.9. Similarly, Uchikova et al. (2005),
reported PT as being significantly shortened in pregnancy compared with the
study control. Significant difference has been reported in PTTK among
pregnant and control subjects, which shows that the level of factors in the
intrinsic pathway are also in creased in normal pregnancy (Durotoye et al .,
2012). They also reported insignificant differences in the result of PTTK in
various trimesters. Blood cells morphorlogy and population has been reported t o
undergo changes in pregnancy (Edlestam et al., 2001 ; Firki et al., 2002;
Hoffbrond et al. 2003).

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Large cross -sectional studies done in pregnancy of healthy women (specifically
excluding any with hypertension) have shown that the platelet count does
decrease during pregnancy, particularly in the third trimest er. This is termed as
‗‗gestational thrombocytopenia‘‘ It is partly due to haemodilution and partly
due to increased platelet activation and accelerated clearance (Shehlata et al .,
1999). The reference range (99% of populati on analyzed) for platelets was
150,000 to 400,000 per cubic millimeter (Ross et al ., 1988) . The reference
range has been confirmed to be the same in the elderly. However, it has been
observed that Men usually have slightly higher mean values when compared to
women (Ross et al., 1988). In this study, the effect of pregnancy on Blood cells
morphorlogy, platel et count, Prothrombin time (PT), International Normalised
Ratio (INR) and Partial thromboplastin time with kaolin (PTTK) will b e
studied.
1.1 Justification of Study
It has been shown that Thrombocytopenia is the second most common
haematological finding in pregnancy after anaemia. It affects 7 -10% of all
pregnant women (Verdy et al., 1997) and that blood clot di sorders are becoming
increasingly frequent in pregnancy.Screeni ng tests used for the investigation of
bleeding; the Partial thromboplastin time with Kaolin and the prothrombin time
(PT) are within normal adult ranges during pregnancy, but in the third trimester
the PT and the APTT are at the lower (shorter) limits of normal or slightly
shortened, and this must be taken into account when assessing coagulation

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screen results from pregnant women (Walker et al ., 1994). However, It is
there fore important to monitor changes in platelet count and haemostatic status
in pregnan t women attending antenatal clinic. This w ill aid the health care
providers to take appropriate steps in ensuring good health in pregnancy.
1.2 Aim of Study
The aim o f this study is to study blood clot and platelet disorder s in pregnant
women attending an tenatal clinic at General Hospital, Esan West Local
Government Area.
1.3 Specific Objectives
To carry out platelet count on the subjects
To determine PT and INR of the subjects
To evaluate PTTK in the subjects
To examine the blood films of the subjects
To determine the differences in the stu died parameters in the subjects
1.4 Research Design
This research wa s designed to study blood clot and platelet disorders in
pregnant women attending antenata l clinic at Ekpoma, Esan west local
government of Edo State. Random samp ling method will be used for this cohort
study.
1.5 Ethical Approval
Permission will be sought from Ethical committee of Edo state hospital
management Board , Benin City; for approval to collect blood samples from

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pregnant women attending a nte-natal clinic at General Hospital Ekpoma, Edo
State .

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CHAPTER TWO
2.0 LITERATURE REVIEW
2.1 PREGNANCY
Pregnancy also known as gestation is the formation and the development of an
offspring , known as an embryo or foetus, in the ute rus of a woman. In hu mans,
pregnancies usually occur as a single foetus however, in most cases multiple
pregnancies occur and they usually involve more than one embryo in a single
pregnancy, as with twins. Childbirth usually occurs about 38 weeks (9 months)
after conception. Women who experience menstrual cycle s of about four weeks
(a month) will likely have an extended pregnancy up to approximate ly 40 weeks
starting from the last normal menstrual period (LNMP) . From day one of
conception to week 8 the co ncept is termed an embryo , beyond this age till
parturition it is termed a foetus (ACOG, 2002).
2.2 STAGES OF PR EGNANCY
Most often , human pregnancy is medically divided into three tri meste rs. A
trimester corresponds to a three month period . These pregnancy periods serve as
a me ans to simplify different developmental stages of prenatal development.
Implantation complications such as miscarriage s are common mostly in the first
trimester . Dur ing the second trimester, the growth and devel opment of the
foetus can b e easily monitored and abnormalities can be diagnosed and
corrected where possible . Further growth and development of the foe tus which

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includes foetal fat stores build up occurs during the third trimester (ACOG,
2002).
2.2.1 FIRST TRIMESTER
During the first trimester the pregnant women will experience many
physiological changes that accompanies pregnancy such as increased minute
ventilation by about 40%, increased size and growth of the uterus to the size of
a lemon at about the 8th week of gestation. Many other sig ns of discomfort such
as nausea , vomiting (early morni ng sickness usually occur to mos t women
(Campbell and Klocke, 2001).
2.2.2 SECOND TRIMESTER
Second trimeste r usually fall between the 13th to 28th week of gest ation.
Physiological changes that accompany this stage include increased weight, fat
redistribution to the breast, energetic, morning sickness which soon fades away
as pregnancy progresses. During the second trimester, the growth and
devel opment of the foetus can b e easily monitored and abnormalities can be
diagnosed and corrected where possible . At this stage the ut erus can expand up
to tw enty (20) times its normal size (Campbell and Klocke, 2001).
2.2.3 THIRD TRIMESTER
This stage corresponds to gestational age of week 28 a nd above, where it is
usually accompanied with physiological symptoms such as enormous weight
gain, a bell -shaped abdomen which usually drops downward as the baby attains

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a downward position getting ready for parturition (Campbell and Klocke, 2001).
At this stage the women usually feels the baby‘s movements. As the foetal head
descends into cephalic presentation, it relieves pressure from the upper
abdomen wi th renewed ease in breathing and may also severely reduce bladder
capacity, and increases pressure on the pelvic floor and the rectum (Stacey et
al., 2011).

Figure 1.1: A diagrammatic representation of a pregnant woman showing the
extension of the uterus into the ab dominal region during pregnancy
(Stacey et al., 2011).

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2.3 HAEMATOLOGICAL CHANGES IN PREGNANCY
Pregnancy is a state characterized by many physiological haematological
changes, which may appear to be pathological in the non -pregnant state. N ormal
pregnancy is characterized by profound changes in almost every organ and
system to accommodate the demands of faetoplacental unit (Harrison, 1966). In
normal pregnancy, the physiological change in haemoglobin concentration
[HGB] and platelet count du ring pregnancy are well known phenomena (Yip,
2000). It is also one of the physiological conditions capable of causing
remarkable and dramatic changes in haematological variables. A pregnancy is
influenced by many factors, some of which include culture, en vironment,
socioeconomic status, and access to medical care. The haematological indices
also have an impact on pregnancy and its outcome (Yip, 2000). The
haematological indices of an individual to a large extent reflect their general
health (WHO, 2004). B lood is a special type of connective tissue composed of
formed elements in a fluid matrix. Many of the haematological indices are
influenced by many factors like sex, seasonal variation, lactation, pregnancy
health, and nutritional status (Smith, 1993).
2.3.1 Red Blood Cells Morphology
During pregnancy, the total blood volume increases by about 1.5 litres, mainly
to supply the demands of the new vascular bed and to compensate for blood loss
occurring at delivery (Ramsay, 2010). Of this, around one litre of blood is
contained within the uterus and maternal blood spaces of the placenta. Increase

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in blood volume is, therefore, more marked in multiple pregnancies and in iron
deficient states. Expansion of plasma volume occurs by 10 –15 % at 6 –12 weeks
of gestati on (Bernstein et al ., 2001; Bjorksten et al ., 1978). Red cell mass
(driven by an increase in maternal erythropoietin production) also increases, but
relatively less, compared with the increase in plasma volume, the net result
being a dip in haemoglobin con centration. Thus, there is Dilutional anaemia.
The drop in haemoglobin is typically by 1 –2 g/dL by the late second trimester
and stabilizes thereafter in the third trimester, when there is a reduction in
maternal plasma volume (owing to an increase in leve ls of atrial natriuretic
peptide). Women who take iron supplements have less pronounced changes in
haemoglobin, as they increase their red cell mass in a more proportionate
manner than those not on haematinic supplements. The red blood cell indices
change little in pregnancy. However, there is a small increase in mean
corpuscular volume (MCV), of an average of 4 fl in an iron -replete woman,
which reaches a maximum at 30 –35 weeks gestation and does not suggest any
deficiency of vitamins B12 and folate. Incre ased production of RBCs to meet
the demands of pregnancy, reasonably explains why there is an increased MCV
(due to a higher proportion of young RBCs which are larger in size). However,
MCV does not change significantly during pregnancy and a haemoglobin
concentration <9.5 g/dL in association with a mean corpuscular volume <84 fl
probably indicates co -existent iron deficiency or some other pathology (Crocker
et al., 2000). Post pregnancy, plasma volume decreases as a result of diuresis,

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and the blood volume returns to non -pregnant values. Haemoglobin and
hematocrit increase consequently. Plasma volume increases again two to five
days later, possibly because of a rise in aldosterone secretion. Later, it again
decreases. Significant elevation has been document ed between measurements of
haemoglobin taken at 6 –8 weeks postpartum and those taken at 4 –6 months
postpartum, indicating that it takes at least 4 –6 months post pregnancy, to
restore the physiological dip in haemoglobin to the non -pregnant values (Taylor
and Lind, 1979).
2.3.2 White Blood Cells
White blood cell count is increased in pregnancy with the lower limit of the
reference range being typically 6,000/mm3. Leucocytosis, occurring during
pregnancy is due to the physiologic stress induced by the pregna nt state
(Fleming, 1975).
Neutrophils are the major type of leucocytes on differential counts
(Konijnenberg et al ., 1997). This is likely due to impaired Neutrophilic
apoptosis in pregnancy (Gatti et al ., 1994). The neutrophils cytoplasm shows
toxic granul ation. Neutrophils chemotaxis and phagocytic activity are
depressed, especially due to inhibitory factors present in the serum of a pregnant
female (Jessica et al ., 2007). There is also evidence of increased oxidative
metabolism in neutrophils during pregn ancy. Immature forms as myelocytes
and metamyelocytes may be found in the peripheral blood film of healthy
women during pregnancy and do not have any pathological significance

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(Karalis et al., 2005). They simply indicate adequate bone marrow response to
an increased drive for erythropoiesis occurring during pregnancy. Lymphocyte
count decreases during pregnancy through the first and second trimesters and
increases during the third trimester. There is an absolute monocytosis during
pregnancy, especially in t he first trimester, but decreases as gestation advances.
Monocytes help in preventing faetal allograft rejection by infiltrating the
decidual tissue (7th –20th week of gestation) possibly, through PGE2 mediated
immunosuppression (Kline et al., 2005). The mo nocytes to lymphocyte ratio are
markedly increased in pregnancy. Eosinophils and basophil counts, however, do
not change significantly during pregnancy (Edlestam et al., 2001). The stress of
delivery may itself lead to brisk Leucocytosis. Few hours after d elivery, healthy
women have been documented as having a WBC count varying from 9,000 to
25,000/mm3. By 4 weeks post -delivery, typical WBC ranges are similar to those
in healthy non -pregnant women.
2.3.3 Platelets
Large cross -sectional studies done in preg nancy of healthy women (specifically
excluding any with hypertension) have shown that the platelet count does
decrease during pregnancy, particularly in the third trimester. This is termed as
‗‗gestational thrombocytopenia‘‘ It is partly due to haemodiluti on and partly
due to increased platelet activation and accelerated clearance (Shehlata et al .,
1999). Gestational thrombocytopenia does not have complications related to
thrombocytopenia and babies do not have severe thrombocytopenia (platelet

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count ≤20,00 0/mm3). The platelet volume distribution width increases
significantly and continuously as gestation advances, for reasons cited before.
Thus, with advancing gestation, the mean platelet volume becomes an
insensitive measure of the platelet size. Post deli very platelet count increases in
reaction to and as a compensation for increased platelet consumption during the
process of delivery.
2.3.4 Haemostatic Profile
Pregnancy is associated with significant changes in the haemostatic profile.
Fibrinogen and clo tting factors VII, VIII, X, XII, vWF and ristocetin co -factor
activity increase remarkably as gestation progresses. Increased levels of
coagulation factors are due to increased protein synthesis mediated by the rising
estrogens levels. In in -vitro experime nts, pregnant plasma has been
demonstrated to be capable of increased thrombin generation (de Boer., et al.,
1989). Thus, pregnancy is a prothrombotic state. In pregnancy, a Partial
Prothrombin Time (PTT) is usually shortened, by up to 4s in the third trim ester,
largely due to the hormonally influenced increase in factor VIII. However, no
marked changes in PT or TT occur (Ramsay, 2010). There are changes in the
levels and activity of the natural anticoagulants also. Levels and activity of
Protein C do not c hange and remain within the same range as for non -pregnant
women of similar age. Levels of total and free (i.e., biologically available)
Protein S, decrease progressively with the advancement of gestation.

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Antithrombin levels and activity are usually stabl e throughout the pregnancy,
fall during labour and rise again soon after delivery.
Acquired activated Protein C (APC) resistance has been found to occur in
pregnancy, even when Factor V Leiden and anti -phospholipid antibodies are not
present (Clark et al., 1998). This has been attributed to the high factor VIII and
factor V activity and low free Protein S levels. Hence, APC sensitivity ratio
does not serve as a screening test for Factor V Leiden during pregnancy.
Coagulation factors remain elevated for up t o 8–12 weeks post partum and
assays for them may be falsely negative during this period. Markers of
haemostatic activity which are clinically relevant are thrombin –antithrombin
complexes (TAT) and prothrombin fragments (F 1 + 2), which reflect in vivo
thrombin formation, as also, tests which demonstrate plasmin degradation of
fibrin polymer to yield fragments, namely D -dimer and fibrin degradation
products (FDP) assay. TAT levels increase with gestation; in early pregnancy
the upper limit of normal is simil ar to the adult range of 2.63 g/L, whereas by
term, the upper limit of normal is 18.03 g/L. D -dimer levels are markedly
increased in pregnancy, with typical reference range being tenfold higher in late
pregnancy than in early pregnancy or in the non -pregna nt state (Ramsay, 2010).
The increase in D -dimers reflects the overall increase in total amount of fibrin
during pregnancy consequent to increased thrombin generation, increased
fibrinolysis or a combination of both (Eichinger, 2004). This also explains wh y

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the D -dimer assay is not reliable for predicting the possibility of venous
thrombo -embolism in pregnant patients (Kline et al., 2005).
2.4 PLATELETS (THROMBOCYTES)
Platelets , which are also known as ―thrombocytes‖, are blood cel ls whose role
(along with the coa gulation factors) is to stop bleeding (Laki, 1972). Platelets as
with erythrocytes do not have nucleus . They are products of the fragment ation
of a matured platelet precursor cell called the mega karyocytes derived from the
bone marrow, and then enter the circulation and at this stage they are in an
inactive form. Platelets are biconvex discoid st ructures , of about 2-3 μm in
diameter (Machlus et al., 2014; Jain, 1975) . Birmingham University physician
Dr Richard Hill Norris in 1880 was the first to describe the action of platelets .
The word thrombus is sometimes used interchangeably with the word clot,
regardless of its composition (white, red, or mixed). In other contexts it is used
to contrast a normal from an abnormal clot: thrombus arises from physiologi c
haemostasis, while thrombosis arises from a pathologic and excessive quantity
of clot (Fu rie and Furie, 2008) . Disease conditions associat ed with
throm bocytes are often caused by two major chang es in the number of
circulating platelets; thrombocytopaenia and thrombocytosis. The reference
range for platelets is 150,000 to 400,000 per cubic millim eter or 150 –400 ×
109 per litre (Ross et al., 1988) . Men usually have slightly higher mean values
than women (Ross et al., 1988).

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2.4.1 Thrombopoiesis
Platelets are products of the fragment ation of a matured platelet precursor cell
called the mega karyocytes derived from the bone marrow, and then enter the
circulation and at this stage they are in an inactive form . Megakaryocyte and
platelet production is mainly regulated by the hormone thrombopoieti n,
produced in the kidneys and liver. Each megakaryocyte can give rise to platelets
between 1,000 and 3,00 0 during its lifetime with an average of 1011 platelets
produced d aily in a healthy adult. Circulating platelets are usually stored in the
spleen (sequestration) , and are released when needed by splenic contraction
induced by the sympathetic nervous system. The average life span of circulating
platelets is 8 to 9 days and it is mainly controlled by the internal apoptotic
regulating pa thway, which has a Bcl -xL timer (Mason et al., 2007 ; Harker et al.,
2000) . Old platelets are destroyed by phagocytosis in the spleen and liver.

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Figure 2 : Haematopoeiesis showing precursor cells of thrombocytes
(Harker et al., 2000).

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Figure 3 : Thrombopoiesis (Harker et al., 2000).

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2.4.2 Thrombocytes and Haemostasis
Platelets will usually interact dependently with thrombin, Factors X, Va, VIIa,
XI, IX, and prothrombin to complete haemostasis via the coagulation cascade
(Bouchard et al., 2010; Ahmad et al., 1992). This means that a reduced in the
number of platelet below normal will result in derangement in blood clotting,
hence bleeding wil l occur. Platelets contain two types of granules, viz: dense
granules, lambda granules and alpha granules. When platelets are activated they
secrete the contents of these granules through their canal icular sys tems to the
exterior which are required for their proper functioning (Ahmad et al., 1992).
Granule characteristics of platelet
 α-granules – contain P-selectin, platelet factor 4, vWF, fibrinogen ,
transforming growth factor – β1, platelet -derived growth factor,
fibronectin, B -thromboglobulin , and coagulation factors V and XIII).
 δ-granul es (dense granule s) – contain ADP or ATP, calcium, and
serotonin .
 γ-granules – similar to lysosomes and also contain several hydrolytic
enzymes.
 λ-granules – contains contents that are involved in clot resorption during
later stages of blood vessel repair ( Boucha rd et al., 2010) .
The coagulation process is a complex series of enzymatic reactions
involving the proteolytic activation of circulating coagulation factors
(zymogens) and activity of co -factors (V, VIII), leading to the production of

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thrombin which conv erts soluble plasma fibrinogen into fibrin (as represented
diagrammatically in Fig. 3). The fibrin enmeshes the platelet plug, forming a
stable thrombus which prevents further blood loss from the damaged vessel
(Cheesbrough, 2006).

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Figure 4 : Diag rammatic representation of blood coagulation. The letter ‘ a’
after a factor indicates activation of factor (Cheesbrough, 2006).

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Figure 5 : Diagram matic representation of the structure of a platelet
showing the various granules (Bouchard et al., 2010) .

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2.5 HAEMOSTATIC CHANGES IN NORMAL PREGNANCY
Normal pregnancy is associated with major changes in the coagulation and
fibrinolytic systems, the concentration of many coagulation factors increasing
and plasma fibrinolytic activity decreasing as the concentrations of plasmin ogen
activator inhibitors progressively raise (Walker et al ., 1994) . Fibrinogen
increases from early pregnancy onwards to almost double its prepregnancy
value by term. Both factor VIII (FVIII) and von Willebrand factor (vWf) rise
steadily throughout pregna ncy. Factors VII and X also increase very
significantly during pregnancy, but the other vitamin K dependent clotting
factors, factors II and IX and factor XII, show a less significant or no rise and
factors XI and XIII may fall slightly (Walker et al., 199 4).
The platelet count does not normally change significantly during
pregnancy,although some authors have reported a slight drop in the count in the
third trimester. The bleeding time remains normal throughout
pregnancy.Screening tests used for the investi gation of bleeding; the activated
partial thromboplastin time (APTT) also called Partial thromboplastin time with
Kaolin and the prothrombin time (PT) are within normal adult ranges during
pregnancy, but in the third trimester the PT and the APTT are at th e lower
(shorter) limits of normal or slightly shortened, and this must be taken into
account when assessing coagulation screen results from pregnant women
(Walker et al., 1994).

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2.5.1 Blood Clotting Disorders in Pregnancy
2.5.2 Inherited bleeding disorde rs
In general, pregnant women with inherited bleeding disorders and the female
partners of men with these defects should be managed by an obstetric unit allied
with a haemophilia centre and a neonatal intensive care unit. The most common
inherited bleeding defects involve factor VIII deficiency (haemophilia A), factor
IX deficiency (haemophilia B), and von Willebrand factor deficiency (von
Willebrand's disease) (Walker et al., 1994).
2.5.3 Haemophilia A and haemophilia B
The prevalences of haemophilia A an d haemophilia B in the United Kingdom
population are 90 per 1 000 000 and 20 per 1 000 000, respectively. Most
female carriers of these X linked recessive disorders do not have major bleeding
problems, but in 10 -20% of carriers extreme Lyonisation results in a substantial
reduction of factor VIII or factor IX concentrations respectively (to < 40
units/dl) and, at the lowermost levels, a significantly increased risk of bleeding.
Statistically, 50% of the male children of carrier females will have haemophilia
and be at risk of serious bleeding. Very rarely, homozygous haemophilia A or B
may occur when a female is the offspring of a haemophilic father and a carrier
mother. These women have the same risk of major haemorrhage as do affected
males (Walker et al., 1994).

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2.5.4 Von Willebrand's disease
Von Willebrand's disease (vWD) is the most common clinically important
inherited abnormality of coagulation affecting women. Because of the very
wide spectrum of clinical presentation, it is difficult to ascertain pre cisely the
prevalence of all forms of vWD but it is more common than generally realised
and may be as high as 1% (Rodeghiero et al., 1987). Broadly, vWD falls into
three classes.
Type 1 vWD : the commonest type (about 75% of patients) is characterised by a
reduction in all forms of vWf, with the highest molecular weight forms
remaining detectable. Its inheritance can be autosomaldominant or recessive and
its expression veryvariable. In non -pregnant patients with type IvWD the vWf
concentrations (by antigen a ndby activity assay) are between 10 -40 units/dlas is
the FVIIHC. The bleeding time may beslightly to moderately prolonged but is
oftennormal and the bleeding tendency is generallymild (Walker et al., 1994).
Type 2 vWD : The feature common to all subvariants of type II vWD is the loss
of the highest molecular weight vWfmultimers. Inheritance is usually autosomal
dominant. In type II vWD the vWf is functionally impaired, the bleeding time is
usually prolonged, and bleeding episodes tend to be more severe and m ore
common especially in type IIA (the commonest type II subvariant -10% of all
patients with vWD) than in type 1. In subtype IIB (7% of all patients with
vWD) the abnormal vWf has enhanced interaction with platelet glycoprotein
(Gp) lb: these patients ofte n have mild to moderate thrombocytopenia. Infusion

26
of desmopressin (1 -desamino -8-Darginine vasopressin, DDAVP) may cause a
further fall in the platelet count in these patients and should therefore be
avoided in type IIB vWD (Walker et al., 1994).
Type 3 vWD : Type III vWD is clinically the most severe. The prevalence in
the United Kingdom of type III vWD is about one in 1 000 000. Patients with
type III vWD have no detectable or only trace amounts of vWf in their
circulation. Probably many patients with type III vWD are homozygous but
some may be compound heterozygotes. The condition is more prevalent in
cultures where consanguinity is common. In general the parents are clinically
unaffected or only mildly affected although their vWf concentrations may be at
or just below the lower limits of normal. Patients with type III vWD have only
1-2 units/dl factors VIIIC and therefore have a bleeding tendency which mimics
moderately severe haemophilia A with the added mucosal and subcutaneous
bleeding characteristic of vWD. Parents who have had a child with type III
vWD require follow up and careful counselling with respect to the management
of future pregnancies and the availability of prenatal diagnosis (Walker et al.,
1994).

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2.5.5 Prothrombin Time in Pregnancy
The P T is a screening test for the extrinsic clotting system, i.e. factor VII. It will
also detect deficiencies of factors, prothrombin, V, X, and fibrinogen
(Cheesbrough, 2006) . It is mainly used to monitor patients receiving warfarin
anticoagulation. As repor ted by Hellgren in 2003, prothrombin complex was
increased in the pregnant women studied, which was expressed as INR of less
than 0.9. Similarly, Uchikova et al ., 2005, reported PT as being significantly
shortened in pregnancy compared with the study contr ol. Also, in the work of
Durotoye et al ., 2012, PT was reported to be shortened in pregnant women
studied compared to the non -pregnant women. They also reported PT reduces as
pregnancy advances which shows that the production of coagulation factors
increas e as pregnancy progresses.
2.5.6 Partial Thromboplastin Time with Kaolin in Pregnancy
The APTT is a screening test of the intrinsic clotting system. It will detect the
inhibition or deficiency of one or more of the following factors: prothrombin, V,
VIII (antihaemophilic factor), IX, X, XI, XII and fibrinogen. The APTT is also
used to monitor patients being treated with heparin (Cheesbrough, 2006).
Significant difference has been reported in PTTK among pregnant and control
subjects, which shows that the le vel of factors in the intrinsic pathway are also
increased in normal pregnancy (Durotoye et al ., 2012). They also reported
insignificant differences in the result of PTTK in various trimesters.

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2.5.7 Blood Cells Morphology in Pregnancy
Physiological anaemi a is the term often used to describe the fall in haemoglobin
concentration that occurs during normal pregnancy results from plasma volume
increases above normal by the end of gestation although the red cell masses
itself increases by some and still leads t o a fall in haemoglobin concentration
with a feature of normocytic and normochromic type of anaemia which shows
normal red blood cells in a well stained smear. However, population of white
blood cells increases in pregnancy which also manifest in the micro scopy of the
stained smear (Hoffbrond et al. 2003). White cell count increased during
pregnancy with a typical reference range of 6 × 109–16 × 109/L, only WBC
more than 16 × 109/L is considered abnormal (Edlestam et al., 2001). Iron
deficiency and megalobl astic anaemia during pregnancy result from inadequate
intake of folate and iron to meet the increase requirements of pregnancy. The
prevalence of megaloblastic anaemia among pregnant is varies in different
populations, apparently depending on the nutrition al status of the population in
well nourished communities, this manifest as larger than normal red blood cells
in a well stained smear (Firki et al., 2002).
Myelocytes and metamyelocytes may be found in the peripheral blood film of
healthy women during pre gnancy and do not have any pathological
significance. Platelet count decreases during pregnancy, particularly in the third
trimester reported by Boehlen et al. 2000 termed ―gestational

29
thrombocytopenia.‖ found to have a platelet count of less than 150 × 109/Llate
in pregnancy (Anne Stiene et al., 1998).
2.5.8 Thrombocytopaenia
Thrombocytop aenia is defined as a platelet count of less than 150 × 103per μL.
It is often discovered incidentally when obtaining a complete blood count
during an office visit. The aetiology usually is not obvious, and additional
investigation is required. Patients with platelet counts greater than 50 × 103per
μL rarely have symptoms. A platelet count from 30 to 50 × 103per μL rarely
manifests as purpura. A count from 10 to 30 × 103per μL may cause bleeding
with mini mal trauma. A platelet count less than 5 × 103per μL may cause
spontaneous bleeding and constitutes a hematolog ic emergency. Patients who
present with thrombocytopenia as part of a multisystem disorder usually are ill
and require urgent evaluation and treatment (Bouchard et al., 2010) .
2.5.8.1 Aetiology and Classification of Throbocytopaenia
Causes of thrombocytop enia can be classified by mechanism and include
decreased platelet production, increased splenic sequestration of platelets with
normal platelet survival, increased plat elet destruction or consumption (both
immunologic and non -immunologic causes), dilution of platelets, and a
combination of these mechanisms.

30
Table 1: Aetiological classification of Thrombocytopaenia
Cause Conditions
Diminished or absent
megakaryocytes in bone
marrow Aplastic anemia, Leukemia, Myelosuppressive drugs
(e g, hydroxyurea, inte rferon alfa -2b, chemotherapy
drugs), Paroxysmal nocturnal hemoglobinuria (some
patients)
Diminished platelet production
despite the presence of
megakaryocytes in bone
marrow Alcohol -induced thrombocytopenia, Bortezomib use,
HIV-associated thrombocytopenia , Myelodysplastic
syndromes (some), Vitamin B 12 or folate (folic acid)
deficiency
Platelet sequestration in
enlarged spleen Cirrhosis with congestive splenomegaly, Gaucher
disease, Myelofibrosis with myeloid metaplasia
Immunologic destruction Connective tissue disorders, Drug -induced
thrombocytopenia, HIV -associated
thrombocytopenia, Immune thrombocytopenia,
Lymphoproliferative disorders, Neonatal
alloimmune thrombocytopenia, Post -transfusion
purpura
Non-immunologic destruction Certain systemic infection s (e g, hepatitis, Epstein –
Barr virus, cytomegalovirus, or dengue virus
infection)
Disseminated intravascular coagulation, Pregnancy
(gestational thrombocytopenia), Sepsis,
Thrombocytopenia in acute respiratory distress
syndrome
Thrombotic thrombocytopenic purpura –hemolytic –
uremic syndrome
Dilution Massive RBC replacement or exchange transfusion
(most RBC transfusions use stored RBCs that do not
contain many viable platelets)
(Bouchard et al., 2010) .

31
2.6 THROMBOCYTOPAENIA IN PREGNANCY
Thrombocytopenia is the second most common haematological finding in
pregnancy after anaemia. It affects 7 -10% of all pregnant women (Verdy et al.,
1997). The normal range of platelets in non -pregnant women is 150,000 –
400,000/μL. Thrombocytopenia is defined as a drop in plat elet count below
150,000/μl. Pregnancy is associated with a physiological fall in the platelet
count with a leftward shift in the platelet count distribution. The cause for the
physiologic decrease in platelet count is multifactorial and is related to
haem odilution, increased platelet consumption, and increased platelet
aggregation driven by increased levels of thromboxane A2 (Fay et al ., 1983).
Platelet count may also be lower in women with twins as compared with
singleton pregnancies, perhaps due to a gre ater increase in thrombin generation
(Tsunoda et al ., 2002). Pregnant women with thrombocytopenia tend to have
fewer bleeding complications than non -pregnant women due to the pro –
coagulant state induced by increased levels of fibrinogen, factor VIII and vo n
Willebrand factor, suppressed fibrinolysis and reduced protein S activity. There
are several other pregnancy -related conditions that can also lead to
thrombocytopenia. Thrombocytopenia in pregnancy is a common reason for
haematology consultation. It is a diagnosis of exclusion, occurring in the late
half of pregnancy, from the mid -second to third trimester. Women are typically
asymptomatic. Platelet count is typically greater than 70,000/μL, with about
two-thirds being 130,000 -150,000/μl. There is usuall y no past history of

32
thrombocytopenia. Gestational thrombocytopenia can recur; the risk of
recurrence, however, is unknown (Burrows and Kelton, 1993).

Fig. 6: Histogram of platelet counts of pregnant women in the third
trimester (n = 6770) compared w ith non -pregnant women (n = 287) (Fay et
al., 1983) .
Thrombocytopaenia in pregnancy may be termed as gestational
thrombocytopaenia, other causes of thrombocytopaenia in pregnancy include;
Immune thrombocytopaenic purpura (ITP),HELLP (haemolysis, elevated l iver

33
enzymes, low platelets) syndrome, Thrombot ic microangiopathy, Acute fatty
liver of pregnancy (AFLP),Miscellaneous causes of thrombocytopaenia
2.6.1 Gestational Thrombocytopaenia
Gestational thrombocytopenia occurs in approximately 8% of all pregnanci es
and accounts for more than 70% of cases with thrombocytopenia in pregnancy.
Although the pathophysiology of gestational thrombocytopenia is unknown, it is
thought to be related to increased activation and peripheral consumption
(Bockenstedt, 2011).
It is a diagnosis of exclusion, occurring in the latter half of pregnancy, from the
mid-second to third trimester. Women are typically asymptomatic. Platelet
count is typically greater than 70,000/μL, with about two -thirds being 130,000 –
150,000/μl (Burrows an d Kelton, 1993). There is usually no past history of
thrombocytopenia. Gestational thrombocytopenia can recur; the risk of
recurrence, however, is unknown.
Gestational thrombocytopenia remains a clinical diagnosis. The main competing
diagnosis is immune th rombocytopenic purpura (ITP), which is usually
considered if the degree of thrombocytopenia is more significant. However,
there are reports of more severe thrombocytopenia that showed no response to
steroids, and which resolved postnatally; consistent with gestational
thrombocytopenia (Win et al ., 2005). Unfortunately, there are no laboratory
tests to differentiate between the two conditions. The existence of pre –
pregnancy thrombocytopenia should rule out gestational thrombocytopenia.

34
However, previous preg nancies complicated by thrombocytopenia would favour
gestational thrombocytopenia. In addition, response to immune modulation with
steroids or immunoglobulins would favour ITP (Usha and Lori, 2013).
During the antepartum period, no treatment is necessary i f the patient is
asymptomatic. Platelet count monitoring is recommended periodically,
depending on the degree of thrombocytopenia. For planning labour and
delivery, it may be helpful to obtain a platelet count at around 36 weeks of
gestation (Usha and Lori , 2013).
The degree of thrombocytopenia is generally not severe enough to increase the
risk of bleeding with delivery. Patients with platelet count greater than 30 –
50,000/μl should be able to undergo vaginal or surgical delivery safely without
increased risk. Altho ugh epidural anaesthesia is relatively safe with platelet
count > 50,000/μl, there remains controversy regarding the threshold above
which epidural anaesthesia is safe. Due to variations in practice, anaesthesia
consultation should be obtained prior to del ivery to determine specific
anaesthesia recommendations. For patients with significant thrombocytopenia,
it is important to counsel them that their individual pregnancy plans may need
to be modified depending on the platelet count at the time of labour. If platelet
counts are below the acceptable level for anaesthesia, a short course of steroids
or intravenous immunoglobulins (IVIg) may be considered, with the thought
that this might be a missed diagnosis of ITP. Gestational thrombocytopenia is
self-limitin g and resolves within 1 to 2 months after delivery. It is not associated

35
with adverse outcomes for the baby. However, given the significant overlap
between gestational thrombocytopenia and ITP, consideration should be made
for evaluating the infant‘s plate let count after delivery (Usha and Lori, 2013).
2.6.2 Immune Thrombocytopenic Purpura (ITP) in Pregnancy
Immune thrombocytopenia occurs in 1 in 1000 -10,000 pregnancies, accounting
for 3% of all thrombocytopenic gravidas. It is the most common cause of
thrombocytopenia in the first and second trimesters (Segal and Powe, 2006).
ITP is an autoimmune disorder caused by development of immunoglobulin G
(IgG) autoantibodies that are directed against several platelet glycoproteins
(Schwartz, 2007). Antibody -boun d platelets are rapidly cleared from maternal
circulation once they bind to specific antibody receptors on macrophages, found
mainly in the spleen and also in the liver (Schwartz, 2007). IgG antibodies can
cross the placenta and have the potential to cause thrombocytopenia in the
infant (Garty et al., 1994).
ITP is a clinical diagnosis and requires ruling out other causes of
thrombocytopenia. The diagnostic approach is not different in pregnant women
compared to non -pregnant women. Women can present with br uising, mucosal
bleeding and petechiae or they may be asymptomatic, with the severity of
symptoms directly proportional to the degree of thrombocytopenia. There is no
specific test that differentiates ITP from other causes of thrombocytopenia. As
primary I TP is a diagnosis of exclusion, causes of secondary thrombocytopenia
such as HIV infection, hepatitis C and autoimmune disease should be ruled out

36
clinically or with laboratory testing. ITP may be indistinguishable from
gestational thrombocytopenia; howeve r, patients with ITP usually have a prior
history of ITP or other immune -mediated disorders (Stavrou and McCrae, 2009;
Burrows and Kelton, 1993; 1988). Antecedent thrombocytopenia prior to
pregnancy, persistence after delivery and response to ITP directed therapy
(steroids, IVIG) makes ITP a more likely diagnosis. ITP is more likely to
present with severe thrombocytopenia earlier on in pregnancy compared with
gestational thrombocytopenia (George et al ., 1996). Since both gestational
thrombocytopenia and ITP are clinical diagnoses and are diagnoses of
exclusion, both deserve close follow up during and after delivery. For both
gestational thrombocytopaenia and ITP, there should be no additional
haematological abnormalities, no microangiopathy, or evidence of d isseminated
intravascular coagulation (DIC) and liver insufficiency (George et al., 1996).
The clinical management of the pregnant woman with ITP requires close
consultation between the obstetrician and the haematologist. The decision to
treat thrombocytop enia is determined by the patient‘s symptoms and the level of
thrombocytopaenia. The goal of therapy is to prevent bleeding, and treatment is
generally not required in patients with platelet counts greater than 20,000 to
30,000/μl if they are not symptomat ic. If the patient is asymptomatic and
platelet count is above 20,000/μl, close monitoring is recommended. Patients
with thrombocytopenia should be evaluated for symptoms and platelet count
monitored once a month during the first and second trimesters. The frequency

37
of serial platelet count monitoring should be increased as term approaches or
thrombocytopenia worsens. Treatment is recommended for women with a
platelet count below 10,000/μl at any time during pregnancy (Kelton, 2002).
Treatment should also b e considered if platelet counts are below 30,000/μl in
the third trimester due to the potential for imminent delivery. Platelet
transfusion alone is not helpful due to the quick destruction of transfused
platelets as evidenced by a poor increment in the po st-transfusion platelet count.
Glucocorticoids such as prednisone are generally the first line of therapy.
Glucocorticoids block antibody production and thereby reduce the phagocytosis
of antibody -coated platelets by the reticuloendothelial system in the s pleen. The
typical starting dose is 1 mg/kg of prednisone based on the pre -pregnancy
weight with a quick taper once a response is achieved.
Response to therapy is not instantaneous, and may take several days to weeks.
Glucocorticoids can cause several uniq ue toxicities in pregnancy, such as
gestational diabetes and pregnancy -induced hypertension. These agents may
also be associated with premature rupture of faetal membranes and placental
abruption (Kelton, 2002). Hence, they should be used sparingly with th e
minimal effective doses employed. IVIg may be used as first line or in steroid
resistant patients. The therapeutic response to IVIg is attributable to several
different immunological mechanisms, including blockage of splenic
macrophages (Baerenwaldt et al ., 2010). High dose IVIg of 1gm/kg over 2 -5
days is effective in raising the platelet count rapidly; however, the effects are

38
transient, generally lasting 1 to 4 weeks. The post -transfusion increment with
platelet transfusion increases markedly after IVIg infusion. In life threatening
bleeding, IVIg followed by platelet transfusion with or without steroids may be
required. Intravenous anti -D could be considered in non -splenectomised Rh
positive patients who are resistant to steroids and IVIg. Anti -RhD is a pooled
IgG product taken from the plasma of RhD -negative donors who have been
immunized to the D antigen. Anti -RhD immunoglobulin binds to maternal red
blood cells. Presentation of the antibody -bound red blood cells to Fc receptors
in the spleen results in preferential splenic phagocytosis of the red blood cells
rather than platelets. Response occurs in 75% of patients within 1 -2 days, with
peak effect at 7 -14 days and the duration of therapy up to 4 weeks (Cooper,
2009). Although experience is limited in p regnant women, the response rates
are comparable to IVIg (Michel et al ., 2003). Anaemia and jaundice in the
infant have been reported. Splenectomy should be deferred if possible as the
severity of the thrombocytopenia may spontaneously improve after delive ry
(Moise, 1991). It is reserved only for severe refractory ITP and is generally
performed in the second trimester, owing to the risks of inducing premature
labour in the first trimester and obstruction of the surgical field by the gravid
uterus in the thi rd trimester (Verdy et al ., 1997). Rituximab is increasingly
being used to treat ITP in non -pregnant women; however, it is classified as a
class C drug in pregnancy. As information is limited in pregnancy it should be
avoided unless there are no other opti ons. Other agents used in the treatment of

39
the non -pregnant women with ITP, such as cytotoxic and immunosuppressive
agents, are discouraged during pregnancy due to the potential teratogenic
effects. The thrombopoietin receptor agonists such as Romiplostim and
Eltrombopag stimulate platelet production by binding to the platelet
thrombopoietin receptor and have been approved for treatment of chronic ITP in
adults. These agents should be avoided in pregnancy as there is no information
on the reproductive effec ts. Episodes of severe bleeding are rare, even with very
low platelets. The most common complication in the peripartum time period
relates to the use of regional anaesthesia during delivery (Verdy et al., 1997).
Vaginal delivery is the preferred method of delivery. Caesarean section should
be reserved for obstetric indications only. The risk for intracranial haemorrhage
in infants of women with ITP is very low. In addition, vaginal delivery does not
increase the risk of intracranial haemorrhage compared to caesarean section
(Kelton, 2002). Regional anaesthesia during labour in thrombocytopenic
patients is controversial due to the increased risk of epidural hematoma; the
decision regarding epidural anaesthesia should be made in consultation with the
anaesthes iologist (van Veen et al., 2010). Thrombocytopenia in infants born to
women with ITP is uncommon. Fortunately, 90% of these infants will not have
significant thrombocytopenia. Only 10% develop more severe
thrombocytopaenia with platelet counts below 50,000 /μl and platelet counts
below 20,000/μl occur in approximately 4% of infants (Burrws and Kelton,
1993). Even when severe thrombocytopenia occurs in the newborn, bleeding

40
problems are rare and can be easily treated. Faetal platelet count monitoring
with sca lp sampling is unreliable and not recommended as the risks of the
procedure outweigh the benefits. However, the platelet count should be
measured in the neonate at birth and for several days following delivery as
faetal platelet counts continue to drop aft er delivery with nadir 1 to 2 days.
Paediatricians or primary care providers should be notified about the mother‘s
hematologic condition so that they can take appropriate steps for management
of the infant (Kelton, 1983).
2.7 PREECLAMPSIA
Thrombocytopenia is usually moderate and platelet count rarely decreases to
less than 20,000/μl. Thrombocytopenia in patients with preeclampsia always
correlates with the severity of the disease (Perepu and Rosenstein, 2013).
2.8 HELLP (HAEMOLYSIS, ELEVATED LIVER ENZYMES , LOW
PLATELETS) SYNDROME
HELLP affects 0.5 -0.9% of all pregnancies and develops in 10% of patients
with preeclampsia (Kirkpatrick, 2010). It is characterized by haemolysis,
elevated liver enzymes and low platelets. The pathophysiology is similar to
preecl ampsia, with endothelial damage and release of tissue factor and
coagulation activation. A recent study identified mutations in genes that
regulate the alternative complement system, suggesting that excessive
complement activation may be involved in pathog enesis similar to atypical
haemolytic uremic syndrome (atypical HUS) (Fakhouri et al ., 2008). The

41
criteria for HELLP syndrome vary among studies, but generally include
microangiopathic haemolytic anaemia, increased AST more than 40 -70 U/ml
and thrombocytop enia with platelet counts less than 100,000/μl (Barton and
Sibai, 2004). HELLP syndrome may represent advanced preeclampsia, although
15-20% presenting with HELLP do not have antecedent hypertension and
proteinuria. It occurs predominantly in the third tri mester between 28 -36 weeks
of gestation, although a small percentage can occur prior to 27 weeks (Haeger et
al., 1992; Haram et al., 2009). HELLP syndrome, like preeclampsia, can occur
postpartum with 30% of patients presenting within 48 hrs of delivery. I t may
even occur up to one week after delivery. Unlike preeclampsia, HELLP is more
common in multiparous women. Patients present with abdominal pain and
tenderness in the epigastrium and right upper quadrant, which may be
accompanied by nausea, vomiting an d malaise. Hypertension and proteinuria
are present in 85% of cases. Generalized oedema precedes the syndrome in
more than half the cases (Haram et al ., 2009). Although thrombocytopenia is
present, bleeding is not typical. HELLP can be difficult to differe ntiate from
preeclampsia; however a typical patient typically does have hypertension and
proteinuria. Thrombocytopenia is much more severe in HELLP than in
preeclampsia. Thrombotic microangiopathies causing thrombocytopenia are
also difficult to distinguis h from HELLP. The PT and PTT are prolonged and
factors V, VIII and fibrinogen are decreased in HELLP syndrome, compared to

42
thrombotic microangiopathies. Delivery of the foetus is the key to the
management of HELLP. Delivery is indicated if pregnancy is gre ater than
34 weeks gestation, there are signs of faetal distress and maternal multiorgan
damage. Patients may require platelet transfusion if there is evidence of
bleeding. The laboratory abnormalities in HELLP syndrome usually peak 24 -48
hours following d elivery. In the absence of other complications, such as renal
dysfunction or DIC, platelet counts tend to rise between the fourth and the sixth
day post -partum. When severe laboratory abnormalities persist after 72 hours,
the use of plasma exchange and glu cocorticoids can be considered. In a small
study28, this approach was demonstrated to induce a rapid remission. For
pregnancies fewer than 34 weeks gestation, where there is no maternal and
faetal distress, glucocorticoids are recommended to accelerate fae tal pulmonary
maturity followed by delivery in 48 hours. Observation alone without a plan for
delivery is not generally recommended because the condition rarely reverses
until delivery of the baby (Perepu and Rosenstein, 2013).

2.9 THROMBOTIC MICROANGIOPA THY
Thrombotic thrombocytopenic purpura (TTP) and haemolytic uremic syndrome
(HUS) are collectively referred to as thrombotic microangiopathies and are not
pregnancy specific, although they occur with increased frequency during
pregnancy, with an incidence of 1 in 25,000 (Dashe et al., 1998). The incidence
is greater in the second and third trimesters. Patients with pregnancy associated

43
TTP are at risk for development of recurrent TTP in subsequent pregnancies.
The exact aetiology is unknown, but endothelia l damage is suspected to be the
initiator. TTP is strongly associated with a severe deficiency of ADAMTS -13, a
metalloproteinase that cleaves ultra -large von Willebrand factor (vWF)
multimers, the most haemostatically active form of vWF. Deficiencies of
ADAMTS -13 are usually acquired, resulting from neutralizing antibodies,
although congenital deficiency accounts for a minority of cases (Tsai and Lian,
1998). Increased levels of ultra -large vWF species promote platelet
agglutination and thrombotic occlusion of the microvasculature. Both TTP and
HUS are essentially clinical diagnoses. While assays are available to measure
functional activity of ADAMTS -13 in most institutions, the results are not
available in a timely enough manner to make them clinically usef ul. Therefore,
to make the diagnosis, one needs to have evidence of microangiopathic
haemolytic anaemia (MAHA) such as the presence of schistocytes, laboratory
evidence of haemolysis and thrombocytopenia. Other hallmarks of advanced
disease are neurologica l dysfunction, fever and renal insufficiency. The
differentiation between TTP, HUS and HELLP can be difficult or even
impossible, especially when the onset is during the second and third trimesters.
Delivery leads to resolution of preeclampsia but not TTP/ HUS. If suspected
preeclampsia/HELLP does not improve within 48 -72 hours after delivery,
TTP/HUS should be considered (Perepu and Rosenstein, 2013).

44
2.10 ACUTE FATTY LIVER OF PREGNANCY (AFLP)
AFLP is a rare disorder with an incidence of 1 in 10,000 -15,000 pregnancies,
maternal mortality of 18% and faetal mortality of 23% (Lee et al ., 2009).
Patients are usually nulliparous and there is an increased incidence in twin
pregnancies.
AFLP is thought to be due to abnormalities in intramitochondrial fatty acid beta
oxidation. Maternal heterozygosity for long chain 3 hydroxyacyl CoA
dehydrogenase deficiency leads to reduced oxidation of the fatty acids. This
combined with dietary factors exacerbate the condition. When a heterozygous
woman carries a foetus that is homozygous, faetal hepatotoxic fatty acids
accumulate and return to maternal circulation, causing maternal liver and
vascular damage (Ibdah et al., 1999).
Patients usually present in the third trimester with nausea, vomiting, malaise,
right upper quadrant pain and cholestatic liver dysfunction (Fesenmeier et al.,
2005). Laboratory findings include normal to low platelet count, normochromic
normocytic anaemia, elevated leukocyte count, prolonged prothrombin time,
and low fibrinogen and antithrombin III (AT III) levels along with raised
transaminases. AFLP is more likely associated with liver and renal failure and
concomitant coagulopathy, hypoglycaemia and encephalopathy than HELLP
syndrome. Half of the patients will have criteria for preeclampsia and some m ay
have features that overlap with HELLP. DIC is the hallmark for AFLP, whereas
only 7% with preeclampsia and 20 -40% with HELLP have DIC. Intensive

45
supportive care with blood product support for the underlying coagulopathy
along with immediate termination of pregnancy is recommended as spontaneous
recovery during pregnancy is unlikely (Perepu and Rosenstein, 2013).
2.11 MISCELLANEOUS CAUSES OF THROMBOCYTOPAENIA
Pseudothrombocytopaenia, Disseminated intravascular coagulation, Nutritional
deficiencies, Hepar in induced thrombocytopenia (HIT), Type 2B von
Willebrand disease, Marrow infiltrative disorders, Autoimmune disease such as
systemic lupus erythematosus and antiphospholipid syndrome, Other inherited
thrombocytopaenia (Perepu and Rosenstein, 2013).
2.12 SYMPTOMS OF THROMBOCYTOPAENIA
Thrombocytopenia (count is below 20,000 per μL ) is usually charac terized by
spontaneous formation of bruises (purpura) and petechiae (tiny bruises), especially
on the extremities, bleeding from soft tissues such as from the nostrils , vaginal,
and gums, and menorrhagia (excessive menstrual bleeding), platelet (Cines and
McMillan, 2005). Severe thrombocytopenia (<10,000 per μL) may result in the
spontaneous formation of haematomas (blood masses) in the mouth or on
other mucous membranes. There is usually a prolonged Bleeding time from
minor lacera tions or abrasions . Haematomas in the C entral nervous system may
lead to stroke and sudden death (Cines and McMillan, 2005).

46
Platelet disorders result in a typical pattern of bleeding:
 Multiple petechiae in the skin (typically most evident on the lower legs)
 Scattered small ecchymoses at sites of minor trauma
 Mucosal bleeding (oropharyngeal, nasal, Gastrointestinal -GI,
Genitourinary -GU)
 Excessive bleeding after surgery.
Heavy GI bleeding and bleeding into the CNS may be life threatening.
However, bleedi ng into tissues (e g, deep visceral hematomas or hemarthroses)
rarely occurs with thrombocytopenia, which causes immediate, superficial
bleeding following an injury. Bleeding into the tissues (often delayed for up to a
day after trauma) suggests a coagulat ion disorder (e g, hemophilia) (Kuter
2014).

Figure 7: Purpura resulting from thrombocytopenia ( Kuter 2014 ).

47
2.12 Diagnosis of Thrombocytopaenia
Tradi tionally, laboratory tests performed should include: full blood count
(FBC) , platelet count, liver enzymes studies , erythrocyte sedimentation
rate, renal functio n test, vitamin B12 levels, folic acid levels , and peri pheral
blood smear. If diagnosis remains unclear, a bone marrow biopsy may be
performed , to differentiate whether the thrombocytopaenia is due to decreased
production or peripheral destruction. Modern diagnosis of thrombocytopaenia
involves the detection of mutation which might have occurred in the gene
coding for platelet production (Kuter 2014).
2.13 Platelet Count
A platelet count may be requested to investigate abnormal skin and mucosal
bleeding which can occur when the platelet count is very low (usually below
20×109/l). Platelet counts are also performed when patients are being treated
with cytotoxic drugs or other drugs which may caus e thrombocytopenia
(Cheesbrough, 2006).
2.14 Prothrombin time (PT) test
The PT is a screening test for the extrinsic clotting system, i.e. factor VII. It will
also detect deficiencies of factors, prothrombin, V, X, and fibrinogen. It is
mainly used to mon itor patients receiving warfarin anticoagulation
(Cheesbrough, 2006) .

48
CHAPTER THREE
3.0 MATERIAL AND METHODS
3.1 Study Area
This study was carried out in the town of Ekpoma, Esan West Local
Government, Edo State Nigeria.
3.2 Study Populatio n:
The study group consist of pregnant women attendi ng antenatal clinic at general
Hospital Ekpom a, Edo State .
3.2.1 Inclusion criteria
Pregnant women (with normal pregnancy, multiple pregnancy and complicated
pregnancy) attending antenatal clinic in Esan W est Local Government Area,
Edo State.
3.2.2 Exclusion criteria
Non pregnant women.
3.3 Sample Collection
5ml of blood was collected(with minimum of stasis) into a plastic tube
containing 1ml of aqueous tri -sodium citrate .without delay ,the sample will be
centrifuged at 1200g for 15minutes.The plasma was pippeted out unto a plain
plastic tube and kept frozen for analysis of clothing disorder. Blood sample was
also collected into EDTA container for platelet count.

49
3.4 LABORATORY ANALYSIS
Laboratory Equip ment: the laboratory equipment used in the analysis of
sample includes;
Microscope
Water bath
Micropipette
Neubauer Counting Chamber
Petri dish
Microscope
Staining rack
Wash Bottle
Glass Slides
3.5 PLATELET COUNT
Relevance of test : platelet count may be r equested to investigate abnormal skin
and mucosal bleeding which can occur when the platelet count is very low
(usually below 20×109/l). Platelet counts are also performed when patients are
being treated with cytotoxic drugs or other drugs which may cause
thrombocytopenia.
Principle of Test : when blood is diluted 1 in 20 in a filtered solution of
ammonium oxalate reagent which the red cells lyses. Platelets are then
countedmicroscopically using an Improved Neubauer ruled countingchamber
and the number of pl atelets per litre of blood calculated.

50
Blood sample: EDTA anticoagulated venous blood was used.
Reagent
Ammonium oxalate 10 g/l (1% w/v) diluting fluid. This was filtered the fluid
before use.
Test method
The test was performed within 2 hours of blood coll ection.
0.38 ml of filtered ammonium oxalate diluting fluid was measured and
dispensed into a tube.
0.02 ml of well-mixed anticoagulated venous blood was added and mixed.
The counting chamber assembled and filled with well-mixed sample.
The chamber was lef t undisturbed for 20 minutes. To prevent drying of the
fluid, the chamber was placed in a petri dish on dampened tissue and cover with
a lid.
The underside of the chamber was dried and placed on the microscope stage.
Using the 10× objective, the rulings of the grid was focused and the central
square of the chamber was brought into view. The platelets were counted in the
designated squares
Platelet calculation details
Platelet count (per Litre) = Counted Cells x 20 x 106
0.2 x 0.1
Where: 20 = dilu tion factor (1 in 20)

51
0.2mm2 = counted area
0.1mm = depth of chamber.
3.5.1 Prothrombin time (PT) test
The PT is a screening test for the extrinsic clotting system, i.e. factor VII. It will
also detect deficiencies of factors, prothrombin, V, X, and fib rinogen. It is
mainly used to monitor patients receiving warfarin anticoagulation.
Principle of the test
Plasma or capillary blood is added to a thromboplastin and c alcium chloride
reagent at 37oC and the time taken for a clot to form is measured. The clo tting
time in seconds is converted to the International Normalized Ratio (INR),
usually by reference to a table provided by the manufacturer of the reagent or
from the formula.
INR = (PT of Patient/PT of Control)ISI
*International sensitivity Index (ISI): This figure is provided by the
manufacturer of the thromboplastin reagent. To obtain the INR, calculate the
prothrombin ratio, log the ratio, multiply by the ISI, and antilog the result.
Reagents
Thromboplastin calcium chloride combined reagent.
Test Pro cedure
0.25 ml of the thromboplastin/calcium reagent was pipetted into a small glass
tube. The tube was placed in a 37oC water bath for 1 –2 minutes.

52
Using a calibrated capillary or delivery pipette, 50 µl(0.05 ml) of capillary
plasma was added, mixed, and started the stop -watch.
The tube was held in the water bath and the mixture tilted back and forth
looking for clot formation. When a clot formed, the stop -watch stopped and the
time was recorded in seconds.
3.5.2 Estimation of PTTK
PTTK is also called Ac tivated partial thromboplastin time (APTT) test
The APTT is a screening test of the intrinsic clottin g system. It will detect the
inhibition or deficiency of one or more of the following factors: prothrombin,
V, VIII (antihaemophilic factor), IX, X, XI, XI I and fibrinogen. The APTT is
also used to monitor patients being treated with heparin.
Principle of test
Kaolin (surface activator) and platelet substitute (phospholipid) are incubat ed
with citrated plasma at 37oC for the time specified in the test method . Calcium
chloride (CaCl 2) is added and the time taken for the mixture to clot is measured
Reagents
Kaolin/platelet substitute mixture
Calcium chloride, 0.025mol/l (25 mM)
Control plasma

53
Test Method
Method using Kaolin/Platelet substitute mixture
0.2 ml of well -mixed kaolin/platelet substitute was pipette into a small glass
tube.
0.1ml of plasma was added , mixed , and incubate at 37 ͦ C for exactly 2 minutes
(tilting the tube at intervals).
0.1 ml 0.025 mol/l calcium chloride was added, mixed and the stop -watch
started.
The tube was held in the water bath and tilted the mixture back and forth,
looking for clot formation.
When a clot forms, the stop -watch was stopped and the time recorded.
3.5.3 Blood Cell Morphology
Value of blood films: Examination of t hin blood films is important in the
investigation and management of anaemia, infections, and other conditions
which produce changes in the appearance of blood cells and differential white
cell count. A blood film report can provide rapidly and at low cost, useful
information about a patient‘s condition.
3.5.3.1 Making, Fixing and Staining Blood Films
Thin blood films were made from well mixed EDTA anticoagulated blood. To
prevent EDTA associated blood changes, blood films from EDTA
anticoagulated blood were made with as little delay as possible, i.e. within 1

54
hour of collecting the blood. The making, fixing, staining and examination of
smears were carried out as described by Cheesbrough (2006) .
3.5.3.2 Making thing film
 A drop of blood was place close to on e end of a clean dry slide
 Using a clean smooth edged spreader, the spreader was drawn back to
touch the drop of blood and allow the blood to extend along the edge of
the spreader. Holding the spreader at an angle of about 45 ͦ, the drop of
blood was sprea d to make a film about 40 –50 mm in length (two thirds of
the slide).
 The films were allowed to air dry and were protected from dust and
insects.
 When completely dried and within a few minutes of making the blood
film, the films were fixed in absolute metha nol.
3.5.3.3 Staining films with Leishman Stain
 The blood films were covered with undiluted Leishman stain but do not
flood the slide.
 After two minutes, the stain on the films was diluted with buffered water
(pH = 6.8) and were left for 8 minutes.
 After which the stain was wash off with buffer and the slides left to air
dry.

55
3.5.3.4 Microscopy
The slides were examined microscopically with x 100(emersion oil objective ).
3.6 Statistical Analysis
The result obtained was analysed using the Statistical Packag e for Social
Sciences (SPSS). P-value of 0.05 was regarded significant. ANOVA, T -test,
correlation was done. Values were represented as mean±SEM (n=300) and
analysed using paired sample T -test at p=0.05; SPSS version 20.

56
CHAPTER FOUR
4.0 RESULTS
Table 2 : measured parameters in pregnant and non -pregnant women

Values with different superscripts are significantly different (p<0.05), across the
row per parameter. The prothrombin time and INR of the test subjects showed a
significant increase compared to those of the control p<0.05. However, the
PTTK and PLT count showed no significant change when compared to that of
the control results at p>0.05 .

Measured variable Pregnant women
(n = 300) Non-pregnant
women (n = 100) P-value
PT (seconds) 17.05 ± 0.05 16.03 ± 0.01 0.041
PTTK (seconds) 41.02 ± 0.10 42.36 ± 0.08 0.060
INR 1.26 ± 0.02 1.15 ± 0.01 0.031
PLT (109/L) 176 ± 30.32 195.34 ± 44.0 0.069

57
Table 3 : PT/PTTK, INR, PLT in pregnant women at different trimesters of
pregnancy.

Values with different superscripts are significantly different (p<0.05), across the
row per parameter. The prothrombin time and INR show ed a significant
increase with duration of pregnancy at p<0.05. However, the PTTK and PLT
count showed no significant change at p>0.05 using Duncan‘s multiple range
and ANOVA. Trimester
Measured variable 1st trimester
n= 100 2nd trimester
n= 100 3rd trimester
n= 100 P-value
PT (seconds) 16.38±0.10a 17.09±0.08b 17.75±0.07c 0.046
PTTK (seconds) 42.29±0.30a 41.63±0.30 a 40.99±0.30 a 0.086
INR 1.16±0.01a 1.21±0.01b 1.28±0.01c 0.044
PLT (109/L) 184.41±2.00a 167.22±4.00a 179.92±3.00a 0.078

58
Table 4 : Plasma platelet levels of the different trimesters and the control
group (mean ± SEM).
Control
n = 100 First trimester
n = 100 Second trimester
n = 100 Third trimester
n = 100
Platelets (109/L) 195.34±44.00a 184.41±2.00a 167.22±4.00a 179.92±3.00a
F-value
(sig.level) 4.193(p=0.069) 6.994(p=0.082) 7.151(p= 6)
Values with d ifferent superscripts are significantly different (p<0.05), across
the row per parameter. The plasma platelet levels showed non -significant
decrease in first and second trimesters, as well as, in the third trimester,
compared to the non -pregnant control gr oup (p<0.05), by paired sample
students‘ t -test and Duncan‘s multiple range analysis of variance.

59

Figure 8 : Trend of Platelets level with duration of pregnancy.
The pl asma platelet levels showed no significant decrease in first and second
trimester s, as well as, in the third trimester, compared to the non -pregnant
control group s (p<0.05).

60
Table 5 : Red Blood cell Morphology in the different trimesters.
Morphology First trimester Second trimester Third trimester
F % F % f %
Microcytic
hypo chromic blood
film platelets 156 52.00 120 40.00 90 30.00
Normocytic
hypochromic blood
film platelets 144 48.00 180 60.00 210 70.00
The microcytic hypochromic blood film showed a steady decrease; whereas the
normocytic hypochromic blood film platelets di splayed the opposite.

61

Figure 9 : Line plot of the morphological status of the different trimesters.
The microcytic hypochromic blood film showed a steady decrease; whereas the
normocytic hypochromic blood film platelets displayed the opposite.

62
Table 6 : Correlation between platelets and PT, APTT, 1NR, in first,
second, and third trimesters of pregnancy.
PARAMETERS 1st TRIMESTER 2ND TRIMESTER 3RD TRIMESTER
r-value p-value r-value P-value r-value p-value
PT (seconds) -0.089 0.122 -0.045 0.433 0.025 0.666
APTT(seconds) -0.064 0.272 0.071 0.223 0.085 0.142
INR -0.203 0.100 0.025 0.665 0.065 0.258
Platelets showed no correlation with any of the parameter.

63
CORRELATION PLOTS:

Figure 10 : Correlation between Plasma platelet leve l and activated partial
thromboplatin time, first trimester of pregnancy. There was no correlation
between both parameters (Pearson’s correlation coefficient, r=0.05;
p>0.05).

64

Figure 11 : Correlation between Plasma platelet level and activated partia l
thromboplatin time second trimester of pregnancy. There was no correlation between
both parameters (Pearson’s correlation coefficient, r=0.007; p>0.05).

65

Figure 12 : Correlation between Plasma platelet level and activated partial
thromboplatin time t hird trimester of pregnancy. There was no correlation between
both parameters (Pearson’s correlation coefficient, r=0.007; p>0.05).

66
CHAPTER FIVE
5.0 DISCUSSION
This study was aimed at evaluating the clotting profile of pregnant woman in
Esan West local government area (A rural community within Edo State,
Nigeria). This research is to evaluate common ca uses of post -partum
haemorrhage which has been reported to be common.
The major findings in this s tudy are that Prothrombin time and INR of the test
subject showed a significant increase compared to those of control group, P<
0.05. However , PTTK and platelet count showed no significant change when
compare to that of the control result at P > 0.05. This observed increase in PT
and INR was su pported by Hellgren (2003) who proposed that during
pregnan cy there was increase Endogenous thrombin generations , acquired
activated protein C resistance and increase porthombin level. Another similar
study in which they also meas ured prothrombin fragments , by C erneca et al.,
(1997) has showed that the parameter showing the greatest variat ions during
pregnancy were PT, Prothombin fragments F1 + 2, the existence of a
hypercoagulable state in levels of factor F1 + 2. In contrast to the results, Lioyd
et al ., (2013) showed that prothrombin time was associated with a significant
increase in the activities of factor VII, VIII, IX and in the concentration of ions
of fibrinogen and antitrypsin.

67
Szecsi et al (2010) proposed that prothrombin time remains unchanged
throughout the three trimester of pregnancy as well as the level of coagulation
factors II, V, X XII and antithrombin and protein C.
This test results showed that the partial thromboplastin time (PTTK) showed no
significant change in the three trimesters c ompared to the control group P >
0.05. These findings agreed with earlier researchers (Obisesan et al 1998) but
disagreed with Prolonged reports from previous studies (Buseri et al 2008).
The platelet count of the pregnant women showed no significant incr ease in the
three trimesters. When compared to the control group, this is contradictory to
the study reported by Boehlen et al (2000) who reported that platelet count
decreases in normal pregnancy possibly due to increased destruction and
haemodilution wi th a maximal decrease in the third trimester.
The Rbc morphology of the test group showed 70%, normocytic normochromic,
while 30% of the test group showed microcytic hypochromic blood film. This
implies that 30% of the pregnant women had iron deficiency a nemia. The
anaemia could be attributed to fetal demand for iron during pregnancy and poor
iron intake among the women in these rural settings.

68
5.1 CONCLUSION

Prothrombin time (PT) and International normalized ratio (INR) increases
progressivel y as the age of pregnancy progresses while activated partial
thromboplastin time( APTT ) and platelet count (PLT) are stable throughout
pregnancy. Red blood cell morphology showed 30% microcytic hypochromic,
while 70 % were normocytic normochromic.
5.2 RECOM MENDATION
Therefore, pregnant women are advised to take iron supplements as well as do a
routine clotting profile check -up throughout pregnancy to avoid disseminated
intravascular coagulopathy and other complications associated with clotting
profile disord ers in pregnancy.

.

69
REFERENCES
Ahmad, S. S., Rawala -Sheik h, R. and Walsh, P. N. (1992). Components and
assembly of the factor X activating complex . Semin. Thromb. Hemost. 18
(3): 311 -323.
Anne Stiene -Martin , E., Cheryl, A. , Lotspeich -Steininger, and John, A. K.
(1998). Clinical Haematology, principle, procedure andcorrelation. 2nd
ed. Philadelphia : Lippincott – Raven. Pp. 302.
Baerenwaldt, A., Biburger, M. and Nimmerjahn, F. (2010). Mechanisms of
action of intravenous immunoglobulins. Expert Rev. Clin . Immunol. 6(3):
425-434.
Barton, J.R. and Sibai, B.M. (2004). Diagnosis and management of hemolysis,
elevated liver enzymes, and low platelets syndrome. Clin Perinatol .
31(4): 807-833.
Beale, L. S. (1864). On the Germinal Matter of the Blood, with Remar ks upon
the Formation of Fibrin .Trans.Micro. Society & J. 12: 47-51.
Bernstein, I.M., Ziegler, W. and Badger, G.J. (2001) . Plasma volume expansion
in early pregnancy. J. Obstet. Gynecol . 97:669-678.
Bizzozero, J. (1882). ExtquotedblÜbereinenneuenForrnbestand teil des Blutes
und dessenRollebei der Thrombose und Blutgerinnung‖. Arch. Pathol.
Anat. Phys. Klin. Med. 3: 261-332.
Bjorksten, B., Soderstrom, T., Damber, M.G., von Schoultz, B. and Stigbrand,
T. (1978). Polymorphonuclear leucocyte function during pregnan cy.
Scand J. Immunol. 5: 257–262.
Bockenstedt, P.L. (2011). Thrombocytopenia in pregnancy. Hematol. Oncol.
Clin. North Am . 24: 293-310.

70
Boehlen , F., Hohfeld , P. And Extermann , P. (2000). Platelet count at
termpregnancy: a reappraisal of thethreshold. Obste r and Gynecol . 95:
29–33.
Bouchard, B. A., Mann, K.G. and Butenas, S. (2010). No evidence for tissue
factor on platelets . Blood. 4(5): 854 -855.
Burrow s, R.F. and Kelton, J.G. (1993) . Pregnancy in patients with idiopathic
thrombocytopenic purpura: assessin g the risks for the infant at delivery.
Obstet. Gynecol. Surv . 6(12): 781-788.
Burrows, R.F. and Kelton, J.G. (1988). Incidentally detected thrombocytopenia
in healthy mothers and their infants. N. Engl. J. Med. 5(3):142 -145.
Burrows, R.F. and Kelton, J.G. (1993).Fetal thrombocytopenia and its relation
to maternal thrombocytopenia. N. EngI. J. Med . (20):1463 -1466.
Burrows, R.F. and Kelton, J.G. (1993).Fetal thrombocytopenia and its relation
to maternal thrombocytopenia. N. EngI. J. Med . (20):1463 -1466.
Campbe ll, L.A. and Klocke, R.A. (2001). Implications for the pregnant
patient‖. Am. J. of Respir. and Critl. Care Med . 163 (5): 1051 –154.
Cerneca, F. , Ricci , G., Simeone, R., et al., (1997). Coagulation and fibrinolysis
changes in normal pregnancy. Increased leve ls of procoagulants and reduced
levels of inhibitors during pregnancy induce a hypercoagulable state, combined
with a reactive fibrinolysis. Eur. J. Obstet Gynecol Reprod Biol. 73(1): 31-36.
Cheesbrough, M. (2006). White blood cells and platelets count. District
Laboratory Practice in Tropical Countries; Part 2.2nd ed. Cambridge
University Press New York. 434.

71
Clark, P., Brennard, J., Conkie, J.A., et al (1998). Activated protein C
sensitivity, protein C, protein S and coagulation in normal pregnancy.
Thro mb. Hemost. 79: 1166 –1170.
Cooper, N. (2009). Intravenous immunoglobulin and anti -RhD therapy in the
management of immune thrombocytopenia. Hematol. Oncol. Clin. North
Am. 23(6):1317 -1327.
Crocker, I.P., Baker, P.N and Fletcher, J. (2000).Neutrophil funct ion in
pregnancy and rheumatoid arthritis. Ann. Rheumat. Dis . 59: 555–564.
Dashe , J.S., Ramin, S.M. and Cunningham, F.G. (1998). The long -term
consequences of thrombotic microangiopathy (thrombotic
thrombocytopenic purpura and hemolytic uremic syndrome) in
pregnancy. Obstet. Gynecol. 91(5): 662-668.
De Boer, K ., ten Cate, J.W., Sturk, A., Borm, J.J. and Treffers, P.E. (1989).
Enhanced thrombin genera tion in normal and hypertensive pregnancy.
Am. J. Obstet. Gynecol. 160(1):95 –100.
Durotoye, I.A., Babatunde, A .S., Olawumi, H.O., Olatunji, P.O. and Adewuyi,
J.O. (2012). Haemostatic parameters during pregnancy in Ilorin, Nigeria.
Tropical J. Health Sci . 19(2): 18 -23.
Edlestam, G., Lowbeer, C., Kral, G., et al., (2001). New reference values for
routine blood sampl es and human neutrophiliclipocalin during third
trimester pregnancy. Scand J. Clin. Lab. Inv . 61:583–592.
Eichinger, S. (2004). D-dimer testing in pregnancy. Pathophysiol. Hemost.
Thromb. 33: 327–329.
Fakhouri, F., Jablonski, M., Lepercq, J., Blouin, J., B enachi, A., Hourmant, M.,
Pirson, Y., Dürrbach, A., Grünfeld, J.P., Knebelmann, B. and Frémeaux –

72
Bacch, V. (2008). Factor H, membrane cofactor protein, and factor I
mutations in patients with hemolysis, elevated liver enzymes, and low
platelet count syndro me. Blood . 112(12): 4542 -4545.
Fay, R.A., Hughes, A.O. and Farron, N.T. (1983). Platelets in pregnancy:
hyperdestruction in pregnancy. Obstet. Gynecol . 61(2): 238-240.
Firki, F. C., Chesterma, C. N., Peningto, D.G. and Rus, B.M. (2002). DeGruchy’s
Clinical Haematology inMedical Practice . 5th ed. London:Blackwell
Publishing Ltd. 1425.
Fleming, A.F. (1975). Hematological changes in pregnancy. Clin. Obstet.
Gynecol. 2: 269-276.
Furie, B. and Furie, B. C. (2008).―Mechanisms of thrombus formation‖. N.
EngI. J. Me d. 359(9): 938 -949.
Garty, B.Z., Ludomirsky, A., Danon, Y.L., Peter, J.B. and Douglas, S.D.
(1994). Placental transfer of immunoglobulin G subclasses. Clin. Diagn.
Lab. Immunol . 1(6): 667-669.
Gatti, L., Tinconi, P.M., Guarneri, D., Bertuijessi, C., Ossola , M.W., Bosco, P.
and Gianotti, G. (1994). Hemostatic parameters and platelet activation by
flow-cytometry in normal pregnancy: a longitudinal study . Internat. J.
Clin. Lab. Res. 24(4): 217–219.
George, J.N., Woolf, S.H., Raskob, G.E., Wasser, J.S., Aledor t, L.M., Ballem,
P.J., Blanchette, V.S., Bussel, J.B., Cines, D.B., Kelton, J.G., Lichtin,
A.E., McMillan, R., Okerbloom, J.A., Regan, D.H. and Warrier, I.
(1996). Idiopathic thrombocytopenic purpura: practice guideline
developed by explicit methods for t he American Society of Hematology.
Blood. 88(1): 3-40.

73
Haeger, M., Unander, M., Norder – Hansson, B., Tylman, M. and Bengtsson, A.
(1992). Complement, neutrophil, and macrophage activation in women
with severe preeclampsia and the syndrome of hemolysis, e levated liver
enzymes, and low platelet count. Obstet. Gynecol . 79(1):19 -26.
Haram, K., Svendsen, E. and Abildgaard, U. (2009). The HELLP syndrome:
clinical issues and management. A Review. BMC Pregnancy Childbirth . 9:8-14.
Harker, L. A., Roskos, L. K., Ma rzec, U. M., Carter, R. A., Cherry, J. K.,
Sundell, B., Cheung, E. N., Terry, D. and Sheridan, W. (2000). ―Effects
of megakaryocyte growth and development factor on platelet production,
platelet life span, and platelet function in healthy human volunteers‖ .
Blood. 95(8): 2514 -2522.
Harrison, K.A. (1996). Blood volume changes in normal pregnant Nigerian
Women. The J. of obstet. and gynaecol. of the Br. Commonwealth. 73(5):
717–723.
Hellgre, M. (2003) . Haemostasis during normal pregn ancy and puerperium.
Throm bosis and Haemostasis . 29: 125 -130.
Hoffbrand, A. and Pettit , V. (2003). Essential haematology fourth edition page.
319-320
Ibdah, J.A., Bennett, M.J., Rinaldo, P., Zhao, Y., Gibson, B., Sims, H.F. and
Strauss, A.W. (1999). A fetal fatty -acid oxidation di sorder as a cause of
liver disease in pregnant women. N. Engl. J. Med. 340(22): 1723 -1731.
Jain, N. C. (1975). ―A scanning electron microscopic study of platelets of
certain animal species‖. Thrombosis etdiathesis haemorrhagica. 33(3):
501-507.

74
Jessica, M., Badger, F., Hseih, .C.C., Troisi, R., Lagiou, P. and Polischman, N.
(2007). Plasma volume expansion in pregnancy: implications for
biomarkers in population studies. Cancer Epidemiol.Biomarkers. 16:
1720 -1729.
Karalis, L., Nadan, S. and Yemen, E.A. (2005). Platelet activation in pregnancy
induced hypertension. Thromb. Res . 116(5): 377–383.
Kelton, J.G. (2002). Idiopathic thrombocytopenic purpura complicating
pregnancy. Blood Rev . 16(1): 43-46.
Kirkpatrick, C.A. (2010 ). The HELLP syndrome. Acta. Clin. Belg . 65(2): 91-97.
Kline, A.J., Williams, G.W. and Hernandez -Nino, J. (2005). D -Dimer
concentration in normal pregnancy: new diagnostic thresholds are
needed . Clin. Chem. 51(5): 825–829.
Konijnenberg, A., Stokkers, E. and van der Post, J. (1997). Extensive platelet
activation in preeclampsia compared with normal pregnancy: enhanced
expression of cell adhesion molecules. Am. J. Obstet. Gynecol . 176(2):
461–469.
Laki, K. (1972). ―Our ancient heritage in blood clotting and some of its
consequences.‖ Annals New York Aca. Sci. 202: 297 -307.
Lancet.(1882). Notes of Gulliver‘s Researches in Anatomy, Physiology,
Pathology, and Botany, 1880; Carpenter‘s Physiology.9th ed. Power. 916.
Lee, N.M. and Brady, C.W. (2009). Liver disease in pregnancy. World J.
Gastroenterol . 15(8): 897-906.
Lloyd, R ., Whitefield, Amol , S., Lele, Gerhard Levy Amherst , et al. ,(2013).
Effect of pregnancy on the relationship between concentration and

75
anticoagulant action of heparin. Clinical pharmacology and therapeutics, 34:
23-28.
Machlus, K.R., Thon, J.N. and Italiano, J. E. (2014). Interpreting the
developmental dance of the megakaryocyte: A review of the cellular and
molecular process es mediating platelet formation . Br. J. Haematol.
165(2): 227 -236.
Mason, K. D., Carpinelli, M. R ., Fletcher, J. I., Collinge, J. E.,Hilton, A. A.,
Ellis, S., Kelly, P. N. and Ekert, P. G. (2007). Programmedanuclear cell
death delimits platelet life span . Cell. 128(6): 1173 -1186.
Michel, M., Novoa, M.V. and Bussel, J.B. (2003). Intravenous anti -D as a
treatment for immune thrombocytopenic purpura (ITP) during pregnancy.
Br.J. Haematol. 123(1): 142-146.
Moise, K.J . Jr. (1991). Autoimmune thrombocytopenic purpura in pregnancy.
Clin. Obstet. Gynecol . 34(1): 51-63.
Obisesan , K. A., Adeyemo , A. A. Okunade , M. A. (1998). Haematological
values in pregnancy in Ibadan, Nigeria . Afr J Med sci, 27: 9-11.
Perepu, U. and Rosenstein, L. (2013). Maternal thrombocytopenia in pregnancy.
Proc . in Obstetr. and Gynecol . 3(1): 6-21.
Ramsay, M. (2010). Normal h aematologica l changes during pregnancy and the
puerperium. Pavord, S. and Hunt, B. ed. The obstetric h aematology
manual, Cambridge University Press, Cambridge 1 –11.
Rodeghiero, F., Castaman, G. and Dini, E. (1987). Epidemiological
investigation of the prevalence of vo n Willebrand's disease. Blood . 69:
454-459.

76
Ross, D. W. Ayscue, L. H., Watson, J. and Bentley, S. A. (1988).―Stability of
hematologic parameters in healthy subjects. Intraindividual versus
interindividual variation‖. Am. J. Clin Path. 90(3): 262 -267.
Schul tze, M. (1865). ―Einheizbarer Objecttisch und seine Verwendungbei
Untersuchungen des Blutes‖. Arch. Mikrosc. Anat. 1(1): 1 -42.
Schwartz, R.S. (2007). Immune thrombocytopenic purpura —from agony to
agonist. N. Engl. J. Med. 357(22): 2299 -2301.
Segal, J.B. an d Powe, N.R. (2006). Prevalence of immune thrombocytopenia:
analyses of administrative data. J. Thromb. Haemost . 4(11): 2377 -383.
Shehlata, N., Burrows, R.F. and Kelton, J.G. (1999).Gestational
thrombocytopenia. Clin. Obstet. Gynecol . 42: 327–334.
Smith, G .F. (1993). ―An investigation into some of the effects of the state of
nutrition of the mother during pregnancy and labour on the condition of
the child at birth and for first few days of life,‖ Nutri. 9(4): 388–392.
Stacey, T., Thompson, J.M., Mitchell, E .A., Ekeroma, A.J., Zuccollo, J.M. and
McCowan, L.M. (2011). ―Association between maternal sleep practices
and risk of late stillbirth: a case -control study‖. BMJ (Clinical research
ed.) 342.
Stavrou, E. and McCrae, K.R. (2009). Immune thrombocytopenia in pregnancy.
Hematol Oncol. Clin. North Am . 23(6): 1299 -1316.
Szecsi P. B. and Jorgensen, M. (2010). Haemostatic reference intervals in
pregnancy, thromb. Haemost. 718 -790.
Taylor, D.J. and Lind, T. (1979). Red cell mass during and after normal
pregnancy. Br. J. Obstet. Gynecol . 86: 364–370.

77
The American College of Obstetricians and Gynecologists (2002).―ACOG
Practice Bulletin: Clinical Management Guidelines for Obstetrcian
Gynecologists: Perinatal care at the threshold of viability‖. Obstet.
Gynecol . 100(3): 617–624.
Tsai, H.M. and Lian, E.C. (1998). Antibodies to von Willebrand factor cleaving
protease in acute thrombotic thrombocytopenic purpura. N. Engl. J. Med .
339(22): 1585 -1594.
Tsunoda, T., Ohkuchi, A., Izumi, A., Watanabe, T., Matsubara, S., Sato, I. and
Minakami, H. (2002). Antithrombin III activity and platelet count are
more likely to decrease in twin pregnancies than in singleton pregnancies.
Acta. Obstet. Gynecol. Scand. 81(9): 840-845.
Uchikova, E.H. and Ledjev, I.I. (2005). Changes in haemost asis during normal
pregnancy. Eur. J. Ghnaecol. Reprod. Biol . 199(2): 185 -188.
Usha, P. and Lori, R. (2013). Maternal thrombocytopenia in pregnancy.
Proceedings in Obstet. and Gynecol. 3(1): 6 -21.
Van Veen , J.J, Nokes, T.J. and Makris, M. (2010). The risk of spinal
haematoma following neuraxial anaesthesia or lumbar puncture in
thrombocytopenic individuals. Br. J. Haematol . 148(1):15 -25.
Verdy, E., Bessous, V., Dreyfus, M., Kaplan, C., Tchernia, G. and Uzan, S.
(1997). Longitudinal analysis of platelet cou nt and volume in normal
pregnancy. Thromb. Haemost . 77(4): 806-807.
WHO. (2004).―Prevention and treatment of malaria during pregnancy,‖
http://pdf.usaid.gov/pdf docs/Pnada621.pdf .

78
Walker, I.D., Wal ker, J.J ., Colvin, B.T., Letsky, E.A., Rivers, R. and Stevens,
R. (1994). Investigation and management of haemorrhagic disorders in
pregnancy. J. Clin. Pathol . 47:100-108.
Win, N., Rowley, M., Pollard, C., Beard, J., Hambley, H. and Booker, M.
(2005). Seve re gestational (incidental) thrombocytopenia: to treat or not
to treat. Hematol . 10(1): 69-72.
Wright, J. H. (1906). ―The Origin and Nature of the Blood Plates‖. The Boston
Med. Surg. J. 154 (23): 643.
Wright, J. H. (1910). ―The histogenesis of blood plate lets‖. J. Morphol. 21: 263 –
278.
Yip, R. (2000). ―Significance of an abnormally low or high h aemoglobin
concentration during pregnancy: special consideration of iron nutrition,‖
The Am. J. of Clin. Nutri. 721: 272–279.