Clinical Practice 4

(CP4)




HAEMATOLOGY HAEMATOLOGY HAEMATOLOGY HAEMATOLOGY IN IN IN IN
CLINICAL PRACTICE CLINICAL PRACTICE CLINICAL PRACTICE CLINICAL PRACTICE

Course Handbook and
Guidelines for Undergraduates



Author: Dr C Dainty
Basic Science Teacher/
Undergraduate Medical Education
Postgraduate Tutor




CONTENT



1 Introduction Page



Haematology Handbook.

This handbook has been designed to give you a basic understanding of science
needed to understand haematological investigations and disease
It is intended to cover the essentials to enable you to formulate further objectives
and reading as appropriate. It is not intended to replace other texts and should be
used alongside your clinical attachments.


Learning Outcomes.

1. List main constituents of blood and their functions.
2. Name conditions that shift oxygen dissociation curve to right and left
3. Compare differences between foetal and adult haemoglobin
4. To explain significance of differential WBC
5. Compare difference between primary and secondary haemostasis
6. List factors needed for coagulation process
7. Classify factors needed for intrinsic, extrinsic and final common pathway.
8. Recognise investigations needed to assess coagulation pathways
9. Name fibrin degradation products
10. List functions spleen and bone marrow.
11. Define and classify different anaemias
12. Read a routine blood count
13. Comment on reticulocyte count
14. Define MCV and discuss its diagnostic and patho physiologic relevance



















Blood

Blood transports life-supporting food and oxygen to every cell of the body and
removes their waste products. It also helps to maintain body temperature,
transports hormones, and fights infections. The brain cells in particular are very
dependent on a constant supply of oxygen. If the circulation to the brain is stopped,
death shortly follows.
Blood has two main constituents. The cells, or corpuscles, comprise about 45
percent, and the liquid portion, or plasma, in which the cells are suspended
comprises 55 percent. Blood temp is approx 38 degrees C, 5X viscous as water,
slightly alkaline Ph av 7.4. An adult male has approx 5-6 L of whole blood(woman
4-5L)
The blood cells comprise three main types:
1. Red blood cells, or erythrocytes;
2. White blood cells, or leukocytes, which in turn are of many different types;
3. and platelets, or thrombocytes.
Each type of cell has its own individual functions in the body.
The plasma is a complex colourless solution, about 90 percent water, that
carries different ions and molecules including proteins, enzymes, hormones,
nutrients, waste materials such as urea, and fibrinogen, the protein that aids in
clotting.


Red Blood Cells
The red blood cells are tiny, round, biconcave disks, averaging about 7.5 microns
(0.003 in) in diameter A normal-sized man has about 5 litres (5.3 qt) of blood in his
body, containing more than 25 trillion red cells. Because the normal life span of red
cells in

the circulation is only about 120 days; more than 200 billion cells are normally
destroyed each day by the spleen and must be replaced. Red blood cells, as well
as most white cells and platelets, are made by the bone marrow by process of
erythopoiesis.
The main function of the red blood cells is to transport oxygen from the lungs to the
tissues and to transport carbon dioxide, one of the chief waste products, it to the
lungs for release from the body.
The substance in the red blood cells that is largely responsible for their ability to
carry oxygen and carbon dioxide is haemoglobin, the material that gives the cells
their red colour. It is a protein complex comprising many linked amino acids, and
occupies almost the entire volume of a red blood cell. Essential to its structure and
function is the mineral iron. It consists of two parts, haem and globin.
Haem is a complex of a porphyrin ring and iron in the ferrous state. Haem is
conjugated with globin a polypeptide
The blood cell type responsible for the transport of oxygen and carbon
dioxide is the ________.
platelet
lymphocyte
white blood cell
red blood cell

A. The blood cell type responsible for the transport of oxygen and carbon dioxide is
the red blood cell (erythrocytes.)

Haemoglobin and transport of Oxygen

The amount of oxygen the blood carries is described as the oxygen content of the
blood. Although vast amount of oxygen is carried bound to haemoglobin, a very
small amount is dissolved in the plasma the total amount of oxygen carried by
blood is given by adding these two amounts together.

Under normal circumstances as blood leaves the lungs Hb is almost fully saturated
with oxygen and each g of Hb carries 1.39mls of oxygen. As it reaches systemic
circulation this falls because of addition of venous blood from lungs and heart.
Therefore in arterial blood 1g of Hb is 98% saturated and contains 1.34mls of O2.
The amount dissolved is proportional to partial pressure.
At 37 Celsius 0.23 mls oxygen dissolves in each L of blood per kPa.

Oxygen content t of blood calculated:

O2 content = (Hb x 1.34 x SaO2) + (0.23 x Pa O2)

= (150 x 1.34x 98%) + (0.23 x 13) = 20.3 ml/l


Oxygen combines reversibly with ferrous ion in Hb molecule to form oxy
haemoglobin.
Hb + O2 >< HbO2

The no of haem units bound with O2 is called Hb saturation If fully loaded it is
100%.
In lungs where partial pressure of O2 is high. Oxygenation of Hb is favoured. In
tissues where PP is low. Reverse happens.
Thus in tissues O2 are released. These are rapid reactions taking less than 0.01
seconds.















This relationship between between saturation of Hb and PP of O2 is;

Oxygen dissociation curve






The curve demonstrates that at a normal Pa O2 (13kPa) Hb is approaching 100%
sat.
Te curve has a sigmoid shape so at low levels of oxygen tension it is difficult for Hb
to combine withO2. Steep section of curve is important as it demonstrates that
below 92% saturation the drop in saturated Hb falls very quickly.

Other factors that effect Curve
Many factors influence the affinity of this binding and alter the shape of the curve:
right shift left shift
temperature high low
DPG high low
p(CO
2
) high low
p(CO) low high
pH (Bohr effect) low (acidosis) high (alkalosis)
type of haemoglobin adult haemoglobin fetal haemoglobin
Left shift of the curve is a sign of hemoglobin's increased affinity to take up oxygen
(eg. at the lungs). Similarly, right shift shows decreased affinity ie releases its
oxygen more easily, as would appear with an increase in body temperature,
hydrogen ion(acidosis), 2,3-diphosphoglycerate or carbon dioxide concentration
(the Bohr effect)De-oxy haemaglobin binds more easily with hydrogen ions than
oxyHb thus as H ions rise, Ph falls Hb affinity for O2 falls
2,3diphosphoglycerate is formed in red blood cells as a product of glycolysis. It is a
highly charged ion. It combines with Hb it displaces O2 and shifts curve to right.
Thus conditions that cause a right shift by encouraging unloading of O2 at
relatively low PP of O2 can be thought of as an exercising muscle.
During exercise- Temp increases/ H ions increase/Co2 and DPG increase = more
O2 to muscles

Carbon monoxide has a much higher affinity for haemoglobin than oxygen does. In
carbon monoxide poisoning, oxygen cannot be transported and released to body
tissues thus resulting in hypoxia.
Fetal Hb is structurally different from adult. This causes it to bind less with DPG.
DPG reduces the amount of O2 that can combine so fetal Hb has a higher affinity
for O2 than adult. This enables fetus to extract O2from maternal blood at placenta.
The oxygen dissociation curve for myoglobin exists even further to the left.

Carbon Dioxide transport


After entering blood CO2 is converted to carbonic acid and bound to Hb in
RBCs(%) and some dissolved in plasma

CO2 + H2O <> H+ + HCO3-

Reversible reaction. Proceeds vigorously in peripheral capillaries tying up large
numbers of CO2 molecules.

H ions bind to HB = carboxy Hb
HCO3 moves into plasma which exchanges another anion CL-. Resulting in
chloride ions moving into RBCs = Chloride shift.
Plasma becomes saturated rapidly and carries only 7% of absorbed CO2






Blood tests and RBCs


Haematocrit(Hct) % of formed elements in whole blood

High= polycythaemia
Norm = 37-54 Low= anaemia

Reticuloctye count % retics High=reticulocytosis
Normal = 0.8% Low=anaemia

Haemaglobin(Hb) Conc of Hb in blood
Normal=12-18g/dl Low-anaemia



RBC count no of RBCs per mu/l blood High-polycythaemia
Normal= 4.2- 6.3 million Low= anaemia



Mean Corpuscular av volume of RBC High= macrocytic
Volume(MCV) Normal= 82-101 Low=microcytic



Mean corpuscular Av amount Hb in RBC High= hyperchromic
Normal=27-34 Low=hypochromic
Haemaglobin(MCHC)



Q. How would haematocrit change after a large haemorrhage?


White Blood Cells
The leukocytes, or white blood cells, are of three types; granulocytes,
lymphocytes, and monocytes. All are involved in defending the body against
foreign organisms.
There are three types of granulocytes: neutrophils, eosinophils, and basophils, with
neutrophils the most abundant. Neutrophils seek out bacteria and phagocytise, or
engulf, them.
The lymphocytes' chief function is to migrate into the connective tissue and build
antibodies against bacteria and viruses.
White blood cells are almost colourless, considerably larger than red cells, have a
nucleus, and are much less numerous; only one or two exist for every 1,000 red
cells. The number increases in the presence of infection.
Monocytes, representing only 4 to 8 percent of white cells, attack organisms not
destroyed by granulocytes and leukocytes.
The granulocytes, accounting for about 70 percent of all white blood cells, are
formed in the bone marrow. The lymphocytes on the other hand are produced
primarily by the lymphoid tissues of the bodythe spleen and lymph nodes. They
are usually smaller than the granulocytes. Monocytes are believed to originate
from lymphocytes. Just as the oxygen-carrying function of red cells is necessary
for our survival, so are normal numbers of leukocytes, which protect us against
infection.
Fighting infection and foreign invaders is one of the primary functions of the
________.
erythrocytes (red blood cells)
leukocytes (white blood cells)
thrombocytes (platelets)
all of the above

Fighting infection and foreign invaders is one of the primary functions of the leucocytes.
























Routine Hematological Tests


Differential WBC Count
There are five different kinds of
WBCs
1.Eosinophils
1. Basophils
2. Neutrophils
3. Lymphocytes
4. Monocytes






Normal Values (Male or Female)
Neutrophils 40 - 75%
Eosinophils 1 - 4%
Basophils 0 - 1%
Lymphocytes 20 - 45%
Monocytes 2 - 8%


Significance Of The WBC Count

When there is an increase in eosinophils it is known as
eosinophilia. It is observed in chronic inflammatory
conditions, asthma, parasitic infestations, and in
hypersensitivity reactions.

When there is a decrease in lymphocytes, it is known as
lymphopenia. It is observed in acute stages of infections
and where there is an excessive irradiation.

When there is an increase in neutrophils, it
is known as neutrophilia. The common
cause is pyogenic (pus forming) bacterial
infections. When there is a decrease in
neutrophils it is known as neutropenia. It is
observed in bacterial infections such as
typhoid, viral infection such as measles, influenza etc. It is
also found in anaemia (aplastic, megaloblastic, iron
deficiency) and in suppression of bone marrow by various
drugs and radiation.

When there is an increase in lymphocytes it is known as
lymphocytosis. Lymphocytosis can be of two types,
relative or absolute.
Relative Lymphocytosis: In this, the actual no of lymphocytes
has not changed, but due to a decrease in neutrophils,
the differential count shows an increase in lymphocytes.
Absolute Lymphocytosis:
It is seen in:
Children.
Also seen when there are infections such
as tuberculosis, typhoid, mumps, measles,
cough, influenza syphilis and other chronic infections.
Infectious mononucleosis.
Chronic lymphatic leukemia.
When there is an increase in the number of monocytes, it is known
as monocytosis. It is observed in tuberculosis, malaria, sub-acute
bacterial endocarditis, typhoid
and in Kala Azar.
A differential count is useful in identifying changes in the distribution of
WBCs, which may be related to specific types of disorders. It also helps to know
the severity of the disease and the degree of the response of the body
Platelet Count
Platelets are very small in diameter - around 3 mm. They help in clotting of
blood.
said to
be adequate. If it is less than that, it is said to be inadequate.
Abnormalities Of Erythrocytes (RBC)
In various types of anaemia and in other diseases such as
thalassemia,
malaria, kidney failure etc, the mature RBCs show significant
changes.
Various changed RBCs seen are termed as, Microcytes,
Macrocytes,
Hypochromic, Spherocytes, Target
cells, Stomatocytes,
Anisocytosis, Poilkilocytosis, sickle
cells, Ovalocytes, Elliptocytes,
Acanthocytes, Burr cells, Siderocytes, Basophilic
stippling, Howell-Jolly Body, Cobot ring, Schi
stocytes, Crescent bodies and
Creneted cells depending upon their morphology.

Abnormalities Of Leucocytes (WBC)
Most abnormalities are seen in Neutrophils.

Different WBCs having abnormalities are termed as Myelocytes,
Promyelocytes,
Blasts, Metamyelocytes, Plasma Cell, Smudge cells etc.




Platelets
Platelets, or thrombocytes, are much smaller than the red blood cells. They are
round or biconcave disks and are normally about 30 to 40 times more numerous
than the white blood cells. The platelets' primary function is to stop bleeding. When
tissue is damaged, the platelets aggregate in clumps to obstruct blood flow.

Blood platelets, or thrombocytes, are not true cells, but rather cytoplasmic
fragments of a large cell in the bone marrow, the megakaryocyte. The central
portion of a platelet stains purple with Wright's stain and is referred to as the
granulomere. The peripheral portion stains clear and is called the hyalomere.
Platelet contents include glycogen granules, the open canalicular system (OCS),
which is composed of canaliculi formed from invaginations of the platelet plasma
membrane, mitochondria, occasional Golgi elements and ribosomes. Platelets
have several types of membrane-bound granules which contain a number of
constituents including fibrinogen and several growth factors (e.g., PDGF).
Platelet activation occurs when injury to the vessel wall exposes sub-endothelial
components, especially collagen. Platelets adhere to the damaged area and
become cohesive to other platelets. This aggregation leads to the formation of a
platelet plug, which prevents further blood loss and allows the repair process to
begin. The micrograph shown below shows activated platelets adhering to some
damaged cells.

Severe reduction in the number of circulating platelets results in a condition known
as thrombocytopenia.
It is a condition which causes spontaneous bleeding as a reaction to minor trauma.
This is due to failure of the platelets to seal over microscopic breaches in blood
vessel walls.
In the skin this is manifest by a reddish-purple blotchy rash. This can be either
small blotch called purpura or larger bruise like areas called ecchymoses.
Cytotoxic drugs used in treatment of cancers may cause this condition. It is also
seen associated with acute leukaemias.
Haemostasis is a complex process which changes blood from a fluid to a solid
state. Intact blood vessels are central to moderating blood's tendency to clot. The
endothelial cells of intact vessels prevent thrombus formation by secreting tissue
plasminogen activator (t-PA) and by inactivating thrombin and adenosine
diphosphate (ADP). Injury to vessels overwhelms these protective mechanisms
and haemostasis ensues. Haemostasis proceeds in two phases: primary and
secondary haemostasis.


Primary haemostasis is characterized by vascular contraction, platelet
adhesion and formation of a soft aggregate plug. It begins immediately after
endothelial disruption. Injury causes temporary local contraction of vascular
smooth muscle. Vasoconstriction slows blood flow, enhancing platelet
adhesion and activation.
o Adhesion occurs when circulating von Willebrand factor(vWf)
attaches to the subendothelium. Next, glycoproteins on the platelet
surface adhere to the "sticky" von Willebrand factor(vWf). Platelets
collect across the injured surface. These platelets are then
"activated" by contact with collagen. Collagen-activated platelets form
pseudopods which stretch out to cover the injured surface and bridge
exposed fibers. The collagen-activated platelet membranes expose
receptors which bind circulating fibrinogen to their surfaces.
Fibrinogen has many platelet binding sites. An aggregation of
platelets and fibrinogen build up to form a soft plug. Platelet
aggregation occurs about 20 seconds after injury.
o Primary haemostasis is short lived. The immediate post injury
vascular constriction abates quickly. If flow is allowed to increase, the
soft plug could be sheared from the injured surface, possibly creating
emboli.
Secondary haemostasis is responsible for stabilizing the soft clot and
maintaining vasoconstriction. Vasoconstriction is maintained by platelet
secretion of serotonin, prostaglandin and thromboxane. The soft plug is
solidified through a complex interaction between platelet membrane,
enzymes, and coagulation factors.
o Coagulation factors are produced by the liver and circulate in an
inactive form until the coagulation cascade is initiated. The cascade
occurs in steps. The completion of each step activates another
coagulation factor in a chain reaction which leads to the conversion
of fibrinogen to fibrin.

The Blood Coagulation Process

Blood coagulation is a process that changes circulating substances within the
blood into an insoluble gel. The gel plugs leaks in blood vessels and stops the loss
of blood. The process requires coagulation factors, calcium and phospholipids.
The coagulation factors (proteins) are manufactured by the liver.
Ionized calcium ( Ca++ ) is available in the blood and from intracellular
sources.
Phospholipids are prominent components of cellular and platelet
membranes. They provide a surface upon which the chemical reactions of
coagulation can take place.

Coagulation can be initiated by either of two distinct pathways.
The Intrinsic pathway can be initiated by events that take place within the
lumen of blood vessels. The Intrinsic pathway requires only elements
(clotting factors, Ca++, platelet surface etc.) found within, or intrinsic to the
vascular system.
The Extrinsic pathway is the other route to coagulation. It requires Tissue
Factor (tissue thromboplastin), a substance which is "extrinsic to", or not
normally circulating in the vessel. Tissue Factor is released when the vessel
wall is ruptured.

Regardless of whether the Extrinsic or Intrinsic pathway starts coagulation,
completion of the process follows a common pathway. The common pathway
involves the activation of factors: X, V, II, XIII and I. Both pathways are required for
normal hemostasis and there are positive feedback loops between the two
pathways that amplify reactions to produce enough fibrin to form a lifesaving plug.
Deficiencies or abnormalities in any one factor can slow the overall process,
increasing the risk of haemorrhage.




The coagulation factors are numbered in the order of their discovery. There are 13
numerals but only 12 factors. Factor VI was subsequently found to be part of
another factor. The following are coagulation factors and their common names:
Factor I - fibrinogen
Factor II - prothrombin
Factor III - tissue thromboplastin (tissue factor)
Factor IV - ionized calcium ( Ca++ )
Factor V - labile factor or proaccelerin
Factor VI - unassigned
Factor VII - stable factor or proconvertin
Factor VIII - antihemophilic factor
Factor IX - plasma thromboplastin component, Christmas factor
Factor X - Stuart-Prower factor
Factor XI - plasma thromboplastin antecedent
Factor XII - Hageman factor
Factor XIII - fibrin-stabilizing factor
The liver must be able to use Vitamin K to produce Factors II, VII, IX, and X.
Dietary vitamin K is widely available from plant and animal sources. It is also
produced by normal intestinal flora. A deficiency is rare but may occur:
in newborns because they must first develop normal flora to produce
Vitamin K, or
when the flora is disturbed by broad-spectrum antibiotics.
At birth and throughout childhood, Factor VIII levels are the same as adult values.
Many other factor levels are below adult levels at birth, some as low as 10% of
adult levels. Theses levels increase toward the adult levels by age 6 months,
although they may remain mildly below adult normal range throughout childhood.
Despite lower levels, newborns and children do not normally experience bleeding.
This confers some level of antithrombolic protection in youth. During pregnancy
Factor XI can decrease, but fibrinogen and factor VIII increase.

Instant Feedback
Of the factors below, which is not produced in the liver?
Factor IV
Factor V
Factor X
Factor VIII


Prothrombin time

The prothrombin time or PT is an assessment of the extrinsic and common
pathways. The PT is the specific and only lab test used to measure the
effectiveness of coumarin-type anticoagulant drugs, such as warfarin sodium
(Coumadin). The most common methods to report the PT are time (in seconds)
and the International Normalized Ratio (INR).
The PT reported as time in seconds, represents how long a plasma sample
takes to clot after a mixture of thromboplastin and calcium are added. If the
patient's blood has less prothrombin than the normal ( or "control"), or a
decrease in other clotting factors that affect the prothrombin time, the PT
time in seconds will be longer than the control values. Normal values for PT
time reported in seconds is between 11 and 13 seconds, depending on the
method used by the laboratory.
To give PT values a consistent basis of comparison from laboratory to
laboratory, the World Health Organization instituted the INR, a uniform value
in which PT is expressed as a ratio. In the last few years, the INR is
becoming a more common method of measuring and reporting the PT.
Targets for the INR vary, depending on the reason for anticoagulation. For
example, a patient having hip surgery, who is being anticoagulated to
prevent deep vein thrombosis, may have a target INR of between 2 and 3.
To prevent arterial thrombosis in a patient with an artificial heart valve, the
INR therapeutic goal may be in the 2.5 to 3.5 range.
As oral coumadin has a relatively long onset of action, it takes several days of
therapy to achieve a therapeutic level. When a patient will be managed on oral
coumadin, he or she may be given both heparin and coumadin, and then heparin is
discontinued. If a PT is increased beyond a therapeutic level, the coumadin dose
may be reduced, or the patient may be given parenteral Vitamin K. Drugs that
increase or prolong the PT time include antibiotics, cimetidine (Tagamet),
salicylates, and sulfonamides. Barbiturates, oral contraceptives, and Vitamin K in
multivitamin preparations or in liquid nutritional supplements decrease the PT time.
When therapy with coumadin is begun, the dose is guided by monitoring the
prothrombin time. Therapeutic levels are generally between 1 1/2 and 2 times
normal, depending on the patient's need for anticoagulation. Expressed in terms of
the INR, the range is between 2.0 and 3.0. A usual dose requirement is between 5
to 7.5 mg. daily. In some patients with coumadin resistance, the dosage may be
much higher. If a patient with a prolonged PT must have surgery, it is important
that the PT be brought within a normal range before surgery. This is often done
with Vitamin K injections. Whole blood or fresh frozen plasma should be available
for the surgical patient with an abnormal PT.


Instant Feedback:
The INR is increasingly becoming an important way to measure and report
prothrombin time.


True
False

Activated Partial Thromboplastin time (aPTT)

The activated partial thromboplastin time (aPTT) is a common screening test done
to evaluate function of the intrinsic clotting system.
It has largely replaced the older PTT, which was unable to incorporate
variables in surface/contact time.
The aPTT now measures the clotting time of plasma, from the activation of
factor XII by a reagent (a negatively charged activator such as silica and a
phospholipid) through the formation of a fibrin clot.
If a patient's aPTT is abnormal, additional tests will be done to determine
the exact cause of the coagulation problem.
Reference values for aPTT vary among laboratories, but generally range between
25 and 38 seconds. The aPTT of a newborn will usually be prolonged and may be
up to 55 seconds at birth. It decreases to the adult range by 6 months of age.
(Note: each lab has its own reference values based upon the equipment and
reagents used.)
The aPTT is the most commonly used test to monitor heparin therapy. The
therapeutic goal for a patient being anticoagulated with heparin, is an aPTT
approximately 1.5 to 2.5 times the mean normal value. Heparin is most often
administered as an initial intravenous bolus followed by a continuous intravenous
infusion. The aPTT is evaluated every 6 hours during the first day of heparin
therapy and 6 hours after any dosage change. If the aPTT is therapeutic, it can be
checked once daily while patients are on heparin. (Note: If low molecular weight
heparin is given for anticoagulation, a prolonged aPTT does not occur, so another
test(s) may be indicated to monitor therapy.)
If the aPTT is increased beyond the therapeutic range, the physician may order the
heparin IV flow slowed or briefly discontinued. As the half-life of heparin is quite
short, these measures will often allow the aPTT to rapidly return to a therapeutic
range. Protamine sulfate may also be given to block the action of heparin. In some
serious situations, such as active bleeding, the physician will order a transfusion of
whole blood or plasma to increase clotting factors. Surgery may be postponed in a
patient who has an increased aPTT, unless it is an emergency procedure.
A prolonged aPTT in non-heparinised patients can occur due to:
salicylates
inherited or acquired intrinsic clotting factor deficiency or abnormality (XII,
XI, X, IX, VII, V, II, I)
massive blood replacement
hemophilia A
lupus anticoagulant
excessive coumarin dosage
A decreased aPTT can occur due to:
digitalis
tetracyclines
antihistamines
nicotine
elevated factor VIII
tissue inflammation or trauma

Instant Feedback:
The aPTT is a measurement of the intrinsic coagulation system.

True
False


Fibrinogen, and fibrin degradation products

Fibrinogen is a circulating plasma protein manufactured by the liver. Thrombin
converts fibrinogen to fibrin in the final stage of blood coagulation. Low fibrinogen
levels can occur as a result of severe liver disease or due to a disorder such as
disseminated intravascular coagulation (DIC). Fibrinogen is quantified by adding
thrombin to a series of successively more dilute plasma samples and comparing
clotting time to a control series. A coagulation analyzer is used to determine
clotting time which will be inversely proportional to the concentration of fibrinogen.
Normal values are approximately 200-400 mg/dl. Fibrinogen can also be measured
directly by immunoassay. In this test a fibrinogen antibody is added to a plasma
sample and then fibrinogen marked with the antibody is measured.


Fibrin degradation products (FDP), also known as fibrin slpit products, are present
in blood when the thrombolytic enzyme plasmin cleaves fibrin or fibrinogen.
Plasmin is produced when the thrombolytic system is activated. D-dimers are an
FDP produced when fibrin is cleaved by plasmin. The presence of D-dimers or
FDP may be used to assist with the diagnosis of DIC, Deep Venous Thrombosis
(DVT) or Pulmonary Embolism (PE). However, because they are produced under a
variety of circumstances their presence alone is not diagnostic.

Instant Feedback:
Plasmin produces D-dimers when which substance is cleaved?

Fibrinogen
Fibrin

Plasma
The plasma is more than 90 percent water and contains a large number of
substances, many essential to life. Its major solute is a mixture of proteins. The
most abundant plasma protein is albumin. The globulins are even larger protein
molecules than albumin and are of many chemical structures and functions. The
antibodies, produced by lymphocytes, are globulins and are carried throughout the
body, where many of them fight bacteria and viruses.
An important function of plasma is to transport nutrients to the tissues. Glucose,
for example, absorbed from the intestines, constitutes a major source of body
energy. Some of the plasma proteins and fats, or lipids, are also used by the
tissues for cell growth and energy. Minerals essential to body function, although
present only in trace amounts, are other important elements of the plasma. The
calcium ion, for example, is essential to the building of bone, as is phosphorus.
Calcium is also essential to the clotting of blood. Copper is another necessary
component of the plasma.
A major source of body energy, transported to the cells by the plasma, is
________.
adenosine triphosphate (ATP)
glucose
oxygen

coenzyme Q-10
A major source of body energy, transported to the cells by the plasma, is glucose.

Spleen
The spleen is an organ that is not essential for life. It can be divided into red
sinuses lined by macrophages and the white pulp which is similar in structure to
follicles.

It has 4 main functions;

1. Sequestrartion and phagocytosis of aged RBCs.
RBcs pass through red pulp and are phagocytosed.

2. Immunological.
Macrophages within spleen can remove antibody coated red cells from blood

3. Blood reservoir.
Up to 1/3 of platelets are sequestrated in spleen. In addition an enlarged spleen
can accommodate RBCs.

4. Extra-medullary haemapoiesis

Spleen is an organ of haemapoiesis for foetus. It also undertakes this role in adult
life at times of haemapoietic crises e.g. haemolytic anaemia, thallassaemia and
myelofibrosis.
A spleen only becomes palpable when it is approximately three times normal size


Hypersplenism
An imprecise term commonly used to refer to a clinical state of;
Reduced RBCs, platelets and granulocytes in any combination
An enlarged spleen from any cause
An adequate cellular bone marrow indicating thee is sufficient cells
Totally or partially correctable by splenectomy. Symptoms include abdo discomfort,
anaemia, infection and bleeding
Bone marrow (or medulla ossea) is the soft tissue found in the hollow interior
of bones. In adults, marrow in large bones produces new blood cells.
Marrow types
There are two types of bone marrow: red marrow (also known as myeloid tissue)
and yellow marrow. Red blood cells, platelets and most white blood cells arise in
red marrow; some white blood cells develop in yellow marrow. The color of yellow
marrow is due to the much higher number of fat cells. Both types of bone marrow
contain numerous blood vessels and capillaries.
At birth, all bone marrow is red. With age, more and more of it is converted to the
yellow type. Adults have on average about 2.6 kg (5.7 lb) of bone marrow, with
about half of it being red. Red marrow is found mainly in the flat bones such as hip
bone, breast bone, skull, ribs, vertebrae and shoulder blades, and in the
cancellous ("spongy") material at the proximal ends of the long bones femur and
humerus. Yellow marrow is found in the hollow interior of the middle portion of long
bones.
In cases of severe blood loss, the body can convert yellow marrow back to red
marrow in order to increase blood cell production.
Types of stem cells
Bone marrow contains two types of stem cells:
Haematopoietic stem cells give rise to the three classes of blood cell that
are found in the circulation: white blood cells (leukocytes), red blood cells
(erythrocytes), and platelets (thrombocytes).
Mesenchymal stem cells are found arrayed around the central sinus in the
bone marrow. They have the capability to differentiate into osteoblasts,
chondrocytes, myocytes, and many other types of cells. They also function
as "gatekeeper" cells of the bone marrow.


Diseases of the Blood
Anaemia
Objectives.
Students must be able to;
1.Define anaemia
2. Read a routine blood report and indicate if anaemia is present
3. Comment on reticulocyte count and indicate if result indicates likelihood of
haemolysis/ loss versus RBC underproduction
4.Define MCV and discuss its diagnostic and pathophysiological relevance
5. Classify anaemia on the basis of MCV into- normocytic/ microcytic/ macrocytic.
Appproach to the diagnosis of anaemia.
Anaemia is a deficiency of haemoglobin in the blood. It can be caused by blood
loss, abnormal destruction of the red cells (haemolysis), and inadequate red cell
formation by the bone marrow.

There are 3 important diagnostic principles:
1.FBC tells you if an anaemia is an isolated abnormality, or it is associated with
another condition.
2.Reticulocyte count tells you whether an anaemia is likely to be due to
underproduction of RBC or haemolysis/loss
3. Mean cell volume(MCV) allows you to classify anaemia by RBC size into
microcytic/ macrocytic/ normocytic.
It is size of RBC that directs your subsequent history and investigations
Anaemia is a reduction in Hb below normal level.Age and sex are important in
consideration of level
.Look at FBC.
Check WBC and platelets and differential WBC.
Abnormally high or low WBC or platelets may be a clue of an underlying blood
disease.
Pancytopenia suggests marrow replacement or hypersplenism
An erythroleukoblastic blood smear(circulating immature RBCs such as
myelocytes or nucleated RBCs) could indicate marrow replacement with
tumour,fibrosis or leukaemia.
When anaemia is only problem look at MCV.
MCV suggests pathophysiology
Mean corpuscular volume is the mean size of RBC as determined by automated
cell counter.
LOW MCV- indicates Hb synthesis is impaired from no of causes.
HIGH MCV- indicates DNA synthesis is impaired or an increase in RBC
membrane.
NORMAL MCV- means no impairment in HB or DNA synthesis.
Microcytic Anaemia- low MCV
Indictes impaired Hb synthesis
1. Iron deficiency- cause to be ascertained
2. Iron unavailabilty as in chronic diseases
3. Impaired globin chain synthesis as in thalassemia
4. impaired haem synthesis as in lead poisoning ad other rare diseases.
Macrocytic Anaemia- high MCV
Classifed as with and without megaloblastosis.
Megaloblastosis; implies impared DNA synthesis as in B12 and folic aid deficiency.
Absence of megaloblasts; implies normal DNA synthesis. There is probably
increased RBC membrane formation as in alcoholism, liver disease and
hypothyroidism.
MCV is also increased in haemolytic anaema due to an increase in reticulocyes
which are larger than erythrocytes.
Normocytic anaemia
Could be underproduction or RBC loss by bleeding or haemolysis.
Reticulocyte count will indicate which.
LOW retic count= underproduction
HIGH retic count= blood loss or haemolysis
Note; response to therapy(b12 or fe) is associated with reticulocytosis


Anaemia caused by acute or chronic blood loss, or abnormal bleeding, results from
the inability of the bone marrow to make new cells as fast as they are needed. In
acute or massive bleeding, the red blood cells and their haemoglobin are normal
but are not abundant. Chronic slow bleeding leads to a deficiency in iron stores
needed for haemoglobin. This results in smaller red blood cells that are paler than
normal.
Abnormal destruction of red cells (the haemolytic anaemias) leads to a shorter
than normal red cell survival. For example, in the hereditary disease Sickle-cell
Anaemia the haemoglobin is built erroneously. Such cells are more fragile and
break more readily in circulation.


Anaemias caused by bone-marrow failure include aplastic anaemia, in which the
bone marrow lacks adequate numbers of some or all types of blood cells. Another
anaemia caused by failure of production of red cells is pernicious anaemia. In this
disease, the person's stomach fails to produce "intrinsic factor," which is
necessary for the normal absorption of vitamin B-12 from the intestines. Because
vitamin B-12 is essential for normal bone marrow function, red cells are not formed
normally. Sublingual vitamin B-12 supplements are available, from which the
vitamin is absorbed through the blood vessels under the tongue, thus avoiding the
potential problems associated with intestinal absorption.
Pernicious anaemia results when the person's stomach fails to produce
intrinsic factor, which is necessary for the normal absorption of vitamin
________.
B-1
B-6
B-12
C

Red Blood Cells - Heads and Tails

Heads..... .....tails
There are numerous red blood cells in the
body
.....because a large amount of oxygen must be
carried from the lungs to all body cells.
Red blood cells are tiny .....because they must fit through extremely
narrow capillaries.

Red blood cells are flexible .....because they must bend and squeeze
through capillaries without being broken.
Red blood cells are packed with
haemoglobin
.....because this protein binds oxygen in the
lungs and releases it where it's needed in the
body.
Red blood cells have no nucleus .....because their job is very simple and more
haemoglobin can be packed in.
Red blood cells have a flat disc shape .....because this provides a large surface area
for oxygen to diffuse across.
Haemoglobin molecules contain iron .....because ions of this metal can bind and
release oxygen.

Red blood cells are broken down in the liver
after only about four months
..but they are continuously made by the red
marrow in certain bones.
Diagnosis of an Abnormal WBC
Given a patient with abnormal WBC, your approach should be very straightforward.
Basically you want to determine
1. Which cell type or types are abnormal? Is the abnormality reactive
(due to infection) if not is it neoplastic? The differential WBC is
essential
2. Is maturation normal? Are immature cells present? Are there any
blasts?
3. Are cell marker studies needed?
4. Are indicators of clonality needed
5. Is there an emergency problem eg hyperleukocytosis or
agranulocytosis?

Indicators of clonality.
In B cells kappa or lambda chain or immunoglobulin heavy chain rearrangents
In T cells T cell receptor gene rearrangements
Plasma cells- serum rotein electrophoresis
Granulocytes- karyotypes.
Causes of Neutropenias
1. Margination
2. Impaired production
3. impaired maturation
4. sequestration/destruction/loss
5. chemotherapy
6. chronic benign neutropenia
7. severe congenital neutropenia
Lymphocytosis and Lymphopenia
Put it into clinical context. Take a history with knowledge of lymphocytosis and
lymphopenia. Do a physical examination
Determine if;
Lymphadenopathy present, location, size and texture of glands
Presence of local inflammatory or neoplastic disase
Is splenomegaly present? Test for monoclonality of lymphocytes.
Causes of abnormal lymphocyte count
Proliferation of normal cells responding to antigen
Chronic leukaemia
\Acute leukaemia
Splenectomy
Causes of Lymphopenia
AIDS
Lymphoma
Splenomegaly
Chemotherapy
Protein losing enteropathy

Further Reading.
Useful website is www.hemeteam.com
A collection of tutorials on basic principles of diagnosis, pathogenesis and
treatment of common blood disorders. It gives you information and option to go
deeper if you wish. Many tutorials have MCQs with instant feedback.