1

BLOOD
Blood is a type of connective tissue composed of liquid extracellular matrix
called blood plasma in which various cells and cell fragments like - Red blood cells, white
blood cells and platelets are suspended. Blood transports oxygen and nutrients from
gastrointestinal tract to every cell of the body. The oxygen and nutrients subsequently diffuse
from blood into the interstitial fluid and than in to cells. Carbon dioxide and other metabolic
wastes moves in reverse direction (i.e. body cells interstitial fluid Blood Transport
to various excretory organs like kidneys, lungs and skin.)
FUNCTIONS OF BLOOD
We can categories function of blood under four headings-
1) Transportation
2) Regulation
3) Protection
4) Other
TRANSPORTATION:-
-It transports oxygen from lungs to cells of the body.
-It transports carbon dioxide from cells to lungs.
- It transports nutrients from g.i tract to cells.
-It transports hormones from endocrine glands to other target cells.
-It transports heat and other metabolic waste products to various organs for elimination.
REGULATION:-
-It regulates homeostasis of all body fluid.
-It regulates pH of body through buffer mechanism.
-It regulates body temperature.

2

PROTECTION:-
-It can clot by an intrinsic and extrinsic mechanism, which protects it against excessive loss
after an injury.
-It protects against disease by caring on phagocytosis (due to WBCs), plasma proteins,
antibodies interferon and complements.
OTHER:-
-It provides color to skin Up to certain extent color of skin depends on skin blood volume,
oxyhemoglobin and blood flow.
AMOUNT OF BLOOD:-
In human body, blood constitutes 20% of extra cellular fluid, amounting to 8%
of the total body weight. In an averaged sized adult male its volume is 5 to 6 liter (1.5 Galen)
and in female is 4 to 5 liter (1.2 Galen).
PROPERTIES OF BLOOD:-
Blood is a denser, viscous and feels slight sticky.
Temperature of blood is 38
0
C (100.4
0
F) i.e. 1
0
C higher than oral and rectal temperature.
Specific gravity - It is the ratio of weight of certain volume of substance to the weight of
an equal volume of water. The specific gravity of blood varies from 1.052 to 1.061.It
depends largely on number on RBCs.
Osmotic pressure - Though the composition of blood under goes continual changes,
although small changes because of passage of water, dissolved food, and waste products
into and out of blood. Therefore, osmotic pressure also gets slightly change but kidneys
rapidly correct this variation. The osmotic pressure of human blood averages about 5900
mm of Hg or 7.8 atmospheres (about that of 0.9 % NaCl solution).
Viscosity - The viscosity of fluid is due to mutual attraction of its molecules, which
thereby offer resistance to flow. The viscosity of blood is five times as great as water. It is
mainly depends on number of blood corpuscles or the amount of proteins.
pH- Ranging from 7.35 to 7.45
3

COMPONENTS OF BLOOD
Extracellular fluid [ECF] Blood 8%


Plasma55% Formed elements & cell fragments 45%
RBCs
Water (91.5%) Protein (7%) Other solutes (1.5%) WBCs
Albumin 54% Electrolytes Platelets
Globulins 38% Nutrients
fibrinogen7% Gases
All others 1% Regulatory substance
Waste products








4

Plasma
Plasma is straw colored liquid portion of blood. It contains 91.5% of water and
8.5% of solutes most of which are protein. The total plasma protein concentration is 64
83g/lit. There are two main principal groups of plasma proteins-
Albumin

1
- Globulin

2
- Globulin
Globulin -Globulin
- Globulin
Fibrinogen
Average normal concentration of main plasma proteins in g/lit
Albumin: Globulin: fibrinogen
54: 38: 07
A number of proteins with specific physiological functions are isolated with
the help of electrophoresis from globulin fraction. These proteins are prothrombin, Plasma
tromboplastin, Isohemaglutinin, Angiotensinogen, immunoglobulin and anterior pituitary
hormones.
Constituent of plasma-
Organic substances-
1) Proteins Albumin ,Globulin ,Fibrinogen
2) Carbohydrates Mainly in the form of Glucose
3) Fats Neutral fats ,Phospholipids ,Cholesterol
4) Amino acids Both essential and non essential amino acid
5) Non-protein nitrogen substances Ammonia, Creatinin, Xanthin, Hypoxanthine, Urea,
Uric acid etc.
6) Internal secretion Many hormones
7) Enzymes Amylase, Carbonic anhydrase, alkaline phosphate, Acid phosphate, Lipase,
Esterase and Transaminase.
8) Antibodies Many Immunoglobulins
5

Inorganic substances of plasma
Sodium Potassium Magnesium
Calcium Chloride Iodide
Iron Phosphate Copper
Oxygen and carbon dioxides gases are present.
Properties of plasma proteins
Molecular weight and shape
Molecular weight of plasma albumin 69000
Molecular weight of fibrinogen 330000
Note - Capillary permeability depends on size and shape of plasma proteins. Albumin passes
more rapidly than others do. In small amount plasma, proteins are also present in lymph.
Osmotic effect Plasma proteins are responsible for osmotic pressure of about 25 mm
Hg and thus influence the exchange of fluid in between blood and tissue spaces.
Specific gravity Specific gravity of plasma proteins are 1.026.
Buffer action Plasma proteins have 1/6 of total buffering action of blood.








6

Table No 1.3
Main features of plasma proteins
M.wt Molecular weight, P.conc. Plasma concentration
Origin of plasma proteins

S.no

Fraction

M.wt.

P.conc.

Function
1 Pre albumin 6.010
4
0.3 Bind thyroxin &
Triodothyronine
2 Albumin 6.910
4
40 Colloid osmotic
pressure ,bind
hormone ,fatty acid
,Bilirubin, drugs
3 1-Globulin 4.510
4
4.0 Includes antiprotease
4 2-Globulin
Ceruloplasmin
Hepatoglobulin

1.610
4

9.010
4

0.4
1.2

Copper transport
Binds hemoglobin
5 -Globulins
Transpherrin

9.010
4


2.4

Iron transport
6 Co of
complement

Plasminogen
Fibrinogen
Prothrombin
2.010
4

1.410
4

3.510
4

6.810
4

1.6

0.7
3.0
1.0


Fibrinolysis
Blood clotting
Blood clotting
7 -Globulins
IgG
IgA
IgM
IgE

1.510
5

1.710
5

1.010
6

2.o10
5



10.0
1.6
1.0
0.1


All Igs are antibodies
7

Albumin, fibrinogen and prothrombin are formed exclusively by liver.
Immunoglobulins are formed in lymphoid tissue by the plasma cells
Red Blood cells (Erythrocytes)
Erythros- Red
Red blood cells or erythrocytes contain the oxygen carrying protein i.e.
hemoglobin, which is a pigment that gives whole blood its red color. These cells are non
nucleated.
Morphology
Normal size Diameter 7.2m
Thickness At periphery it is thick about 2.2m and at center 1m
Shape Biconcave shape (Dumb bell shaped)
Surface area 120sqm
Volume 85 90 cu mm
Color Single RBC is light brown or yellow color but when seen in bulk it appears red in
color.
PROPERTIES OF RBCs
Rouleaux formation When blood is taken out of blood vessels, the RBCs pile up one
above another like the pile coins. The properties of RBCs are called Rouleaux formation.
Specific gravity of RBCs are 1.092 1.101
Packed cell volume When blood is collected in centrifuged tube along with proper
anticoagulant and centrifuged for 30 min. at 3000 rpm. The RBCs settle at the bottom of
tube leaving clear plasma at top. The RBCs form 45% of total blood volume. This is
called as packed cell volume or Hematocrit.
Suspension stability During circulation, the RBCs remain suspended uniformly in the
blood. This property of RBCs are called suspension stability.
Development of RBCs
There are two theories-
Intra vascular
Extra vascular
8

Initially, it was believed that RBCs were formed only in intravascularly from
capillary epithelium.
T Urnbull and Glimour (1941) have shown that they are formed from extra
vascular source (parent cells in an extra vascular cell is known as erythropoiesis).
Site of erythropoiesis
In fetal life During embryonic life erythropoiesis occurs in three stages.
1) Mesoblastic stage During early two months of intra uterine life, the primitive RBCs are
produced from mesenchyme of yolk sac.
2) Hepatic stage During early three months liver is main organ, some erythrocytes are
produced from spleen and lymphoid organs.
3) Myeloid stage During last three months, erythrocytes are produced from bone marrow
and liver.
In Postnatal life and adult In newborn babies, growing children and adults, RBCs are
produced only from red bone marrow.
Up to 5 to 6 year Red bone marrow of all bones.
Up to 6 to 20 years Red bone marrow of long bones and membranous bones.
After 20 years RBCs are produced red bone marrow of long bones Moreover, end of
long bones.
During bone disorder RBCs are produced in spleen.
Stem cells
The stem cells are primitive cells in bone marrow, which give rise to different
blood cells. These are called as pluripotent hemopoietic stem cells (PHSC).The stem cells
are also called as committed stem cells when these are designated to form a particular
type of blood cells. These are of two types-
1) Lymphoid stem cells- give rise to lymphocytes.
2) Colony forming blastocytes give rise to blood cells other than lymphocytes. These are
of three different units.
i. Colony forming unit Erythrocyte(CFU-E)
ii. Colony forming unit Granulocytes /Monocytes (CFU-GM)
iii. Colony forming unit Megakaryocytes (CFU-M)
9


CHANGES DURING ERYTHROPOIESIS
CFU-E passes through different stages and finally become matured red cells.
During these process four important changes occurs-
1) Reduction in size of cells (from diameter 25 m to 7.2 m)
2) Disappearance of nucleoli and nucleus
3) Appearance of hemoglobin
4) Changes in stain property of cytoplasm

STAGES OF DEVLOPMENT
1) Hemocytoblast A big cell of 19 23 m in diameter with a large nucleus, basophilic
cytoplasm (according to intravascular theory this stage starts with endothelial cells).
1. Proerythroblast (Megaloblast) - It is 20 m in diameter. Nucleus is large with two or
more nucleoli and reticular network. Hemoglobin is absent and cytoplasm is basophilic.
2. Early Normoblast About 15 m in diameter, nucleoli disappears condensation of
chromatin network, basophilic cytoplasm. This cell is also called as basophilic
erythroblast.
3. Intermediate Normoblast 10 20 m in diameter. Nucleus is still present; hemoglobin
start appearing .cytoplasm is already basophilic.
4. Late Normoblast Diameter of cell reduced to 8 10 m ink spot nucleus is present due
to condensation of chromatin network. Quantity of hemoglobin increase, cytoplasm
becomes almost acidophilic. The nucleus disintegrates and disappears.
5. Reticulocytes (immature RBCs) It is slightly larger than mature RBCs. It has reticular
network .It is basophilic due to presence of remnant of Golgi apparatus, mitochondria and
other organelles.
6. Mature erythrocytes Reticular network disappears and cell becomes mature RBCs. The
mature cell is biconcave, smaller with diameter 7.2 m .It is with hemoglobin but without
nucleus.
It requires 7 days for development of mature RBCs from Proerythroblast, 5 days
from Reticulocytes and 2 days from Reticulocytes to mature RBCs.


10



Table No 1.4
Stages of RBCs Developments
Stages of Cell
British school American School
Proerythroblast Megaloblast
Early Normoblast Early Erythroblast
Intermediate Normoblast Late Erythroblast
Late Normoblast Normoblast
Reticulocyte Reticulocyte
Erythrocyte Erythrocyte
Factors necessary for erythropoiesis
Various substances are necessary for the development and maturation of RBCs.
General factors
Maturation factors
General factors Hormone erythropoietin (hemopoietin or erythrocytes stimulating) which
are secreted from kidney.
Hypoxia is responsible for the production of erythropoietin. It causes formation and release of
new RBCs into circulation 4 to 5 days after being secreted. Apart from erythropoietin, some
other hormones like thyroxin and testosterone are also important for erythropoiesis.
Other factors Interleukins -3 stimulate growth of stem cells or RBCs. These
are called as growth inducers. Some vitamins like B and C are also important for this.
Maturation factors
11

Vitamin B12
Intrinsic factor of castle
Folic acid
Life Span of RBCs :- 120days
Function of RBCs
Respiration - RBCs carry O
2
and CO
2

Acid base balance It is carried out by hemoglobin and other intra cellular buffer.
Maintenance of iron balance By special permeability of cell membrane RBCs helps to
maintain positive or negative iron balance in the blood.
Viscosity of blood
Various pigments are derived from hemoglobin after disintegration of RBCs e.g.
Bilirubin and Biliverdin etc.

HAEMOGLOBIN
Hemoglobin is most important constituent of Red blood cells. It is a colored
compound protein. It consists of two parts simple protein globins (96%) and red pigment
containing part hem. Hem is an iron containing porphyrin known as iron protoporphyrin IX.
This element is functional part of hem. The iron renders hemoglobin absolutely indispensable
for life. It constitutes about 1/3 part of RBCs. The porphyrins are tetrapyrroles. The pyrrole
rings are number asI, II, III and IV. The carbon of methane bridges are labeled as ,, , and
the position to which side chains are attached are numbered 1- 8 .
The iron in hem is in the ferrous (Fe
+
) form. The iron is attached to N of each
pyrrole ring and to N of iminazole group in associated globulin. A bond is available for loose
union with O
2
and CO. In oxidized Hemoglobin this placed is occupied by an OH group on
ferric (Fe
+
) atom.


12

SYNTHESIS OF HEM
The starting substances for hemoglobin synthesis are succinic acid and Glycine.
COOH COOH COOH
CH
2
(Succinic acid) CH
2
CH
2

CH
2
CH
2
CH
2
CO CO CO
OH (Glycine) HC.NH
2

CH2NH2
+CO
2

H COOH ( amino laevulinic acid)
CH.NH
2
amino keto adipic acid
COOH (Unstable)
These reactions are brought about by enzyme amino laevulinic acid
synthetase (ALA synthetase) with pyridoxal or pyridoxal phosphate as co- enzyme. Two
molecules of amino laevulinic acid condense to form porphobilinogen

COOH COOH COOH
CH2 CH2 CH2
CH2 + H2C ALAdehydrase CH2 CH2
CO OC CH2NH2 C C
CH2 HC C CH2NH2
NH2 N
H
2 molecules of lavulinic acid prophobillinogen
13


Hemoglobin structure
Hemoglobin is made from four polypeptide chains of two types each in
duplicate .It consist of two and two- subunit each associated with one hem group. Adult
hemoglobin in brief is written as Hb A (22).
Physiological Hemoglobin
Adult Hemoglobin :- It is of two types
Hemoglobin A (22) It is preponderant form. It is spheroid molecule with molecular
weight of 68000.
Hemoglobin A2 (22) It is minor component in normal adults. Delta chains have a
slightly different amino acid composition as compare to chains.

Fetal hemoglobin :-
Hemoglobin F (22) - It is present in fetal Red blood cells and it is disappeared
2-3 months after birth. It differs from Hemoglobin A
In - Chain in place of chain
- Greater affinity for oxygen
- More resistance to alkali
Iron metabolism
Functions of iron:-
To carry O
2
as part of hemoglobin.
Brain development and function
Regulation of body temperature
Muscular activity
Catecholamine metabolism
Iron is a critical element in iron-containing enzymes, including the cytochrome system in
mitochondria.
It is important for electron transport and energy metabolism.
14

Table No 1.5
Body Iron Distribution
Body Iron Distribution


Iron Content, mg
Adult Male, 80 kg Adult Female, 60 kg
Hemoglobin 2500 1700
Myoglobin/enzymes 500 300
Transferrin iron 3 3
Iron stores 6001000 0300


The Iron Cycle in Humans
Iron absorbed from the diet or released from stores circulates in the plasma
bound to transferring (blobbed glycoprotein with two iron binding sites), the iron transport
protein. Transpherrin that carries iron exists in two formsmonoferric (one iron atom) or
diferric (two iron atoms). The turnover of Transpherrin-bound iron is very rapid about 6090
min. almost all of the iron transported by Transpherrin is delivered to the erythroid marrow.
The plasma iron level and the erythroid marrow activity affects the clearance time of
Transpherrin bound iron. When erythropoiesis is markedly stimulated, the pool of erythroid
cells requiring iron increases and the clearance time of iron from the circulation decreases.
The half-clearance time of iron in the presence of iron deficiency is as short as 1015 min. in
suppression of erythropoiesis, the plasma iron level typically increases and the half-clearance
time may prolong to several hours. Normally, the iron bound to Transpherrin turns over 10
20 times per day. Assuming a normal plasma iron level of 80100 g/dL, the amount of iron
passing through the Transpherrin pool is 2024 mg/d.
The iron-Transpherrin complex circulates in the plasma until it interacts with
specific Transpherrin receptors of marrow erythroid cells. Diferric Transpherrin has the
15

highest affinity towards it. While apotranspherrin (Transpherrin not carrying iron) has very
little affinity. Transpherrin receptors are found on cells of body. When iron-bearing
Transpherrin interacts with its receptor, the complex is internalized via clathrin-coated pits
and transported to an acidic endosome, here iron is available for hem synthesis while the
Transpherrin-receptor complex is recycled to the surface of the cell. Certain Transpherrin
receptor protein may release into circulation. In erythroid cell, iron is in excess needed for
hemoglobin synthesis. This mechanism of iron exchange also takes place in other cells of the
body expressing Transpherrin receptors, especially liver parenchymal cells. The iron involved
in hemoglobin synthesis enters the circulation as new red cells are released from the bone
marrow. Only after death of RBCs, iron becomes available for reutilization.
0.81.0% of red cells turn over each day. At the end of its life span, the red
cell is recognized as senescent by the cells of the reticuloendothelial (RE) system, and the cell
undergoes phagocytosis. Once within the RE cell, the hemoglobin from the ingested red cell
is broken down, the globin and other proteins are returned to the amino acid pool, and the
iron is return back to the surface of the RE cell, where it is presented to circulating
Transpherrin.
Physiological variation in Hemoglobin and Red cell count
At different periods of life :-
After delivery when the cord is tied hemoglobin and red cell count. After birth, hemoglobin is
higher than 200gm/dl and red cell count is higher than 6.0 10
12
/l.
Postnatal period: - Immediate after postnatal period, hemoglobin falls. Steeply to a min by
about 2
nd
month and cells, become microcytic with the development of physiological iron
deficiency.
Neonate: - In neonates and until about 2
nd
month the average mean cell hemoglobin (MCH) is
34pg.
Puberty:-At puberty, hemoglobin and red cell count rise gradually to almost adult levels by
the time of puberty. In women, this level tends to be significantly lower than those of men
.The factors responsible for that are hormonal influence on hemopoiesis and menstrual blood
loss.
16

Pregnancy:-In normal pregnancy, there is increase in erythropoietic activity. At the same
time, increase in plasma volume occurs .This results in progressive fall in hemoglobin, pack
cell volume (PCV) and red cell count.
Old age: - At old age, hemoglobin is reported to fall progressively in men. By contrast, in
older women this level tends to rise so that a difference of 20g/l in younger age groups is
reduced to 10g/l or less in old age.
Exercise: - Strenuous muscular activity in short term raises Red cell count and
hemoglobin (largely because of reduction in plasma volume and to lesser extent to re-
entry in the circulation of cells previously sequestered in the spleen).
Posture:- There is small but significant increase in hemoglobin and red cell count as
posture changes from lying down to sitting , specially in women.
Athletes:- There is slightly lower in hemoglobin and red cell count in athletes due to
increase plasma volume. It is called as sports anemia.
Seasonal variation:- Minor seasonal variation occurs in hemoglobin and red cell count
but evidence for it is conflicting . One study demonstrate that hemoglobin is lower in
summer than in winter due to 5% increase in plasma volume.
Effect of high altitude :- High altitude raise hemoglobin and red cell count due to less
oxygen concentration in air.
VARIATION IN THE SIZE OF RBCS
1) Microcytes (RBCs of small size)
Iron deficiency anemia
Prolong forced breathing
Increase osmotic pressure in blood
2) Macrocytes (RBCs of large size)
Intrinsic factor deficiency anemia
Folic acid deficiency anemia
Muscular exercise
Decrease osmotic pressure in blood
3) Anisocytes (RBCs of unequal size)
Pernicious anemia
17

VARIATION IN THE SHAPE OF RBCS
Following are abnormal shapes of RBCs some of these are present in different
types of anemia.
Crenation Shrinkage as in hypertonic solution
Spherocytosis Globular form as in hypotonic solution
Elliptocytosis Elliptical shape as in certain types of anemia
Sickle cell Cresentric shape as in sickle cell anemia.
Poikilocytosis Unequal shape due to deformed cell membrane
Table No 1.6
Normal Hematological values

RBCs count
10
12
/lit
Hemoglobin
g/l
WBCs count
10
9
/lit
Platelets
10
9
/lit
Birth 6.0 1.0 180 40 18 8 150- 450
3
rd
Day 5.3 1.3 180 30 15 8 210 500
1
st
Month 4.2 1.2 140 25 12 7 -
2
nd
Month 3.7 0.6 112 18 10 5 210 650
3-6 Month 4.7 0.6 126 15 12 6 200 550
1
st
Year 4.5 0.6 126 15 11 5 200 550
2-6 Year 4.6 0.6 125 15 10 5 200 450
6-12 Year 4.6 0.6 135 20 9 4 180 400


18


WHITE BLOOD CELLS (LEUCOCYTES)
Leuco white
The name it self suggest that WBCs or leukocytes are colorless cell fragments. It has
nuclei but it does not contain hemoglobin. These are classified into two main types
Granulocytes and Agranulocytes, depending on present or absent of cytoplasmic granules.
These are visible only after staining.
Table No 1.7
Types of WBC and Their Morphology
S.no WBC Granules Stain No. of lobes in
Nucleus
Diameter
in m
% Life span
1 Neutrophils Present Neutral Multi lobed 10 12 50 70 02 05
2 Eocinophils Present Acidic Bi lobed 10 14 2 - 4 07 - 12
3 Basophils Present Basic Bi lobed 8 10 0 - 1 12 15
4 Monocytes Absent - Round, oval, kidney
shaped
14 18 2 - 6 02 05
5 Lymphocyte absent - Oval, kidney shaped 10 12 20 30 - 1

Normal count of white blood cells
The Total white blood cell count in normal adult is from 4000 to 11000 / cu mm.



19

Genesis of Leucocytes (Leucopoiesis)
It includes Granulopoesis
Monocytopoiesis
Lymphopoesis
Site Granulocytes and Monocytes In bone marrow only
Lymphocytes Mainly in lymphogenous organ like lymph
Glands, Spleen, Thymus, Tonsil etc
Process
All leucocytes develop from hemopoietic stem cell. The committed stem cell,
which gives origin to granulocyte, and monocytes are same but different from the cell, which
gives raise to lymphocyte. The progenitor cells that develop from committed stem cell
myeloid series GM. Later on, this is divided into two separate cells one is responsible for
granulocyte formation and second is for monocytes.








20

Pluripotent Hemopoietic Stem Cell

Lymphoid stem cell Colony forming blastocytes

Colony forming unit E Colony forming M
Colony forming GM

Lymphoblast Myeloblast

Promyelocyte Monoblast

Neutrophil Basophil Eocinophil
Myelocyte Myelocyte Myelocyte
Neutrophil Basophil Eocinophil
Metamyelocyte Metamylocyte Myelocyte
Lymphcyte Neutrophil Basophil Eocinophil
Monocytes

FUNCTIONS OF WHITE BLOOD CELLS
Neutrophil
Destruction of invading bacteria by the process of phagocytosis and lysis (digestion of
bacteria, the production of hydrogen per oxide, release of myeloperoxidase into
tenphagosome and release of lysozymes, lactoferin and p
H
reduction.
21

Eocinophils
Almost as same as that of Neutrophils.
Phagocytosis of bacteria, destroyed cell and antigen antibody complex. Phagocytic
activities of Eocinophils are less than Neutrophils.
It tends to accumulate at the site of histamine release. It has capacity to antagonize or
inactivate histamines and other chemical mediator of inflammation like 5HT and
bradikinin.
It has wormicidal function.
Basophils
Basophile play an important role in healing process after inflammation and acute allergic
condition. Its count increase during healing process.
It releases some important substances like
Histamine It produces hypersensitivity reaction.
Heparin Prevents intra vascular blood clotting
Hyaluronic acid Important for deposition of ground substances in the basement
membrane.
Protease and myeloperoxidase These enzymes aggravates the inflammatory response.
It has IgE receptors. This helps them to produce inflammatory response.
Monocytes
It play an important role in defense mechanism by phagocytosis action along with
Neutrophils.
It secretes interleukins - (L-1), colony stimulating factors and platelets activating factors
(PAF).
These are precursors of tissue macrophages like kupffers cell in liver, alveolar
macrophages in the lungs and spleen macrophages.


22

Lymphocytes
It plays very important role in immunity. Lymphocytes are of two types T lymphocyte
and B lymphocyte. T lymphocytes are responsible for cell-mediated immunity while B
lymphocytes are responsible for humoral immunity.
Physiological Variation In Total Lucocyte Count
Age In infant WBCs count is about 20,000/cu mm and in children, it is about 10,000 to
15,000/cu mm.
Sex WBCs count is slightly more in male than in female. However, in female, it
increases during menstruation, pregnancy and parturition.
Diurnal variation Difference of 14% for total leucocytes count, 10% for Neutrophils,
14% lymphocyte count, 20% for Eocinophils. Minimum count is found in morning with
subject at rest.
Sleep During sleep, WBCs count is minimum.
Inj. Adrenalin (epinephrine) It causes increases in leucocytes count and all major
types of leucocytes and platelets.
Exercise Random activity may raise the count slightly. Strenuous exercise causes raises
up to 3010
9
/lit.
Emotion It may possibility causes a leucocytes count.
Cigarette smoking- It causes significant increase in leucocytes count. (All types of cell
proportionately affected)
Pregnancy Pregnancy cause moderate leucocytosis up to 15 10
9
/lit owning to
neutrophilia with the peak in 2
nd
trimester.
Environmental factor It may influence leucocytes count. In tropical Africa, there is
tendency for reversal of Neutrophils: lymphocyte ratio in individual with low leukocyte
count.
Genetics Genetics also play important role in low leukocyte count especially in
Neutrophils.
Oral contraceptives Increase total leukocyte count
Vaccination Elder people receiving influenza vaccine show low Leucocyte count
owning to decrease in lymphocytes.

23

Table No 1.8
Pathological causes of variation in WBCs
S.no WBC types High count Low count
1 Neutrophils Bacterial infection
Burn, stress, inflammation
Radiation exposure, drug
toxicity, vit. B12 deficiency
(SLE)
2 Lymphocyte Viral infection ,some leukemia Prolonged illness,
Immunosuppration or t/t with
cortisol
3 Monocytes Viral or fungal infection, some leukemia
other chronic disease
Bone marrow suppression,
t/t with cortisol
4 Eocinophils Allergic reactions, parasitic infestation,
Autoimmune diseases
Drug toxicity, Stress
5 Basophils Allergic reaction, leukemia,
Hypothyroidism
Pregnancy ovulation and stress
or hypothyroidism
Thrombocytes (platelets)
These are small colorless , non nucleated and moderately refractive bodies .Its
diameter is 2 to 4 m and volume ranges from 7 to 8 cubic m . Normally these are spherical
and rod shaped and becomes oval or disc shaped when inactivated. Some time these are
found in dumb bell, comma, cigar or any other unusual shape.
Normal platelets count
Normal platelets count ranges from 2, 00,000 to 4, 00,000 /cu mm and averages about 2,
50,000/cumm.
Life span - 10 days



24

Functions of platelets
Normally platelets are inactive these functions are only when activated.
The platelets are responsible for formation of intrinsic prothrombin activator (substance
responsible for onset of blood clotting).
The cytoplasm of platelet contains contractile proteins like actine, myosin and
thrombosthenin, which is responsible for clot reaction.
It accelerates the process of hemostasis.
Due to adhesive property, the platelets can seal damage blood vessels. By the formation of
temporary plug.
The Platelet derived growth factor (PDGF) formed in cytoplasm of platelets are useful in
repair of capillary endothelium and other structure of ruptured blood vessels.
Platelets encircle the foreign invaders by the property of agglutination and kill them by
the process of phagocytosis.
Physiological variation in platelets count
Age - Platelets are less in infants (1, 50,000 to 2, 00,000 cu mm) and reaches normal at
3
rd
month after birth.
Sex In female, it is 20% higher than that of male.
Menstruation It decreases at the time of menstruation.
Diurnal variation Only slight variation of up to 5% occurs during coarse of day as well
as from day to day.
Exercise Strenuous exercise causes 30 40 % increase in platelet count.
Food The platelets count increases after taking food.
High altitude The platelet count is increased at high altitude.



25

Anemia
Anemia is present in adult if Hematocrit is less than 41% (hemoglobin <
13.5g/dl) in male or 37% (Hemoglobin <12g/dl) in female. A WHO expert Group proposed,
Anemia or iron deficiency should be considered to exist when hemoglobin is below the
following level.
Table No 1.9
Cut Of Points For The Diagnosis Of Anemia

Venous Blood MCHC
Adult males 13gm/dl 34%
Adult females
(Non pregnant)
12gm/dl 34%
Adult Females
(Pregnant)
11gm/dl 34%
Children
(6 months to 6 years)
11gm/dl 34%
Children
(6years to 14years)
12gm/dl 34%
At all ages the normal MCHC should be 34%; the value below this
indicates the RBCs are hypochromic, which occurs in iron deficiency anemia. Personal and
family history is important for diagnosis of congenital anemia. Poor diet results in folic acid
deficiency and contributes to iron deficiency. Bleeding is present universally in iron
deficiency anemia. Physical examination includes attention to sign of primary hematological
diseases (Lymphydenopathy, hepatosplenomegaly, bone tenderness).Mucosal changes such
as a smooth tongue megaloblastic anemia.
26

CLASSIFICATION
ON THE BASIS OF PATHOPHYSIOLOGY
Decrease production
Hemoglobin synthesis
Iron deficiency
Thalassemia
Anemia of chronic disease
DNA synthesis
Megaloblastic anemia
Stem cell
Aplastic anemia
Myeloproloferative leukemia
Bone marrow infiltration
Carcinoma
Lymphoma
Pure red cell aplasia
Increased destruction
Blood loss
Hemolysis (intrinsic)
Membrane Hereditary spherocytosis,Elliptosis
Hemoglobin-Sickle cell anemia, Unstable hemoglobin
Glycolysis Pyruvate kinase deficiency etc.
Oxidation G6PD deficiency
Hemolysis (extrinsic)
Immune Warm antibody, Cold antibody
Microangiopathic Thrombotic thrombocytopenic purpura, Hemolytic uremic syndrome,
Mechanical cardiac valve, Paravalvular leak
Infection Clostridial
Hypersplenism



27


ON THE BASIS OF MCV
Microcytic
Iron deficiency
Thalassemia
Anemia of chronic diseases
Macrocytic
Megaloblastic
Vitamin B12 deficiency
Foliate deficiency
Non-Megaloblastic
Myelodysplasia
Chemotherapy
Liver disease
Increased reticulocytosis
Myxedema
Normocytic
Many causes




28

Iron Deficiency Anemia
Key for diagnosis
Both pathognomonic : Absent bone marrow iron stores or serum ferritin < 12g/l
Caused by bleeding in adults unless proved otherwise.
Response to iron therapy
Iron deficiency is the most common cause of anemia .iron is important for the
formation of Hem and other enzymes. Total body iron ranges between 2 and 4 gram
approximately 50mg/kg in man and 35mg/kg in women. Most of the iron present in
hemoglobin in circulating RBCs. one milliliter of packed RBCS contain approximately one
milligram of iron.
Causes of iron deficiency anemia
Deficient diet
Decreased absorption
Malabsorption from disease (sprue, Crohn's disease)
Malabsorption from surgery (post-gastrectomy)
Acute or chronic inflammations
Increased requirement
Pregnancy
Lactation
Blood loss
Gastrointestinal
Menstrual
Blood donation
Hemoglobinuria
Iron sequestration
Pulmonary hemosyderosis


29

Clinical findings
A symptoms and signs
The only symptoms of iron deficiency anemia are those of anemia itself. (Easy fatigability,
tachycardia, palpitations and tachypnoea on exertion). Severe deficiency causes skin and
mucosal changes, including smooth tongue, brittle nails and cheilosis .dysphagia because of
formation of esophageal webs (Plummer Vinson syndrome) also occurs. Many iron
deficient patient develop pica ,craving for specific foods (ice ,chips ,lettuce etc.) often not
rich in iron.
B. Laboratory findings
Iron deficiency develops in stages. First is depletion of iron store .at this point
there is anemia and no changes in red cell size. The serum ferritin becomes abnormally low.
A ferritin value less than 30g/lit is highly reliable indicator of iron deficiency .The serum
total iron binding capacity rises.
After iron stores has been depleted, RBCs formation will continue with
deficient supply of iron .Serum iron values decline to less than 30 g/dl and transferin
saturation to less than 15%.
In the early stage, MCV remains normal. Subsequently, the MCV falls and
blood smear shows hypochromic microcytic cells. With further progression ,anisocytosis
(variation in RBCs size) and polikilocytosis (variation in shape of RBCs) develop Severe iron
deficiency will produce a bizarre peripheral blood smear ,with severely hypochromic cell,
target cell , hypochromic pencil shaped cells and occasionally small number of nucleated
RBCs. The platelet count is commonly increased.