BLOOD

 Blood is a specialized connective tissue in which cells are

suspended in fluid extracellular material called plasma .
 Propelled mainly by rhythmic contractions of the heart,
about 5 L of blood in an average adult moves unidirectional
within the closed circulatory system.
 The so-called formed elements circulating in the plasma
are erythrocytes (red blood cells), leukocytes (white
blood cells), and platelets .
 CONT...

 Blood is a distributing vehicle, transporting O2, CO2 ,

metabolites, hormones, and other substances to cells throughout
the body.
 O2 is bound mainly to hemoglobin in erythrocytes and is much
more abundant in arterial than venous blood.
 While CO2 is carried in solution as CO2 or HCO3-, in addition
to being hemoglobin-bound.
 Leukocytes have diversified functions and are one of the body’s
chief defenses against infection.
 COMPOSITION OF PLASMA

 Plasma is an aqueous solution, pH 7.4, containing substances of

low or high molecular weight that make up 7% of its volume.
 The major plasma proteins include the following¦
 Albumin , the most abundant plasma protein, is made in the liver
and serves primarily to maintain the osmotic pressure of the
blood.
 α Globulins and β -globulins ,
 COMPOSITION OF PLASMA

 made by liver and other cells, include transferrin and other

transport factors;
 Fibronectin ; prothrombin and other coagulation factors;
lipoproteins and other proteins entering blood from tissues
 γ-Globulins , which are immunoglobulins (antibodies)
secreted by plasma cells in many locations.

5
 COMPOSITION OF PLASMA

 Fibrinogen , the largest plasma protein (340 kD), also made in

the liver, which, during clotting, polymerizes as insoluble,
cross-linked fibers of fibrin that block blood loss from small
vessels.
 Complement proteins , a system of factors important in
inflammation and destruction of microorganisms.
 Plasma Proteins
Plasma
Clear liquid made up of 92% water and 8%
organic and inorganic biochemicals.

albumin globulin

Plasma Proteins

fibrinogen prothrombin 7
 BLOOD CELLS

 Blood cells can be studied histologically in smears prepared

by spreading a drop of blood in a thin layer on a microscope
slide.
 In such films the cells are clearly visible and distinct from one
an other, facilitate observation of their nuclei and cytoplasmic
characteristics.
 Blood smears are routinely stained with special mixtures of
acidic (eosin) and basic (methylene blue) dyes.
 BLOOD CELLS

 These mixtures may also contain dyes called azures that are
more useful in staining cytoplasmic granules containing
charged proteins and proteoglycans.
 Azurophilic granules produce metachromasia in stained
leukocytes like that seen with mast cells in connective tissue.

9
 BLOOD SMEAR PROCEDURE

 Remember that the cells you see in a blood smear have not
been sectioned. Instead you are seeing whole cells dried
down on the glass
 After the smear is made, it is air-dried and then stained.
Common stains are Wright's stain and Giemsa stain.
 The stains generally include two or more dyes, one of them a
basic dye (often methylene blue) and another an acidic dye
(usually eosin).
 BLOOD SMEAR PROCEDURE

 Reddish-blue azures are formed when methylene blue is

oxidized.
 Cells usually stain pink/red with acidic dye and nuclei
stain purple/black with basic dye, while specific
granules stain characteristically.

11
Human blood smear, with RBCs, WBCs and platelets
 CELLS OF THE BLOOD

 Erythrocytes (red blood cells, RBC)

 Platelets (thrombocytes)
 Leukocytes (white blood cells, WBC)
 Granulocytes (with specific granules)
Neutrophil (~60% of WBC)
Eosinophil (~4% of WBC)
Basophil (<1% of WBC)
 Agranulocytes (without specific granules)
Lymphocyte (B-cell, T-cell) (~27% of WBC)
Monocyte (~8% of WBC)
 Erythrocytes

 Erythrocytes (RBCs) are terminally differentiated structures

lacking nuclei and completely filled with the O2 carrying
protein hemoglobin.
 RBCs are the only blood cells whose function does not
require them to leave the vasculature.
 Because of their uniform diameters and their presence in most
tissue sections, RBCs can often be used by histologists as an
internal standard to estimate the size of other cells or
structures.
 Erythrocytes cont.…

 Observations in vivo show that at the angles of capillary

bifurcations, erythrocytes with normal adult hemoglobin
frequently assume a cuplike shape.
 In larger blood vessels RBCs often adhere to one another
loosely in stacks called rouleaux.
 Erythrocytes are normally quite flexible, which permits them
to bend and adapt to the irregular turns and small diameters of
capillaries.

16
 Erythrocytes cont.…

 The plasmalemma of the erythrocyte, because of its ready

availability, is the best-known membrane of any cell.
 It consists of about 40% lipid, 10% carbohydrate, and 50%
protein.
 Most of the latter are integral membrane proteins.
 Erythrocyte cytoplasm lacks all organelles but is densely filled
with hemoglobin, the tetrameric O2 carrying protein that
accounts for the cells’ uniform acidophilia.
 Erythrocytes cont.…

 When combined with O2 or CO2, hemoglobin forms

oxyhemoglobin or carbaminohemoglobin, respectively.
 The reversibility of these combinations is the basis for the gas-
transporting capability of hemoglobin.
 combination of hemoglobin with carbon monoxide (CO) is
irreversible, however, reducing the cells 'capacity to transport O2
 Human erythrocytes normally survive in the circulation for about
120 days.
 Red blood cells in a blood smear

RBC

Platelet
 RBC, transmission electron microscopy
RBC

Platelet
 LEUKOCYTES

 According to the type of cytoplasmic granules and their

nuclear morphology, leukocytes are divided into two groups:
granulocytes and agranulocyte.
 Both types are rather spherical while suspended in blood
plasma, but they become amoeboid and motile after leaving
the blood vessels and invading the tissues.
 LEUKOCYTES CONT.…

 Granulocytes possess two major types of cytoplasmic granules

 1- lysosomes (often called azurophilic granules in blood
cells) .
 2 -specific granules that bind neutral, basic, or acidic stains
and have specific functions
 Granulocytes have polymorphic nuclei with two or more distinct
(almost separated) nuclear lobes and include the
neutrophils,eosinophils, and basophils.

22
 LEUKOCYTES CONT.…

 Agranulocytes do not have specific granules, but they do

contain azurophilic granules (lysosomes), with affinity for the
basic stain azure A.
 The nucleus is spherical or indented but not lobulated. This
group includes lymphocytes and monocytes
 LEUKOCYTES CONT.…

 All leukocytes are key players in the defense against invading

microorganisms, and in the repair of injured tissues.
 Specifically leaving the microvasculature in injured or
infected tissues slowly rolling leukocytes to express integrin
and other adhesion factors that produce firm attachment to the
endothelium .
 In a process called diapedesis (Gr. dia,through + pedesis, to
leap)
 LEUKOCYTES CONT.…

 The leukocytes send extensions through the openings between

the endothelial cells, migrate out of the venules into the
surrounding tissue space,
 The attraction of neutrophils to bacteria involves chemical
mediators in a process of chemotaxis.
 The number of leukocytes in the blood varies according to
age, sex, and physiologic condition

25
 NEUTROPHILS (Polymorph nuclear leukocytes)

 Mature neutrophils constitute 54% to 62% of circulating

leukocytes;
 Circulating immature forms raise this value by 3% to 5%.
Neutrophils are 12-15 µm in diameter in blood smears, with
nuclei having two to five lobes linked by thin nuclear
extensions.
 The cytoplasmic granules of neutrophils provide the cells’s
functional activities and are of two main types.
 NEUTROPHILS (Polymorph nuclear leukocytes) CONT…

Azurophilic primary granules

Resemble lysosomes as large, dense vesicles and have a major
role in both killing and degrading engulfed microorganisms.
They contain proteases and antibacterial proteins, including the
following:
 Myeloperoxidase which generates hypochlorite and other
agents toxic to bacteria.
 Lysozyme, which degrades components of bacterial cell
walls
 Defensins, small cysteine-rich proteins that bind and
disrupt the cell membranes of many types of bacterial
microorganisms
 NEUTROPHILS (Polymorph nuclear leukocytes) CONT…

Specific secondary granules

They are smaller and less dense, stain faintly pink,
They have diverse functions, including secretion of various
ECM-degrading enzymes such as collagenases, of additional
bactericidal proteins to the phagolysosome and insertion of new
cell membrane components.
Neutrophils are short-lived cells with a half-life of 6 to 8 hours
in blood and a life span of 1 to 4 days in connective tissues
before dying by apoptosis.
APPLICATION
 MEDICAL APPLICATION

 Neutrophils look for bacteria to engulf by pseudopodia and

internalize them in vacuoles called phagosomes
 Immediately thereafter, specific granules fuse with and
discharge their contents into the phagosomes which are then
acidified by proton pumps.
 Several kinds of neutrophil defects, often genetic in origin, can
affect function of these cells.EDICAL

29
 Neutrophil,
transmission
electron
micrograph

TEM appearance:
Multi-lobed nucleus
and numerous
specific granules and
lysosomes
(=azurophilic
granules in LM).

Specific
granule
Lysosome
(=azurophilic granule)
 EOSINOPHILS

 Eosinophils are far less numerous than neutrophils,

constituting only 1% to 3% of leukocytes.
 In blood smears, this cell is about the same size as a neutrophil
or slightly larger, but with a characteristic bi-lobed nucleus.
 The main identifying characteristic is the abundance of large,
acidophilic specific granules typically staining pink or red.
 Eosinophils are particularly abundant in connective tissue of
the intestinal lining and at sites of chronic inflammation such
as lung tissues of asthma patients
 MEDICAL APPLICATION

 An increase in the number of eosinophils in blood

(eosinophilia) is associated with allergic reactions and
helminth parasitic infections.
 In patients with such conditions, eosinophils are found in the
connective tissues underlying epithelia of the bronchi,
gastrointestinal tract, uterus, and vagina, and surrounding any
parasitic worms present.
 Eosinophil in a human blood smear

University of Michigan Virtual Slide Collection

LM appearance in smear: About 10-14 µm in diameter. Bilobed nucleus. The cytoplasm has prominent pink/red specific
granules (stained with eosin dye). If the smear is not stained properly, the granules may be brownish.
 BASOPHILS

 Basophils are also 12 to 15 µm in diameter but make up less

than 1% of blood leukocytes and are therefore difficult to find
in normal blood smear.
 The nucleus is divided into two irregular lobes
 But the large specific granules overlying the nucleus usually
obscure its shape. The specific granules (0.5 µm in diameter)
typically
 BASOPHILS cont…

 Stain purple with the basic dye of blood smear stains

 And are fewer, larger, and more irregularly shaped
 Basophils and mast cells also are central to immediate or
type1 hypersensitivity.

35
 MEDICAL APPLICATION

 Basophils and mast cells may rapidly degranulate, producing

vasodilation in many organs a sudden drop in blood pressure
 Other effects comprising a potentially lethal condition called
anaphylaxis or anaphylactic shock.
 Upon subsequent exposure, the allergen combines with the
receptor-bound IgE molecules,
 Release of the inflammatory mediators in this manner can
result in bronchial asthma, cutaneous hives, rhinitis,
conjunctivitis, or allergic gastroenteritis.
Comparison of basophil and eosinophil in a blood smear

Eosinophil

Basophil

.
 LYMPHOCYTES

 The most numerous type of agranulocyte in normal blood smears or

CBCs, lymphocytes constitute a family of leukocytes with spherical
nuclei.
 Lymphocytes can be subdivided into functional groups by
distinctive surface molecules (called “cluster of differentiation” or
CD markers)
 Major classes include B lymphocytes, helper and cytotoxic T
lymphocytes (CD4+ and CD8, respectively), and natural killer
(NK) cells.
 LYMPHOCYTES CONT.…

 Lymphocytes vary in life span according to their specific

functions;
 Some live only a few days and others survive in the circulating
blood or other tissues for many year.

39
 MEDICAL APPLICATION

 Lymphomas are a group of disorders involving neoplastic

proliferation of lymphocytes or the failure of these cells to
undergo apoptosis.
 Although often slow-growing, all lymphomas are considered
malignant because they can very easily become widely spread
throughout the body.
TISSUE PLASMA CELLS (DERIVED FROM B-
LYMPHOCYTES)
 MONOCYTES

 Monocytes are agranulocytes that are precursor cells of

macrophages, osteoclasts, microglia, and other cells of the
mononuclear phagocyte system in connective tissue.
 All monocyte-derived cells are antigen-presenting cells and
have important roles in immune defense of tissues.
 The monocyte nucleus is large and usually distinctly indented
or C-shaped .
 MONOCYTES CONT.…

 Circulating monocytes have diameters of 12 to 15 µm, but

macrophages are somewhat larger .
 The cytoplasm of the monocyte is basophilic and contains
many small lysosomes or Azurophilic granules.
 some of which are at the limit of the light microscope’s
resolution. These granules are distributed through the
cytoplasm, giving it a bluish-gray color in stained smears.
 Mitochondria and small areas of rough ER are present, along
with a Golgi apparatus involved in the formation of
lysosomes.

43
 MEDICAL APPLICATION

 Acute inflammation is usually short-lived as macrophages

undergo apoptosis or leave the site but chronic inflammation
usually involves the continued recruitment of monocytes.
 The resulting continuous presence of macrophages can lead to
excessive tissue damage that is typical of chronic
inflammation.
 MONOCYTE IN A BLOOD SMEAR

LM appearance in smears: About 16 µm in smears, thus the largest

leukocyte. Large, eccentric nucleus either oval, kidney-shaped or
horseshoe-shaped, with delicate chromatin that is less dense than
that of lymphocytes. Pale cytoplasm, often grayish, may contain
occasional stained granules .
MONOCYTE, TRANSMISSION ELECTRON MICROSCOPY

Lysosome
(=azurophilic
Mitochondr
granule)
ion

Centriole

Golgi

Cytoplasm contains mitochondria and some small lysosomes.

 PLATELETS

 Blood platelets (or thrombocytes) are very small non

nucleated membrane-bound cell fragments only 2 to 4 µm in
diameter .
 Platelets originate by separation from the ends of cytoplasmic
processes extending from giant polyploid bone marrow cells
called megakaryocytes.
 Normal platelet counts range from 150,000 to 400,000/µL
(mm) of blood.
 PLATELETS CONT.…

 Circulating platelets have a life span of about 10 days.

 In stained blood smears, platelets often appear in clumps.
 Each individual platelet is generally discoid, with a very
lightly stained peripheral zone, the hyalomere,
 And a darker-staining central zone containing granules, called
granulomere.

48
 PLATELETS CONT.…

 Also in the hyalomere are two systems of membrane channels.

 An open canalicular system of the plasma membrane, which
may facilitate platelets’ uptake of factors from plasma.
 A much less prominent set of irregular tubular vesicles
comprising the dense tubular system is derived from the ER and
stores Ca ions.
 PLATELETS CONT.…

 The role of platelets in controlling blood loss and in wound

healing can be summarized as follows
 Primary aggregation: Disruptions in the micro vascular
endothelium, which are very common, allow the platelet
glycocalyx to adhere to collagen.
 Thus, a platelet plug is formed as a first step to stop
bleeding
 PLATELETS CONT.…

 Secondary aggregation: Platelets in the plug release a

specific adhesive glycoprotein and ADP, which induce further
platelet aggregation and increase the size of the platelet plug.
 Blood coagulation: During platelet aggregation,
 Fibrinogen from plasma, von Willebrand factor and other
proteins released from the damaged endothelium,
 And platelet factor 4 platelet granules promote the sequential
interaction (cascade)

51
 PLATELETS CONT.…

 Clot retraction:
 The clot that initially bulges into the blood vessel lumen
contracts slightly because of the interaction of platelet actin
and myosin.
 Clot removal Protected by the clot, the endothelium and
surrounding tunic are restored by new tissue , and the clot is
then removed, mainly dissolved by the proteolytic enzyme
plasmin
 PLATELETS CONT.…

 Formed continuously through the local action of plasminogen

activators from the endothelium on plasminogen from plasma.

MEDICAL APPLICATION
 Bleeding disorders result from abnormally slow blood
clotting.
 One such disease directly related to a defect in the platelets
is a rare autosomal recessive glycoprotein Ib deficiency.
 Summary
 The liquid portion of circulating blood is plasma.
 Red blood cells or erythrocytes , which make up the hematocrit
portion (~45%) of a blood sample, are enucleated , biconcave discs.
 White blood cells or leukocytes are broadly grouped as
granulocytes(neutrophils, eosinophils, basophils) or agranulocytes
(lymphocytes, monocytes).
 Platelets are small (2-4 μm) cell fragments derived from
megakaryocytes in bone marrow, with a marginal bundle of actin
filaments, alpha granules and delta granules.

54
 Platelets Light Microscopy

Platelets Electron Microscopy

Platelets in
aggregates
 HEMOPOIESIS

INTRODUCTION
 This is the process by which all blood cells
(=haematological system) are produced.
 Platelets, red blood cells, leukocytes of all sorts
 Mature blood cells have a relatively short life span, and
are constantly regenerated in the bone marrow.
 We make 5 x 1011 blood cells daily.
 This is accelerated when there is haematological stress
e.g. infection, need more leukocytes
e.g. high altitude, need more red cells
INTRO……

 In the earliest phase of human embryogenesis, blood cells

arise from the yolk sac mesoderm
 In the second trimester, hemopoiesis (also called
hematopoiesis ) occurs primarily in the developing liver, with
the spleen also playing a role.
 Skeletal elements begin to ossify and bone marrow develops
in their medullary cavities, so that, in the third trimester, bone
marrow increasingly becomes the major hemopoietic organ
 Site of Hematopoiesis
During fetal life After birth
1) Yolk sac: Active (red) BM:
2) Liver & Spleen:
In infancy & childhood
3) Bone marrow (BM):

In adult
 Stem cells, growth factors, & differentiation

 Stem cells are pluripotential cells capable of self-renewal.

Some of their daughter cells form specific, irreversibly
differentiated cell types, and other daughter cells remain stem
cells.
 Hematopoietic stem cells are isolated by using
fluorescence-labeled antibodies to mark specific cell-
surface antigens and a fluorescence-activated cell-sorting
(FACS) instrument.
 Stem cells…..

 Stem cells are also studied by means of experimental

techniques that permit analysis of hematopoiesis in vivo and in
vitro.
 In vivo techniques include injecting the bone marrow of
normal donor mice into lethally irradiated mice whose
hematopoietic cells have been destroyed.
 In these animals, the transplanted bone marrow cells develop
colonies of hematopoietic cells in the spleen.
.
 Stem cells…..

 In vitro techniques involve the use of a semisolid tissue

culture medium made with a layer of cells derived from bone
marrow stroma
 This medium creates favorable microenvironmental conditions
for hematopoiesis.
 Data from several experiments show that under these
favorable microenvironmental conditions, stimulation by
growth factors influences the development of the various types
of blood cells
 Pluripotential Hematopoietic Stem Cells
 It is believed that all blood cells arise from a single type of
stem cell in the bone marrow.
 Because this cell can produce all blood cell types, it is called a
pluripotential stem cell
 These cells proliferate and form one cell lineage that will
become lymphocytes (lymphoid cells) and
 Another lineage that will form the myeloid cells that develop
in bone marrow (granulocytes, monocytes, erythrocytes, and
megakaryocytes).
 Progenitor & Precursor Cells

 The pluripotent stem cells give rise to daughter cells with

restricted potentials called progenitor cells or colony-
forming units (CFUs), since they give rise to:
colonies of only one cell type when cultured or injected into a
spleen.
 There are four types of progenitors/CFUs:
1. Erythroid lineage of CFU-erythrocytes (CFU-E)
2. Thrombocytic lineage of CFU- megakaryocytes (CFU-Meg)
3. Granulocyte-monocyte lineage of CFU-granulocytes-
monocytes (CFU-GM)
4. Lymphoid lineage of CFU-lymphocytes of all types
(CFU-L ).
 Progenitor…..

 All four progenitor/CFUs produce precursor cells or blasts in

which the cells' morphologic characteristics begin to
differentiate, suggesting the mature cell types they
will become

 In contrast, stem and progenitor cells cannot be morphologically

distinguished and resemble large lymphocyte.
 Progenitor…..

 Stem cells divide at a rate only sufficient to maintain their

relatively small population.
 The rate of cell division is accelerated in progenitor and
precursor cells, and large numbers of differentiated, mature
cells are produced (3 x 109 erythrocytes and 0.85 x 109
granulocytes/kg/day in human bone marrow).
 Bone Marrow

 Under normal conditions, the production of blood cells by the

bone marrow is adjusted to the body's needs,
 increasing its activity several-fold in a very short time.
 Bone marrow is found in the medullary canals of long bones
and in the cavities of cancellous bones
 Two types of bone marrow have been described based on their
appearance on gross examination: red, or hematogenous,
bone marrow, whose color is produced by the presence of
blood and blood-forming cells; and
 Bone …..

 yellow bone marrow, whose color is produced by the

presence of a great number of adipose cells.
 In newborns, all bone marrow is red and is therefore active in
the production of blood cells.
 As the child grows, most of the bone marrow changes
gradually into the yellow variety.
 Under certain conditions, such as severe bleeding or hypoxia,
yellow bone marrow is replaced by red bone marrow.
 Bone …..

 Red bone marrow (is composed of a stroma (from Greek,

meaning bed), hematopoietic cords, and sinusoidal
capillaries.
 The stroma of bone marrow contains collagen types I and III,
fibronectin, laminin, and proteoglycans
 Laminin, fibronectin, and another cell-binding substance,
hemonectin, interact with cell receptors to bind cells to the
stroma.
 Maturation of Erythrocytes

 A mature cell is one that has differentiated to the stage at which

it has the capability of carrying out all its specific functions.
 The basic process in maturation is the synthesis of hemoglobin
and the formation of an enucleated, biconcave, small corpuscle,
the erythrocyte.
 During maturation, several major changes take place:
 Cell volume decreases, and the nucleoli diminish in size until
they become invisible in the light microscope
 Maturation …..

 The nuclear diameter decreases, and the chromatin becomes

increasingly more dense until the nucleus presents a pyknotic
appearance and is finally extruded from the cell.
There is a gradual decrease in the number of polyribosomes
(basophilia decreases), with a simultaneous increase in the amount of
hemoglobin (an acidophilic protein) within the cytoplasm.
 Mitochondria and other organelles gradually disappear
 Maturation …..

 The development of an erythrocyte from the first recognizable

cell of the series to the release of reticulocytes into the blood
takes approximately 7 days.
 The hormone erythropoietin and substances such as iron, folic
acid, and cyanocobalamin (vitamin B12 ) are essential for the
production of erythrocytes.
 Erythropoietin is a glycoprotein produced mainly in the
kidneys that stimulates the production of mRNA for globin,
the protein component of the hemoglobin molecule.
 Differentiation

 The differentiation and maturation of erythrocytes involve the

formation (in order) of proerythroblasts, basophilic
erythroblasts, polychromatophilic erythroblasts,
orthochromatophilic erythroblasts (normoblasts),
reticulocytes, and erythrocytes .
 The first recognizable cell in the erythroid series is the
proerythroblast. It is a large cell with loose, lacy chromatin
and clearly visible nucleoli; its cytoplasm is basophilic.
 Differentia…..

 The next stage is represented by the basophilic erythroblast ,

with a strongly basophilic cytoplasm and a condensed nucleus
that has no visible nucleolus
 The basophilia of these two cell types is caused by the large
number of polyribosomes involved in the synthesis of
hemoglobin.
 During the next stage, polyribosomes decrease, and areas of
the cytoplasm begin to be filled with hemoglobin.
 Differentia….. …….
 At this stage, staining causes several colors to appear in the
cell—the polychromatophilic (Gr. polys, many, + chroma,
color, + philein, to love) erythroblast.
 In the next stage, the nucleus continues to condense and no
cytoplasmic basophilia is evident, resulting in a uniformly
acidophilic cytoplasm—the orthochromatophilic (Gr. orthos,
correct, + chroma + philein) erythroblast
 Differentia….. …….

 At a given moment, this cell puts forth a series of cytoplasmic

protrusions and expels its nucleus.
 The expelled nucleus is engulfed by macrophages.
 The remaining cell still has a small number of polyribosomes
that, when treated with the dye brilliant cresyl blue, aggregate
to form a stained network.
 This cell is the reticulocyte, which soon loses its
polyribosomes and becomes a mature erythrocyte.
 Simplified erythropoiesis

bone marrow circulation

pro-
MEP erythroblast* reticulocyte** erythrocyte
erythroblast

EP EPEP
EP

dividing cells committed to non-dividing cells

erythroid lineage: requires EPO (etc)

Hb synthesis
*Thesuffix “-blast” indicates a large, proliferating cell.
**The reticulocyte nucleus is condensed and inactive.
 Granulopoiesis

 The maturation process of granulocytes takes place with

cytoplasmic changes characterized by the synthesis of a number
of proteins that are packed in two organelles: the azurophilic and
specific granules.
 These proteins are produced in the rough endoplasmic reticulum
and the Golgi complex in two successive stages.
 The first stage results in the production of the azurophilic
granules, which stain with basic dyes in the Wright or Giemsa
methods and contain enzymes of the lysosomal system.
 Granulopo….

 In the second stage, a change in synthetic activity takes place

with the production of several proteins that are packed in the
specific granules
 These granules contain different proteins in each of the three
types of granulocytes and are utilized for the various activities
of each type of granulocyte.
 Evidently, a shift in gene expression occurs in this process,
permitting neutrophils to specialize in bacterial destruction
and eosinophils and basophils to become involved in the
regulation of inflammation
 Granulopo….

 The myeloblast is the most immature recognizable cell in the

myeloid series.
 It has a finely dispersed chromatin, and nucleoli can be seen.
 In the next stage, the promyelocyte (L. pro, before, + Gr.
myelos, marrow, + kytos, cell) is characterized by its basophilic
cytoplasm and azurophilic granules.
 Granulopo…

 These granules contain lysosomal enzymes and myeloperoxidase.

 The promyelocyte gives rise to the three known types of
granulocyte.
 The first sign of differentiation appears in the myelocytes, in which
specific granules gradually increase in quantity and eventually
occupy most of the cytoplasm.
 These neutrophilic, basophilic, and eosinophilic myelocytes
mature with further condensation of the nucleus and a considerable
increase in their specific granule content.
 Granulopo…

 Before its complete maturation, the neutrophilic granulocyte

passes through an intermediate stage in which its nucleus has
the form of a curved rod (band cell).
 This cell appears in quantity in the blood after strong
stimulation of hematopoiesis
 Kinetics of Neutrophil Production

 The total time taken for a myeloblast to emerge as a mature

neutrophil in the circulation is about 11 days.
 Under normal circumstances, five mitotic divisions occur in
the myeloblast, promyelocyte, and neutrophilic myelocyte
stages of development
 Neutrophils pass through several functionally and
anatomically defined compartments
 Kinetics of Neutrophil……

 Functional compartments of neutrophils. (1) Medullary

formation compartment. (2) Medullary storage (reserve)
compartment. (3) Circulating compartment. (4) Marginating
compartment. The size of each compartment is roughly
proportional to the number of cells
Kinetics of Neutrophil……
 Kinetics of Neutrophil……

 The marginating compartment is composed of neutrophils

that are present in blood but do not circulate.
 These neutrophils are in capillaries and are temporarily
excluded from the circulation by vasoconstriction, or—
especially in the lungs—they may be at the periphery of
vessels, adhering to the endothelium, and not in the main
bloodstream.
 Kinetics of Neutrophil……

 The marginating and circulating compartments are of about

equal size, and there is a constant interchange of cells between
them.
 The half-life of a neutrophil in these two compartments is 6–7h.
 The medullary formation and storage compartments together
are about 10 times as large as the circulating and marginating
compartments.
 Kinetics of Neutrophil……

 Neutrophils and other granulocytes enter the connective tissues by

passing through intercellular junctions found between endothelial
cells of capillaries and postcapillary venules (diapedesis).
 The connective tissues form a fifth compartment for
neutrophils, but its size is not known.
 Neutrophils reside here for 1–4 days and then die by apoptosis,
regardless of whether they have performed their major function of
phagocytosis
 MEDICAL APPLICATION

 Changes in the number of neutrophils in the blood must be

evaluated by taking all these compartments into consideration.
 Thus, neutrophilia, an increase in the number of neutrophils in
the circulation, does not necessarily imply an increase in
neutrophil production.
 Intense muscular activity or the administration of epinephrine
causes neutrophils in the marginating compartment to move
into the circulating compartment, causing an apparent
neutrophilia even though neutrophil production has not
increased.
 MEDICAL ……
 However, glucocorticoids (adrenal gland hormones) increase
the mitotic activity of neutrophil precursors in the marrow and
increase the blood count of neutrophils.
 Neutrophilia may also result from liberation of greater
numbers of neutrophils from the medullary storage
compartment.
 This type of neutrophilia is transitory and is followed by a
recovery period during which no neutrophils are released.
 MEDICAL ……

 The neutrophilia that occurs during the course of bacterial

infections is due to an increase in production of neutrophils
and a shorter duration of these cells in the medullary
storage compartment.
 In such cases, immature forms such as band cells, neutrophilic
metamyelocytes, and even myelocytes may appear in the
bloodstream.
 MEDICAL ……

 The neutrophilia that occurs during infection is of longer

duration than the neutrophilia that occurs as a result of intense
muscular activity

 The appearance of large numbers of immature neutrophils

(band cells) in the blood is called a shift to the left and is
clinically significant, usually indicating bacterial infection.
 Maturation of Lymphocytes & Monocytes

 Study of the precursor cells of lymphocytes and monocytes

is difficult, because these cells do not contain specific
cytoplasmic granules or nuclear lobulation, both of which
facilitate the distinction between young and mature forms of
granulocytes.
 Lymphocytes and monocytes are distinguished mainly on the
basis of size, chromatin structure, and the presence of
nucleoli in smear preparations.
 Maturation….

 As lymphocyte cells mature, their chromatin becomes more

compact, nucleoli become less visible, and the cells
decrease in size.
 In addition, subsets of the lymphocyte series acquire
distinctive cell-surface receptors during differentiation that
can be detected by immunocytochemical techniques
 Lymphocytes

 Circulating lymphocytes originate mainly in the thymus and

the peripheral lymphoid organs (eg, spleen, lymph nodes,
tonsils).
 However, all lymphocyte progenitor cells originate in the
bone marrow.
 Some of these lymphocytes migrate to the thymus, where they
acquire the full attributes of T lymphocytes.
 Subsequently, T lymphocytes populate specific regions of
peripheral lymphoid organs.
 Lympho….
 Other bone marrow lymphocytes differentiate into B
lymphocytes in the bone marrow and then migrate to
peripheral lymphoid organs, where they inhabit and multiply
in their own special compartments
 The first identifiable progenitor of lymphoid cells is the
lymphoblast, a large cell capable of incorporating [3H]
thymidine and dividing two or three times to form
prolymphocytes.
 Lympho…

 Prolymphocytes are smaller and have relatively more

condensed chromatin but none of the cell-surface antigens that
mark prolymphocytes as T or B lymphocytes.
 In the bone marrow and in the thymus, these cells synthesize
cell-surface receptors characteristic of their lineage, but they
are not recognizable as distinct B or T lymphocytes in routine
histological procedures.
 Using immunocytochemical techniques makes the distinction
 Monocytes

 The monoblast is a committed progenitor cell that is almost

identical to the myeloblast in its morphologicalcharacteristics.
 Further differentiation leads to the promonocyte, a large cell up
to 18 μm in diameter) with a basophilic cytoplasm and a large,
slightly indented nucleus.
 The chromatin is lacy, and nucleoli are evident. Promonocytes
divide twice in the course of their development into monocytes.
 Mono….
 A large amount of rough endoplasmic reticulum is present, as
is an extensive Golgi complex in which granule condensation
can be seen to be taking place
 These granules are primary lysosomes, which are observed as
fine azurophilic granules in blood monocytes.
 Mature monocytes enter the bloodstream, circulate for about 8
h, and then enter the connective tissues, where they mature
into macrophages and function for several months.
 MEDICAL APPLICATION

 Abnormal bone marrow can produce diseases based on cells

derived from that tissue.
 Leukemias are malignant clones of leukocyte precursors. They
occur in lymphoid tissue (lymphocytic leukemias) and in bone
marrow (myelogenous and monocytic leukemias).
 In these diseases, there is usually a release of large numbers
of immature cells into the blood.
 MEDICAL…..

 The symptoms of leukemias are a consequence of this shift in

cell proliferation, with a lack of some cell types and excessive
production of others (which are often abnormal in function).
 The patient is usually anemic and prone to infection
 A clinical technique that is helpful in the study of leukemias and
other bone marrow disturbances is bone marrow aspiration.
 Origin of Platelets

 The megakaryoblast is 25 to 50 μm in diameter and has a large

ovoid or kidney-shaped nucleus with numerous nucleoli.
 The nucleus becomes highly polyploid (ie, it contains up to 30
times as much DNA as a normal cell) before platelets begin to
form.
 Erythropoietin (EPO: aka epoietin) produced by kidneys
specifically stimulates erythropoiesis
 Thrombopoiesis

bone marrow circulation

megakaryocyte platelets
MEP

requires TPO (etc)

Platelets (1,000s) bud off surface of megakaryocyte.

 Megakaryocytes

 The megakaryocyte is a giant cell (35–150 μm in diameter) with

an irregularly lobulated nucleus, coarse chromatin, and no visible
nucleoli.
 The cytoplasm contains numerous mitochondria, a well-
developed rough endoplasmic reticulum, and an extensive Golgi
complex.
 Platelets have conspicuous granules, originating from the Golgi
complex, that contain biologically active substances, such as
platelet-derived growth factor,
 Megakary……

 Fibroblast growth factor, von Willebrand's factor (which

promotes adhesion of platelets to endothelial cells), and platelet
factor IV (which stimulates blood coagulation).
 With maturation of the megakaryocyte, numerous invaginations
of the plasma membrane ramify throughout the cytoplasm,
forming the demarcation membranes.
 This system defines areas of a megakaryocyte's cytoplasm that
shed platelets, extruding them into the circulation.
 SUMMARY OF KEY POINTS

 Pluripotent stem cells for blood cell formation, or

hemopoiesis,occur in the bone marrow of children and adults.
 Progenitor cells, committed to forming each type of mature
blood cell, proliferate and differentiate within micro
environmental niches of stromal cells, other cells, and ECM
with specific growth factors.
 These progenitor cells are also known as colony-forming units
(CFUs).
 SUMMARY…….

 Red bone marrow is active in hemopoiesis; yellow bone

marrow consists mostly of adipose tissue
 Erythropoietic islands or cords within marrow contain the red
blood cell lineage: proerythroblasts, erythroblasts with
succeeding developmental stages called basophilic,
polychromatophilic,
and orthochromatophilic that reflect the cytoplasmic transition
from RNA-rich to hemoglobin-filled.
 SUMMARY…….

 At the last stage of erythropoiesis cell nuclei are extruded,

producing reticulocytes that still contain some polyribosomes
but are released into the circulation
 Granulopoiesis includes myeloblasts, which have large
nuclei and relatively little cytoplasm; promyelocytes, in
which lysosomal azurophilic granules are produced;
 myelocytes, in which specific granules for one of the three
types of granulocytes are formed
 SUMMARY…….

 metamyelocytes, in which the characteristic changes in

nuclear morphology occur.
 Immature neutrophilic metamyelocytes called band (stab)
cells are released prematurely when the compartment of
circulating neutrophils is deleted during bacterial infections
 SUMMARY…….

■ Megakaryocytes, large polyploid cells of red bone marrow,

produce:
platelets, or thrombocytes, by releasing them from the ends
of cytoplasmic processes called proplatelets.
■ All these formed elements of blood enter the circulation by
crossing the discontinuous endothelium of sinusoids in the
red marrow
• Haematopoiesis is a complex business but the
outlines are pretty clear.!!