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7/28/2019 Sub Module 5 (Granulocytes Disorders)
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SUB MODULE 5:
GRANULOCYTES DISORDERS
TEACHER GUIDANCE
Contributors :
Budiman, dr., SpPK(K)
Budi Darmawan M, dr., SpPD-KHOM
Dr. Endang Sri Wahyuni, dr., MS
Setyohadi, drg., MS
Djoko Heri Hermanto, dr., SpPD
Susanto Nugroho, dr., SpA
FACULTY OF MEDICINEUNIVERSITY OF BRAWIJAYA
MALANG
2008
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SUB MODULE 5:GRANULOCYTE
DISORDERSI. DESCRIPTION
Definition andOverview
The white blood cells (leukocytes) may be divided into twobroad group: the phagocytes and the immunocytes.Granulocytes, which include three types of cell: neutrophils(polymorphs), eosinophils and basophils, together withmonocytes comprise the phagocytes.The function of phagocytes and immunocytes in protectingthe body against infection is closely connected with twosoluble protein systems of the body, immunoglobulins and
complement. These proteins, which may also be involved inblood cell destruction in a number of diseases.Granulopoiesis, the function of granulocytes, andgranulocytes disorders are discussed in this sub module.
Competency Area Area of competence : 3rd of the Doctor CompetenciesStandard from Indonesian Medical Council
CompetencyComponent
To apply the concept and principles of granulopoiesis, themorphology and the function of granulocytes, granulocytesdisorders, and also clinical approach to diagnose these.
Clinical Competence 1. Student can describe granulopoiesis2. Student can describe the morphology and the function of
granulocytes3. Student can describe causes of granulocytes disorders4. Student can do the clinical approach to diagnose causes of
granulocytes disorders
Learning Methode Active learning with modul task, group discussion, expertlecture, and skill development (history taking)
Equipment Classroom, worksheet, computer, LCD and screen
Time Active learning with modul task 3x50 minutes; groupdiscussion 2x50 minutes; expert lecture 1x50 minutes; andskill development (history taking) overall 3x50 minutes.
Lecturer 1. Budiman, dr., SpPK(K)2. Budi Darmawan M, dr., SpPD-KHOM
3. Dr. Endang Sri Wahyuni, dr., MS4. Setyohadi, drg., MS5. Djoko Heri Hermanto, dr., SpPD6. Susanto Nugroho, dr., SpA
Evaluation Middle exams at the end of modul programmes with multiplechoice questions, and final exams at the end of semesterwith OSCE.
Suggested Refferences 1. Hoffbrand AV, Pettit JE, Moss PAH. The White Cells 1:Granulocytes, Monocytes and their Benign Disorders. In:Essential Haematology. 4th ed. London: BlackwellScience. 2001.
2. Holland SM, Gallin JI. Disorders of Granulocytes and
Monocytes. In: Fauci AS, Kasper DL, Longo DL et al, eds.Harrisons Principles of Internal Medicine. 17th ed. NewYork: The McGraw-Hill Companies. 2008.
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II. LECTURE CONTENTS
OVERVIEW
The white blood cells (leukocytes) may be divided into two broad group: the phagocytes
and the immunocytes. Granulocytes, which include three types of cell: neutrophils (polymorphs),
eosinophils and basophils, together with monocytes comprise the phagocytes.
The function of phagocytes and immunocytes in protecting the body against infection is
closely connected with two soluble protein systems of the body, immunoglobulins and
complement. These proteins, which may also be involved in blood cell destruction in a number of
diseases. Granulopoiesis, the function of granulocytes, and granulocytes disorders are
discussed in this sub module.
GRANULOCYTES
Neutrophils (polymorph)
This cell has a characteristic dense nucleus consisting of between two and five lobes,
and a pale cytoplasm with an irregular outline containing many fine pink-blue (azurophilic) or
grey-blue granules (Fig.1.a). The granules are divided into primary, which appear at the
promyelocyte stage, and secondary (specific) which appear at the myelocyte stage and
predominate in the mature neutrophil. Both types of granule are lysosomal in origin; the primary
contains myeloperoxidase, acid phosphatase and other acid hydrolases, the secondary contains
collagenase, lactoferrin and lysozyme. The lifespan of neutrophils in the blood is only about 10
hours.
Neutrophil precursors
These do not normally appear in normal peripheral blood but are present in the marrow.
The earliest recognizable precursor is the myeloblast, a cell of variable size which has a large
nucleus with fine chromatin and usually two to five nucleoli. The cytoplasm is basophilic and nocytoplasmic granules are present. The normal bone marrow contains up to 4% of myeloblasts.
Myeloblasts give rise by cell division to promyelocytes which are slightly larger cells and have
developed primary granules in the cytoplasm. These cells then produce myelocytes which have
specific or secondary granules. The nuclear chromatin is now more condensed and nucleoli are
not visible. Separate myelocytes of the neutrophil, eosinophil, and basophil series can be
identified. The myelocytes give rise by cell division to metamyelocytes, non-dividing cells, which
have an idented or horseshoe-shaped nucleus and a cytoplasm filled with primary and
secondary granules. Neutrophil forms between the metamyelocyte and fully mature neutrophil
are termed band, stab or juvenile. These cells may occur in normal peripheral blood. They do
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not contain the clear fine filamentous distinction between nuclear lobes which is seen in mature
neutrophils.
Monocytes
These are usually larger than other peripheral blood leucocytes and possess a large
central oval or idented nucleus with clumped chromatin (Fig.1.b). The abundant cytoplasm stains
blue and contains many fine vacuoles, giving a ground-glass appearance. Cytoplasmic granules
are also often present. The monocyte precursors in the marrow (monoblasts and promonoblasts)
are difficult to distinguish from myeloblasts and monocytes.
(a) (b)
(c) (d)
Fig.1. White blood cells (leucocytes): (a) neutrophil (polymorph);(b) monocyte; (c) eosinophil; (d) basophil
Eosinophils
These cells are similar to neutrophils, except that the cytoplasmic granules are coarser
and more deeply red staining and there are rarely more than three nuclear lobes (Fig.1.c).
Eosinophil myelocytes can be recognized but earlier stages are indistinguishable from neutrophil
precursors. The blood transit time for eosinophils is longer than for neutrophils. They enter
inflammatory exudates and have a special role in allergic responses, defence against parasites
and removal of fibrin formed during inflammation.
Basophils
These are only occasionally seen in normal peripheral blood. They have many dark
cytoplasmic granules which overlie the nucleus and contain heparin and histamine (Fig.1.d). In
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the tissues they become mast cells. They have immunoglobulin E (IgE) attachment sites and
their degranulation is associated with histamine release.
GRANULOPOIESIS
Granulopoiesis (or granulocytopoiesis) is hematopoiesis of granulocytes. The blood
granulocytes and monocytes are formed in the bone marrow from a common precursor cell
(Fig.2). In the granulopoietic series progenitor cells, myeloblasts, promyelocytes and myelocytes
form a proliferative or mitotic pool of cells while the metamyelocytes, band and segmented
granulocytes, make up a postmitotic maturation compartment (Fig.3). Large numbers of band
and segmented neutrophils are held in the marrow as a reserve pool or storage compartment.
The bone marrow normally contains more myeloid cells than erythroid cell in the ratio of 2:1 to
12:1, the largest proportion being neutrophils and metamyelocytes. In the stable or normal state,
the bone marrow storage compartment contains 10-15 times the number of granulocytes found
in the peripheral blood. Following their release from the bone marrow, granulocytes spend only
6-10h in the circulation before moving into the tissues where they perform their phagocytic
function. In the bloodstream there are two pools usually of about equal size the circulating pool
(included in the blood count) and the marginating pool (not included in the blood count). It has
been estimated that they spend on average 4-5 days in the tissues before they are destroyed
during defensive action or as the result of senescence.
Fig.2. Diagrammatic representation of the bone marrow pluripotent stem celland the cell lines that arise from it
Control of granulopoiesis: myeloid growth factors
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The granulocytes series arises from bone marrow progenitor cells which are increasingly
specialized. Many growth factors are involved in this maturation process including interleukin-1
(IL-1), IL-3, IL-5 (for eosinophils), IL-6, IL-11, granulocyte-macrophag colony-stimulating factor
(GM-CSF), granulocyte CSF (G-CSF) and monocyte CSF (M-CSF) (Fig.4). The growth factors
stimulate proliferation and differentiation and also affect the function of the mature cells on which
they act (e.g. phagocytosis, superoxide generation and cytotoxicity in the case of neutrophils;
phagocytosis, cytotoxicity and production of other cytokines by monocytes).
Increased granulocyte and monocytes production in response to an infection is induced
by increased production of growth factors from stromal cells and T lymphocytes, stimulated by
endotoxin, IL-1 or tumour necrosis factor (TNF).
Fig.3. Neutrophil kinetics
THE FUNCTION OF GRANULOCYTES
The normal function of neutrophils and monocytes may be divided into three phases:
1. Chemotaxis (cell mobilization and migration)
The phagocyte is attracted to bacteria or the site of inflammation by chemotactic
substances released from damaged tissues or by complement components and also by the
interaction of leucocyte adhesion molecules with ligands on the damaged tissues.
2. Phagocytosis
The foreign material (bacteria, fungi, etc.) or dead or damaged cells of the host are
phagocytosed (Fig.5). Recognition of a foreign particle is aided by opsonization with
immunoglobulin or complement because both neutrophils and monocytes have Fc and C3breceptors. Opsonization of normal body cells (e.g. red cells or platelets) also makes them
liable to destruction by macrophages of the reticuloendothelial system, as in autoimmune
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hemolysis, idiopathic (autoimmune) thrombocytopenic purpura or many of the drug-induced
cytopenias.
Fig.4. A diagram of the role of growth factors in normal hemopoiesis.
Macrophages have a central role in antigen presentation processing and presenting
foreign antigens on human leucocyte antigen (HLA) molecules to the immune system. They
are also secrete a large number of growth factors which regulate inflammation and immune
responses.
Fig.5. Phagocytosis and bacterial destruction
Chemokines are chemotactic cytokines of which there are two main classes CXC ()
chemokines, small (8-10.000 MW) pro-inflammatory cytokines which mainly act on
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neutrophils, and CC () chemokines such as macrophage inflammatory protein (MIP)-1 and
RANTES which act on monocytes, basophils, eosinophils and natural killer (NK) cells.
Chemokines may be produced constitutively and control lymphocyte traffic under
physiological conditions; inflammatory chemokines are induced or up-regulated by
inflammatory stimuli. They bind to and activate cells via chemokine receptors and play an
important part in recruiting appropriate cells to the sites of inflammation. Chemokine
receptors have been identified as coreceptors for human immunodeficiency virus (HIV) entry
into cells.
3. Killing and digestion
This occurs by oxygen-dependent and oxygen-independent pathways. In the oxygen-
dependent reactions, superoxide (O2-), hydrogen peroxide (H2O2) and other activated
oxygen (O2) species, are generated from O2 and reduced nicotinamide adenine dinucleotide
phosphate (NADPH). In neutrophils, H2O2 reacts with myeloperoxidase and intracellular
halide to kill bacteria; activated oxygen may also be involves a fall in pH within phagocytic
vacuoles into which lysosomal anzymes are released. An additional factor, lactoferrin an
iron-binding protein present in neutrophil granules is bacteriostatic by depriving bacteria of
iron (Fig.5).
Defects of phagocytic cell function
1. Chemotaxis
These defects occur in rare congenital abnormalities (e.g. lazy leucocyte syndrome)
and in more common acquired abnormalities either of the environment, e.g. corticosteroid
therapy, or of the leucocytes themselves, e.g. in acute or chronic myeloid leukemia,
myelodysplasia and the myeloproliferative syndromes.
2. Phagocytosis
These defects usually arise because of a lack of opsonization which may be caused
by congenital or acquired causes of hypogammaglobulinemia or lack of complement
components.
3. Killing and digestion
This abnormality is clearly illustrated by the rare X-linked or autosomal recessive
chronic granulomatousdisease which results from abnormal leucocyte oxidative metabolism.
There is an abnormality affecting different elements of the respiratory burst oxidase or its
activating mechanism. The patients have reccuring infections, usually bacterial but
sometimes fungal, which present in infancy or early childhood in most cases.
Other rare congenital abnormalities may also result in defects of bacterial killing e.g.
myeloperoxidase deficiency and the Chediak-Higashi syndrome. Acute or chronic myeloid
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leukemia and myelodysplastic syndromes may also be associated with defective killing of
ingested microorganisms.
LEUCOCYTOSIS AND MONOCYTOSIS
Neutrophil leucocytosis
An increase in circulating neutrophils to levels greater than 7.5x10 9/l is one of the most
frequently observed blood count changes. The causes of neutrophil leucocytosis are given in
Table 1. Neutrophil leucocytosis is sometimes accompanied by fever as a result of the release of
leucocyte pyrogens. Other characteristic features of reactive neutrophilia may include:
a. A shift to the left in the peripheral blood differential white cell count, i.e. an increase in
the number of band forms and the occasional presence of more primitive cells such as
metamyelocytes and myelocytesb. The presence of cytoplasmic toxic granulation and Doehle bodies
c. An elevated neutrophil alkaline phosphatase (NAP) score
For this the strength of the staining of each of 100 neutrophils is scored between 0 and 4. The
maximum score is therefore 400; a normal score is between 20-100.
Table 1. Causes of neutrophil leucocytosis
Bacterial infection (especially pyogenic bacterial, localized or generalized)
Inflammation and tissue necrosis, e.g. myositis, vasculitis, cardiac infarct, traumaMetabolic disorders, e.g. uremia, eclampsia, acidosis, gout
Neoplasms of all types, e.g. carcinoma, lymphoma, melanoma
Acute hemorrhagic of hemolysis
Corticosteroid therapy (inhibits margination)
Myeloproliferative disease, e.g. chronic myeloid leukemia, polycythemia vera, myelosclerosis
Treatment with myeloid growth factors, e.g. G-CSF, GM-CSF
The leukemoid reaction
The leukemoid reaction is a reactive and excessive leucocytosis usually characterized bythe presence of immature cells (e.g. myeloblasts, promyelocytes and myelocytes) in the
peripheral blood. Occasionally lymphocytic reactions occur. Associated disorders include severe
or chronic infections, severe hemolysis or metastatic cancer. Leukemoid reactions are often
particularly marked in children. Granulocyte changes such as toxic granulation and Doehle bodie
and a high NAP score help to differentiate the leukemoid reaction from chronic myeloid leukemia
(in which the NAP score is low).
Eosinophilic leucocytosis (eosinophilia)
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The causes of an increase in blood eosinophils above 0.4x109/l are listed in Table 2. Sometimes
no underlying cause is found and if the eosinophil count is elevated (>1.5x109/l) for over 6
months and associated with tissue damage then the hypereosinophilic syndrome is diagnosed.
Table 2. Causes of eosinophilia
Allergic diseases, especially hypersensitivity of the atopic type, e.g. brochial asthma, hay fever, urticaria
and food sensitivity
Parasitic diseases, e.g. amoebiasis, hookworm, ascariasis, tapeworm infestation, filariasis,
schistosomiasis and trichinosis
Recovery from acute infection
Certain skin diseases e.g psoariasis, pemphigus and dermatitis herpetiformis
Pulmonary eosinophilia and the hypereosinophilic syndrome
Drug sensitivity
Polyarteritis nodosa
Hidgkins disease and some other tumoursMetastatic malignancy with tumour necrosis
Eosinophilic leukemia (rare)
Treatment with GM-CSF
Basophilic leucocytosis (basophilia)
An increase in blood basophils above 0.1x109/l is uncommon. The usual cause is a
myeloproliferative disorder such as chronic myeloid leukemia or polycythemia vera. Reactive
basophil increases are sometimes seen in myxoedema, during smallpox or chickenpox infection,and in ulcerative colitis.
Monocytosis
A rise in blood monocyte count above 0.8x109/l is infrequent. The conditions listed in Table 3
may be responsible.
Table 3. Causes of monocytosis
Chronic bacterial infections: tuberculosis, brucellosis, bacterial endocarditis, typhoid
Protozoan infections
Chronic neutropenia
Hodgkins disease and other malignancies
Myelodysplasia (especially chronic myelomonocytic leukemia)
Treatment with GM-CSF or M-CSF
NEUTROPENIA
The lower limit of the normal neutrophil count is 2.5x109/l. When the absolute neutrophil
level falls below 0.5x109/l the patient is likely to have recurrent infections and when the count
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falls to less than 0.2x109/l the risks are very serioud, particularly if there is also a functional
defect. Neutropenia may be selective or part of a general pancytopenia (Tabel 4).
Table 4. Causes of neutropenia
Selective neutropenia
Congenital
Kostmanns syndrome
Acquired
Drug induced
Anti-inflammatory drugs (aminopyrine, phenylbutazone)
Antibacterial drugs (chloramphenicol, co-trimoxazole, sulfaslazine, imipenem)
Anticonculsants (phenytoin, carbamazepine)
Antithyroids (carbimazole)
Hypoglicemics (tolbutamide)Phenothiazines (chlorpromazine, thioridazine)
Psychotropics and antidepressants (clozapine, mianserin, imipramine)
Bark derivatives, e.g. paclitaxil
Miscellaneous (glod, penicillamine, mepacrine, amodiaquine, ticlopidine, furosemide, etc)
Benign (racial or familial)
Cyclical
Immune
Autoimmune
Systemic lupus erythematosusFeltys syndrome
Hypersensitivity and anphylaxis
Large granular lymphocytic leukemia
Infections
Viral, e.g. hepatitis, influenza, HIV
Fulminant bacterial infection, e.g. typhoid, military tuberculosis
Part of general pancytopenia
Bone marrow failure
Splenomegaly
a. Congenital neutropenia
Kostmanns syndrome is an autosomal recessive disease presenting in the first year of
life with life-threatening infections. Most cases are due to mutation of the gene coding for
neutrophil elastase. G-CSF produces a clinical response although marrow fibrosis and acute
myeloid leukemia may supervene.
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b. Drug-induced neutropenia
A large number of drugs have been implicated (Table 4) and may induce neutropenia
either by direct toxicity or immune-mediated damage.
c. Cyclical neutropenia
This is a rare syndrome with 3-4 weeks periodicity. Severe but temporary neutropenia
occurs. Monocytes tend to rise as the neutrophils fall. Mutation of the gene for neutrophil
elastase underlies some cases.
d. Autoimmune neutropenia
In some cases of chronic neutropenia an autoimmune mechanism can be demonstrated.
The antibody may be directed against one of the neutrophil-specific antigens (NA, NB, etc.).
e. Idiopathic benign neutropenia
An increase in the marginating fraction of blood neutrophils and a corresponding
reduction in the circulating fraction is one cause of benign neutropenia.
Clinical features
Severe neutropenia is particularly associated with infections of the mouth and throat. Painful and
often intractable ulceration may occur at these sites (Fig.6), on the skin or the anus. Septicemia
rapidly supervenes. Organisms carried as commensals by normal individuals, such as
Staphylococcus epidermidis or Gram-negative organisms in the bowel, may become pathogens.
Diagnosis
Bone marrow examination is useful in determining the level of damage in granulopoiesis, i.e.
whether there is reduction in early precursors or whether there is reduction only of circulating
and marrow neutrophils with late precursors remaining in the marrow. Marrow aspiration and
trephine biopsy may also provide evidence of leukemia, myelodysplasia or other infiltration.
Fig.6. Ulceration of the tongue in severe neutropenia
Algorithms of leucocytosis, eosinophilia, and neutropenia (see Fig.7, 8 and 9)
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III. MODUL TASK
1. Describe in brief three types of granulocytes (neutrophils, eosinophils, basophils)
and monocytes !
2. Describe in brief the normal function of neutrophils and monocytes into three
phases !3. Describe in brief defects of phagocytic cell function !
IV. SUGGESTED READINGS
3. Hoffbrand AV, Pettit JE, Moss PAH. The White Cells 1: Granulocytes, Monocytes and theirBenign Disorders. In:Essential Haematology. 4th ed. London: Blackwell Science. 2001.
4. Holland SM, Gallin JI. Disorders of Granulocytes and Monocytes. In: Fauci AS, Kasper DL,Longo DL et al, eds. Harrisons Principles of Internal Medicine. 17 th ed. New York: TheMcGraw-Hill Companies. 2008.