sub module 5 (granulocytes disorders)

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
http://en.wikipedia.org/wiki/Granulocyteshttp://en.wikipedia.org/wiki/Granulocytes


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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.

sub module 5 (granulocytes disorders)

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