CHAPTER 4

Leukocytic Disorders

1
 Learning Objectives

At the end of this session, students will be able

to:
• Describe the classification of leukocytic disorders
• Describe quantitative leukocytic disorders
• Describe qualitative/functional leukocytic disorders
• Identify the morphologic characteristics of normal and
abnormal WBC

2
 Outline

• Definition of terms
• Classification of leukocytic disorders
• Non malignant leukocytic disorders
• Malignant leukocytic disorders

3
 Definition of Terms
• Leucocytosis: an increased number of leucocytes in the
peripheral blood
• Cytopenia: reduction in the number of cells in the peripheral
blood, e.g. neutropenia, thrombocytopenia, pancytopenia
• Leukemoid reaction: leukemoid reaction a reactive condition
which simulates or closely resembles leukemia
• Leukaemia: a myeloid or lymphoid neoplasm, characterized by
circulating neoplastic cells but also encompassing similar cases
in which there are neoplastic cells in the bone marrow.
• Vacuole: a fluid-filled cavity within a cell which, in the case of
phagocytes, is formed by invagination of the surface membrane
• Toxic granulation: increased staining of neutrophil granules
occurring as a response to infection and inflammation but also as
a physiological change during pregnancy
4
 Definition of Terms Cont’d
• Auer rod: a rod-shaped crystalline structure derived from
primary granules, found in the cytoplasm of cells of granulocytic
and, less often, monocytic lineage, observed in acute myeloid and
monocytic leukemias
• Döhle body: a weakly basophilic inclusion within the cytoplasm
of a neutrophil composed of ribosomes toxic granulation
increased staining of neutrophil granules occurring as a response
to infection and inflammation but also as a physiological change
during pregnancy
• Malignant: very injurious; in relation to neoplasms, aggressive
and characterized by local invasion, distant spread (metastasis)
and cytological differences from the equivalent normal tissue

5
 Leukocyte disorders

• Classification: 1. Non-Malignant 2. Malignant

• 1. Non-malignant (Reactive) Quantitative Disorders
• Leukocytosis Vs Leukopenia
• Leukocytosis occurs mostly due to Neutrophilia because of
their predominance in the circulating blood

6
 Reactive Leukocyte disorders cont’d

Non-malignant leukocytosis may be caused by:

• Increased movement of immature cells out of the bone marrow
proliferation compartment

• Increased mobilization of cells from the maturation storage

compartment of the bone marrow to the peripheral blood

• Decreased movement of mature cells from the circulation into

tissues

7
 Reactive Neutrophilia

 Assess for toxic changes and multilineage abnormalities.

Toxic changes such as Toxic granulation, Dohle bodies,
prominent cytoplasmic Vacoules, may be found when
sepsis is present and in exceptional cases, intra- and
extracellular bacteria and fungi may be identified.
 Evaluate for bacterial infections
 Blood findings with clinical features & chemical analysis,
cytogenetic analysis if a neoplastic disorder is suspected as
the presumptive diagnostic consideration

8
 Quantitative Leukocyte Disorders
• Neutrophil leukocytosis
– Infections, drugs, inflammation
• Note: Leukemoid Reaction needs to be differentiated
from Chronic Myeloid Leukemia
– Leukocytosis with marked left shift of immature
neutrophils; bands, myelocyte, metamyelocyte
– Infection, other cancer, burn, hemolysis, hemorrhage
– Differential: chronic myelocytic leukemia (CML)
• Increase in granulocytes both, Toxic granulation
in LR, Dohle body, vaculation in granulocytes,
increased Leukocyte Alkaline Phosphatase (LAP)
score in LR
• Neutropenia: decrease in neutrophils
 Quantitative Leukocyte Disorders
• Eosinophilia
– Allergy, parasites, Chronic Myelocytic Leukemia (CML),
Hodgkin’s lymphoma
• Basophilia
– Myeloproliferative disorder (CML)
• Lymphocytosis
– Viral infection, autoimmune disease
– Chronic lymphocytic leukemia (CLL)
• Monocytosis
– Infection, autoimmune disease,
– Acute monocytic leukemia
• Lymphopenia
– Viral infection
 Morphological abnormalities of mature
granulocytes (especially neutrophils)
Frequently observed morphological abnormalities: are
acquired
• Toxic granulation
• Döhle bodies (Inclusion of RNA origin)
• Vacuolation
• Hypersegmentation
Rarely observed genetic disorders
• Pelger-Hu acquired Pelger–Huët anomaly (bi-lobed, or non-
segmented nuclei)-no significant functional abnormalities
• May-Hegglin anomaly (large blue cytoplasmic inclusion
resembling Döhle bodies)
• Chediak-Higashi syndrome (giant cytoplasmic granules)
• Alder-Reilly inclusions 11
Reactive WBC disorders

• Morphology of neutrophils A
in infection
A. Toxic granulation C
B. Döhle bodies
C. Cytoplasmic
vacuolization

B
 Septicemia

Peripheral blood film of a patient with

Septicemia showing a Döhle body in a
neutrophil.
 Morphological changes
• Hypersegmented neutrophil (B12,
Folatic Acid deficiency)
• Hyposegmented, bilobed
– Pelger-Huet anomaly
• Inherited Autosomal Dominant,
asymptomatic
• Acquired Pelger-Heut-(e.g.
Myelodysplastic syndrome
(MDS)
• drugs
Acquired Pelger–Huët anomaly
Fig. 3.64 Peripheral
blood film of a patient
a with the
Acquired Pelger–Huët
anomaly as part of c
therapy-related
myelodysplastic
syndrome showing:
b (a)neutrophil with non-
lobed nucleus;
(b) anisocytosis,
poikilocytosis and
C neutrophil with bilobed
nucleus.
(c) neutrophil with
 Inherited Leukocyte
Disorders
• May Hegglin anomaly
– pale inclusion (Dohle body),
thrombocytopenia, giant
platelet
• Alder-Reilly
– Numerous granules, violet
color(round/comma)
– muccopolysaccharidoses
• Chediak-Higashi
– albinism, infection, bleeding,
– Defect in chemotaxis,
phagocytosis, giant granules
– Lysosomal inclusions
Peripheral blood film of a patient with the
Chédiak–Higashi syndrome showing a
neutrophil with giant and abnormally staining
granules.
 Fig. 3.56 Peripheral blood film of
a healthy subject showing a
normal polymorphonuclear
neutrophil and normal small
lymphocyte. The disposition of
the nuclear lobes around the
circumference of a circle
is apparent.

Peripheral blood film of a

healthy female showing a
normal neutrophil with a
drumstick
 Reactive or Atypical lymphocyte . Note the
abundant cytoplasm that “hugs” erythrocytes
 Large granular lymphocyte.

Large granular lymphocytes: A small number of

lymphocytes in normal blood are slightly larger than
resting lymphocytes, with reddish-purple (azurophilic)
granules. This appearance generally corresponds to
natural killer (NK) cells
Activated T- lymphocytes as seen in
Infectious Mononucleosis
HEMATOLOGIC MALIGNANCIES

22
Objectives

At the end of this chapter, the student shall be

able to:
 Define hematologic malignancies
 Explain the mechanisms of malignant transformation
 Describe the classification of leukemia
 Explain the diagnostic methodologies for the leukemias
 Describe myelodysplastic syndromes
 Define malignant lymphoma
 Characterize multiple myeloma
 Describe myeloproliferative disorders

23
Introduction
 Hematological malignancies are clonal diseases that
derive from a single cell in the marrow or peripheral
lymphoid tissue which has undergone a genetic
alteration
 A combination of genetic and environmental factors
determine the risk of developing malignancy:
 Inherited factors – genetic diseases increase the
incidence of leukemia
Down’s syndrome, Bloom’s syndrome, Fanconi’s
anemia, ataxia telangiectasia
 Environmental influences
Chemicals, drugs, radiation, infection
24
The genetics of malignant transformation
 Malignant transformation - accumulation of genetic
mutations in cellular genes
 The genes involved in the development of cancer are
divided broadly into two groups:
 Oncogenes
Arise because of gain-of-function mutations in
normal cellular genes called proto-oncogenes
Oncogenic versions are generated when the
activity of proto-oncogenes is increased or they
acquire a novel function through:
Translocation
Mutation
Duplication
 Tumour suppressor genes
May acquire loss-of-function point mutation or
deletion leading to malignant transformation
25
Leukemia
 The leukemias are a group of disorders characterized by
the accumulation of abnormal white cells in the bone
marrow
 These abnormal cells may cause:
 Bone marrow failure
 A raised circulating white cell count
 Infiltration of organs
 Thus common but not essential features include:
 Abnormal white cells in the peripheral blood
 A raised total white cell count
 Evidence of bone marrow failure (e.g. anemia,
neutropenia, thrombocytopenia) in the acute leukemias
 Involvement of other organs (e.g., liver, spleen, lymph
nodes, meninges, brain, skin or testes) 26
Leukemia cont’d

 Although viruses cause several forms of leukemia in

animals, their role in humans is uncertain
 Only two viral associations are identified
Epstein-Barr virus, a DNA virus, associated with
Burkitt's lymphoma
Human T-cell lymphotropic virus type I, called
human T-cell leukemia/lymphoma virus, an RNA
retrovirus, associated with some T-cell leukemias
and lymphomas
 Exposure to ionizing radiation and certain chemicals
(e.g., benzene, some anti-neoplastic drugs) is
associated with an increased risk of leukemia
27
Leukemia cont’d

 Some genetic defects (e.g., Down syndrome, Fanconi's

anemia) also predispose to leukemia
Classification of leukemia
 The main classification is into acute and chronic
leukemia
 On the basis of morphology and cytochemistry, acute
leukemia is further subdivided into:
 Acute myeloid (myeloblastic/myelogenous) leukemia
(AML)
 Acute lymphoblastic (lymphocytic) leukemia (ALL)
 AML is further subdivided into eight variants on a
morphological basis according to the French-American-
British (FAB) scheme (M0 – M7)
28
Classification of Leukemia cont’d

 ALL is subdivided on a morphological basis according to

the French-American-British (FAB) classification into L1,
L2, and L3
 The chronic leukemias comprise two main types:
 Chronic myeloid leukemia (CML)
 Chronic lymphocytic (lymphatic) leukemia (CLL)
 Other chronic types include:
 myeloproliferative
 Hairy cell leukemia
 Lymphoproliferative leukemia
 Various leukemia/lymphoma syndromes
29
Leukemias FAB classification

30
 Cont...
– Comparison of acute and chronic leukemias:
Acute Chronic
Age all ages usually adults
Clinical onset sudden insidious
Course (untreated) 6 mo. or less 2-6 years
Leukemic cells immature >20% blasts more mature cells
Anemia prominent (known) mild
Thrombocytopenia prominent mild
WBC count variable increased
Lymphadenopathy mild present;often
Splenomegaly mild present;often,prominent

31
 The FAB classification system AL
M1 Myelogenous Blasts & Promyelocyte without further maturation
M2 Myelogenous Myelogenous cells with maturation beyond blast
and promyelocyte stage
M3 Promyelocytic Promyelocytes predominate in the bone marrow
M4 Myelomonocyitic Both myelogenous and monoctic cells are present (at least 20% of
the total leukocytes)
M5 Monocytic Most cells monocytic
M6 Erythroleukemia Known as Di Guglielmo syndrome; abnomal proliferation of both
erythroid and granulocytic precursors; may include abnormal
megakaryocytic and monocytic proliferations
M7 Megakaryocytic Large and small megakaryoblast with a high nuclear-cytoplasm ratio;
pale agranular cytoplasm
L1 Homogenous One population of cells
Small cells predominant;
Nuclear shape is regular with an occasional cleft; Chromatin pattern
is homogenous &
Nucleoli are rarely visible; Cytoplasm moderately basophilic
L2 Heterogenous Large cells
Nuclear shape irregular; cleft in the nucleus common
1 large Nucleoli, Cytoplasm varies in colour
L3 Burkitt’s Cells are large and homogenous in size
lymphoma type Nuclear shape round or oval;
1-3 prominent nucleoli
Cytoplasm is deeply basophilic; often with prominent vacuoles32
The Acute Leukemias
 The leukemic cell population in ALL and AML
probably result from clonal proliferation by successive
divisions form a single abnormal stem or progenitor
cell
 There are over 20% myeloblasts or lymphoblasts in the
bone marrow at clinical presentation, and these blast
cells fail to differentiate normally but are capable of
further divisions
Replacement of the normal hemopoietic precursor
cells of the bone marrow by myeloblasts or
lymphoblasts and, ultimately in bone marrow
failure
33
The Acute Leukemias cont’d

 The clinical condition of the patient can be correlated

with the total number of leukemic cells in the body
When the abnormal cell number approaches 1012
the patient is usually gravely ill with severe bone
marrow failure
 Peripheral blood involvement by the leukemic cells
and infiltration of organs such as the spleen, liver and
lymph nodes may not occur until the leukemic cell
population comprised 60% or more of the marrow
cell total
34
The Acute Leukemias cont’d
 The disease may be recognized by conventional
morphology only when blast (leukemic) cells in the
marrow exceed 5% of the cell total unless the blast
cells have some particular abnormal feature(e.g. Auer
rods in myeloblasts and monoblasts).
 The clinical presentation and mortality in acute
leukemia arises mainly from:
Neutropenia
Thrombocytopenia, and
Anemia because of bone marrow failure
Organ infiltration (e.g. meninges or testes ,less
common) 35
The Acute Leukemias cont’d

 The acute leukemias comprise over half of the

leukemias seen in clinical practice.
 ALL is the common form in children
Its incidence is highest at 3-4 years
Falls off by 10 years
There is a lower frequency of ALL after 10 years
of age with a secondary rise after the age of 40
 AML occurs in all age groups
It is the common form of acute leukemia in adults
including the elderly
36
Laboratory Features in Acute Leukemias
 A normochromic normocytic anemia
 The total white cell count variable may be decreased,
normal or increased up to 200x109/l or more
 Thrombocytopenia in most cases, often extreme in
AML
 Blood film examination typically shows variable
numbers of blast cells
In AML, the blasts my contain Auer rods and
other abnormal cells may be present, e.g.,
promyelocytes, myelocytes, agranular neutrophils,
pseudo-Pelger cells or myelomonocytic cells
 ALL must be differentiated from infectious
37
mononucleosis and other causes of lymphocytosis
Auer rods may be seen in AML (shown
by arrows)

38
Laboratory features of Acute Myeloid
Leukemias cont’d
 In AML M6 (erythroleukemia) many erythroblasts may be
found and these may also be seen in smaller numbers in
other forms. Megaloblastic changes may also be seen in
bone marrow
 The bone marrow is hypercellular with a marked
proliferation of leukemic blast cells which amount to
over 50% and typically over 75% of the marrow cell total
 In ALL the marrow may be difficult to aspirate because of
increased reticulum fiber
 In AML M7 the patient typically has an acute onset of
pancytopenia with marrow fibrosis

39
 L1 ALL
Note: Nucleoli present in all six
lymphoblasts

Note: Peripheral blood film of patient with ALL-L1. Please

note nucleoli(lightly stained area in the nucleus) in
lymphoblasts
40
 L2 ALL

Peripheral blood film from patient with ALL L2

note: large irregularly shaped lymphoblasts

41
L3 ALL Leukemic Counterpart of Burkitt’s
Lymphoma

Peripheral blood film from patient with ALL L3

please note vacuoles in extremely basophilic cytoplasm
of lymphoblasts 42
AML M0

Peripheral blood film from patient with AML M0

43
AML M1

Peripheral blood film from patient with AML M1

note: please note Auer rods present in the blast in center
of the field.
44
AML M2

Peripheral blood film from patient with AML M2

45
AML M3

Peripheral blood film from patient with AML M3

Multiple Auer Rods may be seen in the cytoplasm of the
46
Promyelocytic cells
AML M4

Peripheral blood film from patient with AML M4

note: both myeloblasts and monoblasts are usually present

47
AML M5

Peripheral blood film from patient with AML M5

48
AML M6

Bone marrow film from patient with AML M6

49
50
Blast cells from a patient with acute leukemia that
have been accidentally transferred to the blood film
of another patient by the use of an inadequately
cleaned spreader.
Acute Plasmacytic Leukemia

Peripheral blood film from patient with acute plasmacytic

leukemia
52
Differentiation of ALL from AML

 In most cases, the clinical features and morphology on

routine staining separate ALL from AML
 In ALL the blasts show no differentiation (with the
exception of B-cell ALL)
 In AML some evidence of differentiation to granulocytes
or monocytes is seen in the blasts or their progeny
 Special test (e.g., cytochemistry, gene rearrangement
studies and chromosome analysis) are needed when the
cells are undifferentiated to:
 Confirm the diagnosis of AML or ALL, and
 Subdivide cases of AML or ALL into their different
subtypes
53
 Cytochemical Reactions in Acute
Leukemias
Type MPO SBB CAE NSE PAS AP

AML + + + + +/- +/-

(M4, M5) (M5, M6, M7)

ALL - - - - +/- +

MPO=Myeloperoxidase; SBB=Sudan black B; CAE=Chloroacetate esterase;

NSE=nonspecific esterase (using α-naphtol acetate or α- naphtol butyrate as
substrates to identify monocytic leukemias, do not react with myeloblasts);
PAS=periodic acid Schiff; AP=acid phosphatase
(MPO have best sensitivity & specificity compared to SBB & CAE)
54
Differentiation of ALL from AML cont’d

PAS PAS POSITIVE PEROXIDASE NEGATIVE

POSITIVE L1 L2 ALL POSITIVE L1 ALL
ALL

55
AML M2 PEROXIDASE

56
 Myeloproliferative Disorders
 A group of conditions characterized by clonal
proliferation of one or more hemopoietic components in
the bone marrow and, in many cases, the liver and
spleen
 Polycythemia vera (PV)
 Essential thrombocythemia
 Myelofibrosis with myeloid metaplasia
 Chronic Myeloid Leukemia

57
The Chronic Leukemias

Chronic Myeloid Leukemia (CML)

 One of the Myeloproliferative disorders
 Comprises <20% of all the leukemias and is seen most
frequently in middle age
 In over 95% of patients there is a replacement of normal
bone marrow by cells with an abnormal chromosome- the
Philadelphia or Ph chromosome
 This is an abnormal chromosome 22 due to the
translocation of part of a long (q) arm of chromosome
22 to another chromosome, usually 9, with translocation
of part of chromosome 9 to chromosome 22(t(9:22))

58
Conti....

59
Chronic Myeloid Leukemia (CML) cont’d
 It is an acquired abnormality of hemopoietic stem cells
that is present in all dividing:
 Granulocytic cells
 Erythyroid cells
 Megakaryocytic cells in the marrow
 And also in some B-cell lymphocytes and probably a
minority of T lymphocytes
 A great increase in total body granulocyte mass is
responsible for most of the clinical features
 In at least 70% of patients there is a terminal
metamorphosis to acute leukemia (myeloblastic or
lymphoblastic) with an increase of blast cells (blast
crisis) in the marrow to 50% or more

60
Chronic Myeloid Leukemia (CML) cont’d

 It occurs in either gender (male: female = 1.4:1), most

frequently between the ages of 40 and 60 years
 It may occur in children and neonates and in the elderly
 In most cases there are no predisposing factors but the
incidence was increased in survivors of the atom bomb
exposures in Japan
Laboratory findings in CML
 Leukocytosis is usually >50x109/l and sometimes
>500x109/l
 A complete spectrum of myeloid cells is seen in the
peripheral blood
 The levels of neutrophils and myelocytes exceed those
of blast cells and promyelocytes 61
Laboratory findings in CML cont’d

 Philadelphia (Ph) chromosome on cytogenetic analysis

of blood or bone marrow
 Hypercellular bone marrow with granulopoietic
predominance
 Leukocyte alkaline phosphatase score is invariably low
 Increased circulating basophils
 Normochromic, normocytic anemia is usual
 Platelet count may be increased (most frequently),
normal or decreased
 Serum vitamin B12 and vitamin B12-binding capacity are
increased
 Serum uric acid is usually raised
62
 Comparison of Reactive Neutrophilia
with CML
Parameter Reactive Neutrophilia CML
WBC Usually <30 x 109/L Usually >50 x 109/L

Toxic granulation Present Usually absent

Left shift Includes myelocytes Includes blasts

Basophilia Absent Present

Platelet count Variable, with sepsis Increased

NRBC Absent Present

Splenomegaly Absent Present
Fever Usually present Usually absent
Uric acid* Normal Increased
LAP** Increased Low except in Pts with 2o infection
Karyotype Normal Philadelphia chromosome***
*a chemical indicator of increased cell turnover
**LAP =Leukocyte Alkaline Phosphatase, is 2 o granules’ cytolytic enzyme. Primary granules contain
numerous cytolytic enzymes and the most notable one is myeloperoxidase
***Philadelphia chromosome (Ph1) = a translocation involving chromosomes 22 and 9. It occurs63 in the
precursors and megakaryocytes of patients with CML
Polycythemia vera (PV)
 Stem cell disorder, a clonal malignancy with excessive
production of mature hematopoietic cells, especially
RBCs
 Refers to a pattern of blood cell changes that includes:
 An increase in hemoglobin above 17.5g/dl in adult
males and 15.5g/dl in females
 An accompanying rise in red cell count (above 6.0 x
1012/l in males and 5.5x1012/l in females)
 Hematocrit (above 55% in males and 47% in females)
 The increase in red cell volume is caused by
endogenous myeloproliferation
 The stem cell origin of the defect is shown in many
patients by an over-production of granulocytes and
platelets as well as of red cells
 This is a disease of older subjects with equal incidence 64
in males and females
Laboratory findings in PV

 The hemoglobin, hematocrit and red cell count are

increased
 A neutrophil leukocytosis is seen in over half the
patients, and some have increased circulating basophils
 Increased platelet count is present in about half the
patients
 The leukocyte alkaline phosphatase (LAP) score is
usually increased above normal
 Increased serum vitamin B12 and vitamin B12-binding
Capacity

65
Laboratory findings in PV cont’d

 The bone marrow is hypercellular with increased

presence of megakaryocytes
Best assessed by a trephine biopsy
Clonal cytogenetic abnormalities may occur,
but there is no single characteristic change
 Circulating erythroid progenitors are increased
and grow in vitro independently of added
erythropoietin
 Blood viscosity is increased
 Plasma urea is often increased
66
Essential Thrombocythemia

 Megakaryocyte proliferation and overproduction of

platelets is the dominant feature of this condition
 There is sustained increase in platelet count above
normal (400x109/l)
 Recurrent hemorrhage and thrombosis are the principal
clinical features
 Splenic enlargement is frequent in the early phase but
splenic atrophy due to platelets blocking the splenic
mirocirculation is seen in some patients
 There may be anemia due to:
 Iron deficiency from chronic gastrointestinal or uterine
hemorrhage
 The marrow disorder itself 67
Laboratory findings in Essential
thrombocythemia

 Abnormal large platelets and megakaryocyte fragments

may be seen in the blood film

 The bone marrow picture is similar to that of

Polycythemia Vera

 Platelet function tests are consistently abnormal

68
Myelofibrosis with Myeloid Metaplasia

 There is the gradual replacement of the bone marrow by

connective tissue
 A prime feature is extramedullary hematopoieis
 Patients will typically have an enlarged spleen and liver
 Typically affects patients more than 50 years old
Laboratory findings in myelofibrosis
 Anemia is usual but a normal or increased hemoglobin
level may be found in some patients
 The white cell and platelet counts are frequently high at
the time of presentation
 Later in the disease leukopenia and
thrombocytopenia are common
69
Laboratory findings in myelofibrosis
cont’d
 A leuko-erythroblastic blood film is found
 The red cells show characteristic tear-drop poikilocytes
 Bone marrow is usually unobtainable by aspiration
 Trephine biopsy may show a hypercellular marrow with an
increase in reticulin-fibre pattern
 Low serum and red cell folate levels, raised serum vitamin B 12
and vitamin B12-binding capacity, and an increased leukocyte
alkaline phosphatase (LAP) score are usual
 High serum uric acid, LDH and hydroxybutyrate
dehydrogenase levels reflect the increased but largely
ineffective turnover of hemopoietic cells
 Transformation to acute myeloid leukemia occurs in 10-20%
of patients
70
 Lymphoproliferative neoplasma
chronic lymphocytic leukemia.
Hair cell leukemia.
Hemophagocytic lymphohistiocytosis (HLH)
lymphomas.
multiple myeloma

71
Chronic lymphocytic leukemia
 It accounts for 25% or more of the leukemias seen in
clinical practice
 It occurs in older subjects and is rare before 40 years
 The male to female ratio is 2:1
 The accumulation of large numbers of lymphocytes to
50-100 times the normal lymphoid mass in the blood,
bone marrow, spleen, lymph nodes and liver may be
related to immunological non-reactivity and excessive
lifespan
 The cells are a monoclonal population of B
lymphocytes
 With advanced CLL:
 Bone marrow failure
 A tumorous syndrome with generalized discrete
lymphadenopathy 72
Chronic lymphocytic leukemia cont’d

 Immunological failure results from reduced humoral

and cellular immune processes with a tendency to
infection
Laboratory findings in CLL
 Lymphocytosis
 The absolute lymphocyte count is >5 x 109/l and may
be up to 300x109/l or more
 Between 70% and 99% of white cells in the blood film
appear as small lymphocytes
 Smudge or smear cells are also present

73
Laboratory findings in CLL

 Normocytic normocytic anemia is present in later

states due to marrow infiltration or hypersplenism
 Autoimmune hemolysis may also occur
 Thrombocytopenia occurs in many patients
 Bone marrow aspiration shows lymphocytic replacement
of normal marrow elements
 Lymphocytes comprise 25-95% of all the cells
 Reduced concentrations of serum immunoglobulins
 More marked with advanced disease
 Rarely a paraprotien is present

74
 CLL

Peripheral blood film from patient with CLL

75
Hairy Cell Leukemia
• Also known as leukemic reticuloendotheliosis B/c of
marked marrow reticulin fibrosis, the marrow
frequently is difficult or impossible to aspirate
 It is a slow growing leukemia
 It is most common in older white males
 It is an unusual disease of peak age 40-60 years
 Men are affected nearly four times as frequently as
women
 It is a type of chronic lymphoid leukemia
 The disease is characterized clinically by features of
Pancytopenia
 The spleen may be moderately enlarged, but lack
lymphadenopathy. 76
Hairy Cell Leukemia cont’d

 There is a monoclonal proliferation of cells with an

irregular cytoplasmic outline (‘hairy’ cells, a type of B
lymphocyte) in:
 The peripheral blood
 Bone marrow
 Liver and other organs
 The number of hairy cells in the peripheral blood is
variable
 The bone marrow trephine shows a characteristic
appearance of mild fibrosis and a diffuse cellular infiltrate
 A serum paraprotein may be present and patients may
have arthritis, serositis or vasculitis
77
Hairy Cell Leukemia

Peripheral blood film from patient with Hairy Cell Leukemia

78
Malignant Lymphomas

 This group of diseases is divided into:

 Hodgkin’s disease
 Non-Hodgkin’s lymphomas
 In both, there is replacement of normal lymphoid
structure by collections of abnormal cells
 Hodgkin’s lymphoma is characterized by the presence
of Reed-Sternberg (RS) cells
 The non-Hodgkin’s lymphomas are characterized by
diffuse or nodular collections of abnormal
lymphocytes or, rarely, histiocytes

79
Hodgkin’s Disease

 It is a type of lymphoma characterized by the presence

of Reed-Sternberg cells (see Figure)
 In many patients, the disease is localized initially to a
single peripheral lymph node region and its subsequent
progression is by contiguity within the lymphatic system
 RS cells and the associated abnormal and smaller
mononuclear cells are neoplastic and the associated
inflammatory cells represent a hypersensitivity response
by the host
 After a variable period of containment within the lymph
nodes, the natural progression of the disease is to
disseminate to involve non-lymphatic tissue

80
Hodgkin’s disease cont’d

 The disease can present at any age but is rare in

children
 It has bimodal age incidence
 First peak in young adults (age 20-30 years)
 A second after the age of 50
 In developed counties the ratio of young adults to child
cases and of nodular sclerosing disease to other types is
increased
 There is an almost 2:1 male predominance

81
Laboratory findings in Hodgkin’s disease

 Normochromic, normocytic anemias is most common

 One-third of patients have a leucocytosis due to a
neutrophil increase
 Eosinophilia is frequent
 Advanced disease is associated with lymphopenia
 The platelet count is normal or increased during early
disease, and reduced in later stages
 The ESR is usually raised and is useful in monitoring
disease progress

82
Laboratory findings in Hodgkin’s disease
cont’d

 Bone marrow involvement is unusual in early disease

 It may be demonstrated by trephine biopsy, usually in
patients with disease at many sites
 There is progressive loss of immunologically competent
T lymphocytes with reduced cell-mediated immune
reactions
 Infections are common, particularly:
 Herpes zoster
 Cytomegalovirus
 Fungal, e.g., Cryptococcus and Candida
 Tuberculosis may occur
83
Laboratory findings in Hodgkin’s disease
cont’d
 Patients with bone disease may show:
 Hypercalcemia
 Hypophosphatemia
 Increased levels of serum alkaline phosphatase
 Serum lactate dehydrogenase (LDH) is raised initially in
30-40% of cases an indicates a poor prognosis
 Elevated levels of serum transaminase may indicate liver
involvement
 Serum bilirubin may be raised due to biliary obstruction
caused by large lymph nodes at the bile duct
 Hyperuricemia may occur
84
 Reed-Sternberg Cell

Reed-Sternberg cell present in the bone marrow of a

patient with Hodgkin’s lymphoma
85
Non-Hodgkin’s lymphomas
 The clinical presentation and natural history of these
malignant lymphomas are more variable than in
Hodgkin’s disease
 Pattern of spread is not regular
 A greater proportion of patients present with extra nodal
disease or leukemic manifestations
Laboratory findings in Non-Hodgkin’s lymphoma
 A normochromic normocytic anemia is usual but auto-
immune hemolytic anemia may also occur
 In advance disease with marrow involvement there may
be neutropenia, thrombocytopenia especially if the
spleen is enlarged or leuko-erythroblastic features

86
Laboratory findings in non-Hodgkin’s
lymphomas cont’d
 Lymphoma cells (also referred to as cleaved follicular
lymphoma or blast cells) with variable nuclear
abnormalities may be found in the peripheral blood in
some patients
 Trephine biopsy of marrow shows focal involvement,
usually paratrabecular(Lymphoid aggregate/infiltrate)in
20% of cases
 Elevation of serum uric acid may occur
 Abnormal liver function tests suggest disseminate
disease
 The serum LDH level is raised in more rapidly
proliferating and extensive disease and may be used as
87
a prognostic marker
Multiple Myeloma

 A neoplastic monoclonal proliferation of bone marrow

plasma cells characterized by:
 Lytic bone lesions.
 Plasma cell accumulation in the bone marrow, and
 The presence of monoclonal protein in the serum and
of urine or both
 98% of cases occur over the age of 40 with a peak
incidence in the seventh decade of life

88
Laboratory finding in multiple myeloma
 In 98% of patients monoclonal protein occurs in the serum or
urine or both
 The serum paraprotein is IgG in two-thirds
 IgA in one-third
 Rare IgM or IgD or mixed cases
 Normal serum immunoglobulins (IgG, IgA and IgM) are
depressed
 The urine contains Bence-Jones protein in two-thirds of cases
 The bone marrow shows increased plasma cells often with
abnormal forms referred to as myeloma cells
 Immunological testing shows these cells to be monoclonal
B cells
 Express the same immunoglobulin heavy and light chains
as the serum monoclonal protein
89
Laboratory finding in multiple myeloma
cont’d
 Usually a normochromic, normocytic or macrocytic
anemia
 Rouleaux formation is marked in most cases
 Neutropenia and thrombocytopenia occur in advanced
disease
 Abnormal plasma cells appear in the blood film in 15% of
patients
 Leuko-erythroblastic changes are occasionally seen
 High ESR
 Serum calcium elevation occurs in 45% of patients

90
Laboratory finding in multiple myeloma
cont’d
 The blood urea is raised above 14mmol/l and serum
creatinine raised in 20% of cases
 Proteinaceous deposits from heavy Bence-Jones
proteinuria, hypercalcaemia, uric acid, amyloid and
pyelonephritis may all contribute to renal failure
 A low serum albumin occurs with advance disease
 Serum β2-microglobulin (the light chain of the HLA class
1 antigens) is a useful indicator of prognosis
 It partly reflects renal function
 Levels less than 4mg/l imply a relatively good
prognosis.
91
Myelodysplastic Syndromes
(MDS)/Myelodysplasia
 A heterogeneous group of disease states that usually
present as peripheral blood cytopenias with a
hypercellular bone marrow
 Most common in the elderly and males are more
commonly affected
 There is a tendency to progress to acute myeloid
leukemia, although death often occurs before this
develops
 The fundamental disorder is the clonal proliferation of
stem cells that produce progeny that fail to mature
normally
 The maturation defect is more subtle; mature forms
develop but they are often morphologically atypical
(“dysplastic”) and frequently dysfunctional as well 92
Myelodysplastic Syndromes
(Myelodysplasia) cont’d
 The MDS are classified into five subgroups:
 Refractory anemia (RA)
( any anaemia that does not respond to therapy)
 RA with ring sideroblasts (RARS)
 RA with excess blasts (RAEB)
 RAEB in transformation (RAEB-t)
 Chronic myelomonocytic leukemia (CMML)

93
 FAB cooperative group criteria for myelodysplastic syndrome
subtypes

Monocyte RS-blasts,
BM blasts, PB blasts,
Type Auer rods s >15% of
percent percent
>1000/µL NRBCs
RA <5 ≤1 No No No
RARS <5 ≤1 No No Yes
RAEB 5-20 <5 No No ±
CMML ≤20 <5 No Yes ±
RAEB-T* 21-30 OR ≥5 ± ± ±

FAB: French-American-British; RA: refractory anemia; RARS: refractory anemia with

ringed sideroblasts; RAEB: refractory anemia with excess blasts; CMML: chronic
myelomonocytic leukemia; RAEB-T: refractory anemia with excess blasts in
transformation.
Myelodysplastic Syndromes
(Myelodysplasia) cont’d
Laboratory features of MDS
 Peripheral blood
 Pancytopenia is a frequent finding
 The red cells are usually macrocytic or dimorphic but
occasionally hypochromic; normoblasts may be
present
 The reticulocyte count is low (CRC<1%
 Granulocytes are often reduced
Show lack of granulation
Their chemotactic, phagocytic and adhesive
functions are impaired
95
Laboratory features cont’d

 The Pelger Huet-type abnormality (single or bilobed

nucleus in neutrophils)) is often present
 In CMML, monocytes are >1.0x109/l in the blood
 The total white blood count may be >100x109/l with left
shift
 The platelets may be unduly large or small and are
usually decreased in number but in 10% of cases are
elevated
 Leukoerythroblastic blood picture
 Variable numbers of myeloblasts in blood indicate poor
prognosis

96
Laboratory features cont’d

Bone marrow
 The cellularity is usually increased
 Ring sideroblasts may occur in all five FAB types
 Multinucleate normoblasts and other dyserythropoietic
features are seen
 The granulocyte precursors
 Show defective primary and secondary granulation,
and
 Cells which are difficult to identify as either agranular
myelocytes, monocytes or promonocytes are frequent
 Megakaryocytes are abnormal with micro, small
binuclear or polynuclear forms
 Bone marrow biopsy shows fibrosis in 10% of cases 97
Review Questions

1. Briefly describe the classification of leukemia

2. Explain the laboratory diagnosis of different types of
leukemia
3. Define myelodysplastic syndrome (MDS) and indicate
the hematological findings associated with MDS
4. What is the distinguishing morphological characteristic of
Hodgkin’s lymphoma?
5. What are the laboratory findings of patients with multiple
myeloma?
6. Define myeloproliferative disorders

98
 Bibliography
• MA Lichtman, E Beutler, U Seligsohn, K Kaushansky,
TO Kipps (Editors). William’s Hematology. 7th Ed.
McGraw-Hill Co. Inc. 2008.
• Dacie, John V and Lewis, S.M. Practical Hematology 10th
Edition Churchill-Livingstone 2006.
• Wintrobe, Maxwell M. Clinical Hematology 11th Edition
Lea and Febiger, Philadelphia 2003.

99