Leukocytes constitute the cellular components of the innate and adaptive immune system and are critical for host defense. These cells mediate acute and chronic inflammation, modulate immune responses, and protect the host against numerous pathogens. Disorders affecting leukocytes can be divided broadly into malignant disorders (tumors of leukocytes or their progenitors) and non-malignant disorders. The malignant disorders are uncommon but clinically important entities Non- malignant leukocyte disorders can involve any any of the leukocytes (neutrophils, eosinophils, basophils, monocytes, B cells, T cells, and natural killer cells) but the disorders of greatest clinical relevance affect neutrophils; these will be our major focus.

1. Leukocyte Disorders
Asst. Prof. Aşkın K. KAPLAN M.D.
Maltepe University Faculty of Medicine

2. Leukocyte Disorders
• Leukocytes constitute the cellular components of the innate and
adaptive immune system and are critical for host defense. These
cells mediate acute and chronic inflammation, modulate immune
responses, and protect the host against numerous pathogens.
• Disorders affecting leukocytes can be divided broadly into
malignant disorders (tumors of leukocytes or their progenitors) and
non-malignant disorders.
• The malignant disorders are uncommon but clinically important
entities
• Non- malignant leukocyte disorders can involve any any of the
leukocytes (neutrophils, eosinophils, basophils, monocytes, B cells,
T cells, and natural killer cells)
• but the disorders of greatest clinical relevance affect neutrophils;
these will be our major focus.

3.  eosinophilia
 basophilia
 monocytosis
 leukemoid reaction
 leukemia
 leukocytosis  leukopenia
 neutrophilia  neutropenia
 lymphocytosis  lymphopenia
Leukocyte disorders

4. WHAT HAPPENS IN BONE MARROW?
• Neutrophils, red cells and megakaryocytes all descended
from common precursor cell called myeloid stem cell
• Stem cells differentiate into primitive cells called blasts,
which are precursors of each cell type
• Blasts divide and mature under the influence of proteins
called growth factors; as they mature they lose the ability to
divide
• Mature neutrophils and red cells enter the blood
• Megakaryocytes break into small fragments (platelets),
which enter blood

5. Hematopoiesis
Mature blood cells differentiate from pluripotent hematopoietic stem cells.

6. Granules in Neutrophils: Primary granules: promyelocyte stage , Secondary granules: myelocyte stage and predominate therafter
Myelopoiesis
Myelopoiesis: As granulocytes develop and mature, they go through
a series of recognizable morphologic stages that correlate with the
expression of genes that confer the specific functions indicated on
the time line.
HSC, hematopoietic stem cell; PMN, polymorphonuclear leukocyte.
Life span of neutrophil in Blood is 6-10 hours

7. Morphology of mature leukocytes in Periferic Blood (PB)

8. myeloblast
neutrophil
promyelocyte myelocyte
Band form
metamyelocyte
eosinophil
basophil
Myelopoiesis.

9. Distribution of Macrophages

10. Granulopoiesis Neutrophil kinetics
G-CSF : Granulocyte Colony-Stimulating Factors, SCF: Stem Cell Factor, IL-3:Interleukin 3,
GM-CSF: Granulocyte-macrophage colony-stimulating factor

11. myeloblast
neutrophil
promyelocyte myelocyte
Band form
metamyelocyte
eosinophil basophil
monoblast
monocyte lymphoblast lymphocyte

12. Normal white cell counts

13. Polymorphonuclear Leukocytes
• Polymorphonuclear leukocytes, also called
granulocytes because their cytoplasm contains
granules, include
– Neutrophils
– Eosinophils
– Basophils
• Polymorphonuclear leukocytes occur in the
circulation and have multilobed nuclei.

14. Morphology of mature leukocytes in Periferic Blood (PB)

15. Neutrophils
• Neutrophils constitute 40 to 70% of total circulating WBCs;
they are a first line of defense against infection. Mature
neutrophils have a half-life of about 2 to 3 days.
• During acute inflammatory responses (eg, to infection),
neutrophils, drawn by chemotactic factors and alerted by the
expression of adhesion molecules on blood vessel
endothelium, leave the circulation and enter tissues.
• Their purpose is to phagocytose and digest pathogens.
• Microorganisms are killed when phagocytosis generates lytic
enzymes and reactive oxygen compounds and triggers release
of granule contents.

16. Eosinophils
• Eosinophils are granulocytes derived from the same
progenitor cells as monocytes-macrophages,
neutrophils, and basophils. They are a component of
the innate immune system. Eosinophils have a
variety of functions, including
– Defense against parasitic infections
– Defense against intracellular bacteria
– Modulation of immediate hypersensitivity reactions

17. Eosinophils
• Eosinophils constitute up to 5% of circulating WBCs.
• They target organisms too large to be engulfed; they
kill by secreting toxic substances (eg, reactive oxygen
compounds similar to those produced in neutrophils),
major basic protein (which is toxic to parasites),
eosinophil cationic protein, and several enzymes.
• Eosinophils are also a major source of inflammatory
mediators (eg, prostaglandins, leukotrienes, platelet-
activating factor, many cytokines).

18. Eosinophil production and function
• Eosinophil production appears to be regulated by
T-Cells through the secretion of the hematopoietic
growth factors:
– granulocyte-macrophage colony-stimulating factor (GM-
CSF),
– interleukin-3 (IL-3), interleukin-5 (IL-5).
– Although GM-CSF and IL-3 also increase the production of
other myeloid cells, IL-5 increases eosinophil production
exclusively.

19. Eosinophil production and function
• Eosinophil granules contain major basic protein and
eosinophil cationic protein;
– toxic to several parasites and to mammalian cells.
– These proteins bind heparin and neutralize its anticoagulant
activity.
• Eosinophil-derived neurotoxin can severely damage
myelinated neurons.
• Eosinophil peroxidase, generates oxidizing radicals in
the presence of hydrogen peroxide and a halide.
– Charcot-Leyden crystals are primarily composed of
phospholipase B and are located in sputum, tissues, and stool
in disorders in which there is eosinophilia
(eg, asthma, eosinophilic pneumonia).

20. Normal peripheral blood
eosinophil count
• Diurnal levels vary inversely with plasma cortisol levels;
the peak occurs at night and the trough in the morning.
• The eosinophil count can decrease with stress, with the
use of beta-blockers or corticosteroids, and sometimes
during bacterial or viral infections.
• The count can increase (eosinophilia) in allergic
disorders, during certain infections (typically parasitic),
and as a result of numerous other causes.
• The circulating half-life of eosinophils is 6 to 12 h, with
most eosinophils residing in tissues (eg, the upper
respiratory tract, GI tract, skin, uterus).

21. Normal peripheral blood
eosinophil count
• Generally accepted that a count > 500/mcL is
elevated.
• Peripheral eosinophilia is characterized as
– Mild: 500 to 1500/mcL
– Moderate: 1500 to 5000/mcL
– Severe: > 5000/mcL

22. Basophils
• Basophils constitute < 5% of circulating WBCs and
share several characteristics with mast cells
• Both have high-affinity receptors for IgE called Fc-
epsilon RI (FcεRI).
• When these cells encounter certain antigens, the
bivalent IgE molecules bound to the receptors become
cross-linked, triggering cell degranulation with release
of preformed inflammatory mediators (eg, histamine,
platelet-activating factor) and generation of newly
synthesized mediators (eg, leukotrienes,
prostaglandins, thromboxanes).

23. Monocytes
• Monocytes in the circulation are precursors to tissue
macrophages.
• Monocytes migrate into tissues, where over about 8 h,
they develop into macrophages under the influence of
macrophage colony-stimulating factor (M-CSF),
secreted by various cell types (eg, endothelial cells,
fibroblasts).
• At infection sites, activated T cells secrete cytokines
(eg, interferon-gamma [IFN-gamma]) that induce
production of macrophage migration inhibitory factor,
preventing macrophages from leaving.

24. Macrophages
• Macrophages are activated by IFN-gamma and
granulocyte-macrophage colony-stimulating factor
(GM-CSF).
• Activated macrophages kill intracellular organisms and
secrete IL-1 and tumor necrosis factor-alpha (TNF-
alpha).
• These cytokines potentiate the secretion of IFN-gamma
and GM-CSF and increase the expression of adhesion
molecules on endothelial cells, facilitating leukocyte
influx and destruction of pathogens.
• Based on different gene expression profiles, subtypes
of macrophages (eg, M1, M2) have been identified.

25. Lymphocytes
• The 2 main types of lymphocytes are
– B cells (which mature in bone marrow)
– T cells (which mature in the thymus)
• They are morphologically indistinguishable but have
different immune functions.
• They can be distinguished by antigen-specific surface
receptors and molecules called clusters of
differentiation (CDs), whose presence and absence
define some subsets.
• More than 300 CDs have been identified
• Each lymphocyte recognizes a specific antigen via
surface receptors.

26. B cells
• About 5 to 15% of lymphocytes in the blood are B
cells; they are also present in the bone marrow,
spleen, lymph nodes, and mucosa-associated
lymphoid tissues.
• B cells can present antigen to T cells and release
cytokines, but their primary function is to
develop into plasma cells, which manufacture
and secrete antibodies.
• Patients with B-cell immunodeficiencies (eg, X-
linked agammaglobulinemia) are especially
susceptible to recurrent bacterial infections.

27. T cells
• T cells develop from bone marrow stem cells
that travel to the thymus, where they go
through rigorous selection. There are 3 main
types of T cell:
• Helper
• Regulatory (suppressor)
• Cytotoxic

28. Helper T (Th) cells
• Helper T (Th) cells are usually CD4 but may be CD8. They
differentiate from Th0 cells into one of the following:
• Th1 cells: In general, Th1 cells promote cell-mediated
immunity via cytotoxic T cells and macrophages and are
thus particularly involved in defense against intracellular
pathogens (eg, viruses). They can also promote the
production of some antibody classes.
• Th2 cells: Th2 cells are particularly adept at promoting
antibody production by B cells (humoral immunity) and
thus are particularly involved in directing responses aimed
at extracellular pathogens (eg, bacteria, parasites).
• Th17 cells: Th17 cells promote tissue inflammation.

29. Regulatory (suppressor) T cells
• Regulatory (suppressor) T cells mediate suppression of
immune responses and usually express the Foxp3
transcription factor.
• The process involves functional subsets of CD4 or CD8
T cells that either secrete cytokines with
immunosuppressive properties or suppress the
immune response by poorly defined mechanisms that
require cell-to-cell contact.
• Patients with functional mutations in Foxp3 develop
the autoimmune disorder IPEX
syndrome (immunodysregulation, polyendocrinopathy,
enteropathy, X-linked syndrome).

30. Cytotoxic T (Tc) cells
• Cytotoxic T (Tc) cells are usually CD8 but may be CD4; they
are vital for eliminating intracellular pathogens, especially
viruses. Tc cells play a role in organ transplant rejection.
• Tc-cell development involves 3 phases:
• A precursor cell that, when appropriately stimulated, can
differentiate into a Tc cell
• An effector cell that has differentiated and can kill its
appropriate target
• A memory cell that is quiescent (no longer stimulated) but
is ready to become an effector when restimulated by the
original antigen-MHC combination
• Fully activated Tc cells, like NK cells, can kill an infected
target cell by inducing apoptosis.

31. Mast Cells
• Mast cells are tissue-based and functionally similar to
basophils circulating in the blood.
• Mucosal mast cell granules contain tryptase and
chondroitin sulfate; connective tissue mast cell
granules contain tryptase, chymase, and heparin. By
releasing these mediators, mast cells play a key role in
generating protective acute inflammatory responses;
basophils and mast cells are the source of type I
hypersensitivity reactions associated with atopic
allergy. Degranulation can be triggered by cross-linking
of IgE receptors or by the anaphylatoxin complement
fragments C3a and C5a.

32. Definitions White Cell Numbers
• Leukocytosis: increase in the numbers of circulating white
cells: >12,000/uL
• Leukopenia: decrease in the numbers of circulating white
cells: < 4,000/uL
• Left Shift – increased circulating numbers of immature
neutrophils
• Leukoerythroblastic Reaction – leukocytosis with a left
shift accompanied by nucleated red cells: seen in
malignancy.
• Leukemoid Reaction – benign excessive leukocytosis
accompanied by an exaggerated neutrophilia and a left
shift in response to an infection; the WBC > 50 x 109/L

33. Definitions White Cell Numbers
• Neutrophilia >7.5 x 109/L
• Other defining features:
– Left shift – Increased band forms
– “toxic” cell appearance
• Dohle bodies
• Vacuoles
• Intra-cellular microbes

34. Causes of Neutrophilia
• Infections
– (primarily bacterial)
• Drugs/Hormones
– Epinephrine
– corticosteroids
– lithium
• venoms/poisons/toxins
– Tissue necrosis
• acute gout
• burns
• trauma
• infarcts
Other
• autoimmune disorders
• stress
• severe physical activity
• pregnancy
• smoking
• acute hemorrhage
• post-splenectomy
• myeloproliferative disorders
• Metabolic
• Ketoacidosis
• uremia
• eclampsia
• thyrotoxicosis

35. Basophilia
• Basophilia Absolute count >0.15 x 109/L
Causes:
– CML
– Allergies
– inflammatory disorders
– İrradiation
– viral infections

36. Eosinophilia
• Eosinophilia Absolute count >0.5 x 109/L
• Causes:
– Parasites
– Helminths
– drug treatments
– Allergies
– İnfections
– Neoplasms
– Chronic myeloid leukemia (CML)
– autoimmune disorders

37. Monocytosis
Monocytosis Absolute count >0.8 x 109/L
Most commonly seen in conditions with increased
cell damage:
– Chronic infection [TB, syphilis, protozoal infections,
rickettsial infections]
– Recovery from agranulocytosis
– Post-splenectomy
– Strenuous exercise
– Subacute bacterial endocarditis

38. Lymphocytosis
Lymphocytosis Absolute count >5.5 x 109/L
Normally:
– 60-80% circulating lymphs are T-cells
– [2:1 CD4/CD8]
– 10-20% are B-lymphs
– 5-10% are natural killer or NK cells
Infections
Viral
Infectious mononucleosis
Bacterial
Pertussis
Thyrotoxicosis
Recovery from acute infections
Neoplasm
Leukemias
Lymphomas
Causes

39. Lymphocytosis
• Epstein-Barr Virus
– Infectious mononucleosis
Reactive/atypical/variant
lymphocytes

40. Infectious Mononucleosis
• Acute, self-limiting, febrile infection of B-cells
• Circulating reactive lymphocytes are primary CD8 T-cells
• Typically occurs in those age 10-25 years
– Fever
– Sore throat
– Lymphadenopathy
– Lethargy
Positive serology
Heterophile antibodies

41. Leukopenia
• is a reduction in the circulating WBC count to < 4000/μL. It is usually
characterized by a reduced number of circulating neutrophils, although a
reduced number of lymphocytes, monocytes, eosinophils, or basophils
may also contribute. Thus, immune function can be generally decreased.
• Neutropenia is a reduction in blood neutrophil count to < 1500/μL in
whites and < 1200/μL in blacks. It is sometimes accompanied by
monocytopenia and lymphocytopenia, which cause additional immune
deficits.
• Lymphocytopenia, in which the total number of lymphocytes is < 1000/μL
in adults, is not always recognized as a decrease in the total WBC count
because lymphocytes account for only 20 to 40% of the total WBC count.
The consequences of the lymphopenia can depend on the lymphocyte
subpopulation(s) that are decreased.
• Monocytopenia is a reduction in blood monocyte count to < 500/μL.
Monocytes migrate into the tissues where they become macrophages,
with specific characteristics depending on their tissue localization.

42. Neutropenia
Neutropenia < 1.5 x 109/L
Definition: less than the normal absolute count; greatly
influenced by patient age and race. African and Middle Eastern
populations Subclasses include mild, moderate and severe:
• Reactions to Drugs
• BM ablative therapy
• Infections
• HIV/Hepatitis
• Typhoid/ miliary TB
• Malaria
• Immune Disorders
• Systemic lupus erythromatous (SLE)
• Neoplasm
• BM Failure
• Megaloblastic Anemia
• Aplastic Anemia
• Hypersplenism
• Idiopathic (of unknown cause).
Causes

43. Neutropenia
• Neutropenia is absolute neutrophil count (ANC)
<1500 cells/microL
• Severe neutropenia is ANC <500/µL, or an ANC
that is expected to decrease to <500/µL over the
next 48 hours
ANC= WBC x (PMN % + Bands %)
Aşkın K. KAPLAN M.D.

44. Eosinopenia
• Eosinopenia is a form of agranulocytosis where the
number of eosinophil granulocytes in perepheral
blood is lower than expected < 40/cmm
– Bacterial infection
– By stress reactions
– Cushing's syndrome
– By the use of steroids
– Pathological causes include burns and acuteinfections.

45. Neutropenic Fever
• The Infectious Diseases Society of America
defines neutropenic fever as
– a single oral temperature of >38.3°C (101°F) or
– a temperature of >38.0°C (100.4°F) sustained for
>1 hour
Aşkın K. KAPLAN M.D.

46. Neutropenia Causes
• Defects inside or outside the Bone Marrow
– Decreased proliferation [failure of cells - aplasia]
– Decreased maturation [insufficient number of
precursors undergoing abnormal maturation]
– Decreased survival [increased destruction and/or
rapid removal of cells]
– Distribution [total body pools are normal,
circulating numbers are reduced]

47. Lymphopenia
• Lymphopenia Absolute lymphocyte count <0.6
x 109/L
• There are three types of abnormalities:
– Decreased production
– Increased destruction
– Changes in distribution

48. Lymphopenia
Decreased production
– SCID = severe combined
immunodeficiency
– Protein-calorie malnutrition
– Zinc deficiency
Increased destruction
– HIV infection
– Radiation therapy
– Neoplastic chemotherapy
– SLE
Redistribution
• Glucocorticoid therapy
• Anesthesia
• TB
• Influenza
• Burns
Other
• Hodgkin’s
• Myasthenia gravis

49. Monocytopenia
• is a reduction in blood monocyte count
to < 500/μL. Risk of certain infections is
increased. It is diagnosed by CBC with
differential. Treatment with hematopoietic
stem cell transplantation may be needed.

50. Qualitative WBC Disorders

51. Abnormal WBC

52. Altered Cell Morphology
• Nuclear
– Pelger-Huet
– Hypersegmentation
• Cytoplasmic
– Alder, Reilly, Chediak-Higashi
– May-Hegglin
– Gaucher , Niemann-Pick
– Mucopolysacchridoses
– Toxic Granulation
– Vacuolization
– Dohle Bodies
– Necrobiosis
– Peripheral Blood, Wright’s, 100x.
• Think Neutrophilia due to Severe Bacterial Infection

53. Pelger Huet vs band neutrophil
•Pelger Huet – an inherited condition
resulting in hyposegmentation of
granulocyte nuclei with increased density
and coarseness of the chromatin.. Don’t
confuse this anomaly with a neutrophilic
left shift!
•May-Hegglin - a rare syndrome
characterized by leukopenia, variable
thrombocytopenia, GIANT PLATELETS, and
gray-blue cytoplasmic inclusions in the
neutrophils and monocytes [dohle-like
bodies]

54. Alder-Reilly vs Chediak-Higashi
• Alder-Reilly - an inherited trait characterized
by the presence of abnormally large
azurophilic and basophilic granules
resembling neutrophilic toxic granulation.
• Chediak-Higashi - is a genetic disorder that
has an equivalent syndrome in mink, cattle,
mice, cats, & killer whales. Affected
individuals display partial albinism, are very
susceptible to common infectious agents, and
have white cells demonstrating giant
cytoplasmic granules.

55. Definitions
• Gaucher & Niemann-Pick are characterized by the lack of or
defective activity of enzymes.
• In Gaucher disease, there is a lack of beta-glucocerebrosidase
and macrophages become laden with glucocerebrosides.
• In Niemann-Pick, there is deficient activity of lysosomal
hydrolase and sphingomyelinase resulting in the accumulation
of cholesterol and sphingomyelin in mononuclear phagocytes.
• Mucopolysaccharidoses are a group of genetically determined
deficiencies of specific enzymes involved in the degradation of
mucopolysaccharides. Examples: Hurlers, Hunter, Sanfilippo

56. Pelger-Huet & Hypersegmentation
Bilobed and occasional
unsegmented neutrophils
Autosomal recessive disorder
Rare autosomal dominant condition
Neutrophil function is essentially normal
Pelger-Huet anomaly Neutrophil hyper-segmentation

57. May Hegglin
Neutrophils contain basophilic inclusions of RNA
Occasionally there is associated leucopenia, Thrombocytopenia
and giant platelet are frequent.

58. Alder Reilly
Alder’s anomaly
Granulocytes, monocytes and lymphocytes contain granules
which stain purple with Romanowsky stain
Granules contain mucopolysaccharides

59. Chediak Higashi
Autosomal recessive disorder
Giant granules in granulocytes, monocytes and lymphocytes
Partial occulocutaneous albinism, Depressed migration and degranulation ,
Recurrent pyogenic infections , Lymphoproliferative syndrome may develop
Treatment is BMT
BMT Bone Marrow Transplant

60. Mucopolysacharidoses

61. Toxic Granulation & Vacuolization

62. Dohle Bodies and Necrobiosis
Necrobiotic WBC displays nuclear
degradation or karyorrhexis. Indicates
cell death in chemotherapy or a poorly
preserved specimen.
Single or multiple blue cytoplasmic
inclusions. They represent remnants of
rough endoplasmic reticulum from earlier
maturational stages.
They are associated with myeloid "left
shifts" and are seen in conjunction with
toxic granulation.

63. Niemann Pick

64. gaucher

65. Lymphocytosis
Bordatella pertussis
Whooping Cough –BUTT CELL

66. Maltepe University Faculty of Medicine
Thank You