BIO3CBH – White Blood

Cells and their disorders

Lecturer
Helen Irving
Intended learning outcomes
By the end of this topic, you should be able to:

•Distinguish different types of white cells in peripheral blood (consider size,

organelles, etc.)
•Describe the roles of different white cells and how they may be affected by
diseases and disorders
•Analyse how malfunctions in granulopoiesis, monocytopoiesis and lymphopoiesis
can contribute to disease states and white cell disorders
•Analyse blood results in relation to granulocytes, monocytes and lymphocytes and
disease states
•Explain how chronic conditions can affect haematological results – particularly
white cells
What do these results tell you about the blood of a patient?

CBC/FBE Do the white cell CBC results

• WBC 16.8 4.0 - 12 x 109/L correlate to the blood film?
• Neutrophils 15 1.8-7.5 x 109/L
• Lymphocytes 1.4 1.5 -3.5 x 109/L
• Eosinophils 0.3 0.04-0.4 x 109/L
• Hb 102 115-175 g/L
• MCV 86 80-95 fL
• Platelet count 1400 150-400 x 109 L
Image source: https://webpath.med.utah.edu/HEMEHTML/HEME006.html
BIO3CBH – Granulocyte Formation
(Focussing on Neutrophils)
Lecturer
Helen Irving

Basophil
Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
Leukocytes (white blood cells) in normal peripheral blood
Task: note some features.
Neutrophils (~ 1,500-8,000/ μL)

Eosinophils (~ 0-450 / μL)

Basophils (~ 0-200 / μL)

Monocytes (~ 200-950 / μL)

Blood film stained with Romanowsky type dyes (e.g. eosin

and methylene blue) to differentiate white cell features
Haemopoiesis

Source: Figure 1.2, Hoffbrand & Moss (2016) Hoffbrand’s essential Haematology 7th ed
 Growth factors responsible for haemopoiesis
Committed
progenitor cells Erythropoietin GM-CSF IL-3
Erythrocytes
IL-3 GM-CSF Thrombopoietin
Megakaryocyte Platelets

IL-1, IL-3, GM-CSF IL-3 M-CSF Monocytes

Monocyte-
IL-6, GM-CSF
granulocyte
GM-CSF, GM-CSF G-CSF IL-3
progenitor Neutrophils
Pluripotent SCF
stem cell GM-CSF IL-3 IL-5 Eosinophils

IL-3 Basophils

B lymphocytes

Interleukins (IL) 1, 2, 4, 6, 7, 12
Thymus T lymphocytes
SCF = stem cell factor, GM-CSF = granulocyte-monocyte colony stimulating factor, G-CSF = granulocyte colony stimulating factor, M-CSF = monocyte colony stimulating factor
 Stages of maturation of granulocytes

Key features distinguishing

myeloblast and mature
segmented cells include cell
size and the degrees of
development of the nucleus

Segmented and to a leeser

extent band neutrophils are
found in blood; the other
types are restricted to bone
marrow

Stages are the same for basophils, eosinophils and neutrophils, only the granules are different
How long does granulopoiesis take?

Diameter 16 24 10-12 μm

Maturation time 14 days

Mitotic yes no
Granules no azurophilic Specific 20% 80%
 Numbers and Lifespan of Neutrophils
Neutrophil populations in the
blood are normally quite high
ranging from 1.5-8x109/L

Neutrophils are also found in

“marginated pools,” vascular
pools located in the lungs,
spleen, and liver.

Key:
DAMPs – damage associated molecular
pattern molecules (AMPs)
PAMPs – pathogen AMPs
LPS – lipopolysaccharide
fMLF - formyl-methionyl-leucyl phenylalanine
HMGB1 - high-mobility group box-1

Carlos Silvestre-Roig et al. Blood 2016;127:2173-2181

©2016 by American Society of Hematology

Why are there so many neutrophils?
• Neutrophils are phagocytes
• Involved in first line of defence against invading pathogens
• Important mediators of inflammation induced injuries

• Recent work suggests neutrophils are actually heterogenous population with

roles in shaping adaptive and innate immune responses (Silvestre-Roig et al. Blood
2016;127:2173-2181)

• Neutropenia is defined by levels of neutrophils below 1.5 x 109/L. It is

associated with increased risks of infection

• Neutrophil leucocytosis (neutrophilia) refers to excess neutrophils (>8 x109/L)

and is usually associated with increased numbers of immature or band
neutrophils
 Disorders of Neutrophils
Helen Irving

Reference text: Hoffbrand & Moss (2016) Hoffbrand’s Essential Haematology

particularly chapters 8 and 9

Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
Neutrophils and Monocytes are Phagocytes
bacterium

• Bacteria are engulfed by neutrophil Phagocytosis

and initial endosome fuses with
lysosome to form phagosome Phagosome granule
• Enzymes in lysosome/phagosome
attack bacteria beginning
degradation
H2O2 activated O2 species
• Secondary granules also fuse with
lysosome/phagosome and secrete NO generated compounds
additional enzymes
• Activated O2 species also help Residual
destroy bacterium bacterial body
• Residual bacterial components are
excreted by exocytosis Exocytosis
Image: Volker Brinkmann; (2005) PLoS Pathogens Issue Image | Vol. 1(3) November 2005.
PLoS Pathogens 1(3): ev01.i03. https://doi.org/10.1371/image.ppat.v01.i03
 Neutrophils and Neutrophilia
Neutrophils (NEU): 50 - 70% WBC differential = 1.8-7.5 x 109/L

• Neutrophilia or increased levels of neutrophils can be due to:

• Bacterial infections (sometimes accompanied by changes in appearance).
• Chronic inflammation and wounds can also generate increases in neutrophils
as can cardiac infarcts.
• Other causes include pregnancy, metabolic disorders, acute haemorrhage,
neoplasms, treatment with G-CSF, etc.

Arrow points to Dohle bodies Toxic granulation Image sources:Chris Bradley

Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014".
WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
 Benign disorders – changes in neutrophil morphology

Source: Chris Bradley Source: John Lazarchick; Ginell R Post; ASH Image bank image #3567

Pelger-Huet anomaly May-Hegglin anomaly

• Both these conditions are autosomal dominant conditions with relatively insignificant
effects on the patient
• Activity: Compare and contrast the neutrophils? What makes them abnormal?
 Neutrophils and Neutropenia
Neutrophils (NEU): 50 - 70% WBC differential = 1.8-7.5 x 109/L

• Decreased neutrophils levels below 0.5 x 109/L can be very serious as

accompanied by increased susceptibility to infections.
• Neutropenia is often clinically associated with painful infections (e.g.
intractable ulcers) of the mouth and throat
• Decreased neutrophil levels can be due to benign ethnic neutropenia
• West African and middle eastern populations tend to carry polymorphisms in
the DARC gene resulting in lower neutrophil levels. The polymorphisms are
related to an increased resistance to malaria
• Many drugs can reduce neutrophils– particularly cytotoxics
• Some chronic autoimmune diseases cause neutropenia such as SLE
(systemic lupus erythyromatous)
Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
 Eosinophil and Basophils
Helen Irving

Reference text: Hoffbrand & Moss (2016) Hoffbrand’s Essential Haematology

Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
Eosinophils
Eosinophils (EOS): 1 - 5% WBC differential = 0.04-0.44 x 109/L

• Eosinophils enter inflammatory exudates and

are involved in local immune responses and
tissue repair
• Eosinophils particularly have roles in
• allergic responses
• defence against parasites
• removal of fibrin
Normal
• Note pink granules and bilobed nucleus. It is eosinophil
attached to a small lymphocyte
• such associations sometimes have diagnostic
importance

Image sources: https://library.med.utah.edu/WebPath/HEMEHTML

Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014".
WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN
2002-4436.
Basophils
Basophils (BASO): up to 1% WBC differential = 0.01-0.1 x 109/L

• Basophils rarely seen in normal peripheral

blood but are rapidly recruited to sites of
inflammation
• When stimulated, basophils release their
contents, including histamine, and
cytokines to aid immune responses, Normal
particularly against parasites basophil
• Note the large dense granules that
obscure the bilobed nucleus.
• Other cells are a band form neutrophil and a
monocyte.
Image sources: https://library.med.utah.edu/WebPath/HEMEHTML
Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014".
WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
BIO3CBH – Monocytes and
Macrophages
Lecturer
Helen Irving

monocyte
Source: ttps://la.m.wikipedia.org/wiki/
Fasciculus:Blausen_0649_Monocyte_(crop).png
 Growth factors responsible for haemopoiesis
Committed
progenitor cells Erythropoietin GM-CSF IL-3
Erythrocytes
IL-3 GM-CSF Thrombopoietin
Megakaryocyte Platelets

IL-1, IL-3, GM-CSF IL-3 M-CSF Monocytes

Monocyte-
IL-6, GM-CSF
granulocyte
GM-CSF, GM-CSF G-CSF IL-3
progenitor Neutrophils
Pluripotent SCF
stem cell GM-CSF IL-3 IL-5 Eosinophils

IL-3 Basophils

B lymphocytes

Interleukins (IL) 1, 2, 4, 6, 7, 12
Thymus T lymphocytes
SCF = stem cell factor, GM-CSF = granulocyte-monocyte colony stimulating factor, G-CSF = granulocyte colony stimulating factor, M-CSF = monocyte colony stimulating factor
Comparison of formation of monocyte and basophil phagocytes

Source: Figure 8.2, Hoffbrand & Moss (2016) Hoffbrand’s essential Haematology 7th ed
 Monocytes and their derivatives
• Monocytes develop in bone marrow and spend some time in peripheral circulation
before entering tissues where they mature to macrophages with different tissue
determined functions (i.e. phagocytosis, antigen presentation, and cytokine production)
• Macrophages can self replicate and can live for months to years.
• Monocytes can also form dendritic cells involved in antigen presentation

Normal monocyte

Kidney: Lungs: Spleen sinus: Lymph node

Brain: Serosal Liver: Bone marrow:
Intraglomerular Alveolar Macrophages, Macrophages,
microglia macrophages Küpffer cells macrophages, APC
mesangial cells macrophages APC APC
APC = antigen presenting cell
Monocytosis

• Normal levels of monocytes are: 200-950/μL or 0.2-1.0 x 109/L

• Monocytosis is the raised level of monocytes and is associated with
chronic bacterial or parasitic infections and various connective tissue
disorders.
 Monocyte disorders
Helen Irving

Reference text: Hoffbrand & Moss (2016) Hoffbrand’s Essential Haematology

particularly chapters 8 and 9

Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
 Monocytes and their fates

Normal monocyte

Kidney: Lungs: Spleen sinus: Lymph node

Brain: Serosal Liver: Bone marrow:
Intraglomerular Alveolar Macrophages, Macrophages,
microglia macrophages Küpffer cells macrophages, APC
mesangial cells macrophages APC APC
APC = antigen presenting cell

Monocyte migration to specific tissues and their differentiation occur upon

stimulation by different cytokines, interleukins and/or other factors cocktail.
Fate of monocytes – monocyte derivatives
• Depending on location, monocytes
become:
• interstitial dendritic cells
• macrophages
• micro-glial cells

• Dendritic cells are involved in

antigen presentation to T cells

Le Douce et al. (2010) Retrovirology

Normal and abnormal monocytes
• Monocytosis is relatively rare but is
induced by:
Normal Dysplastic
• Chronic bacterial infections such as
monocyte neutrophil
tuberculosis, typhoid, bacterial
endocarditis
• Connective tissue diseases – SLE (lupus),
rheumatoid arthritis, temporal arthritis
Abnormal
monocytes • Protozoan infections
• Chronic neutropenia
Pro- monocyte
• Various malignancies – Hodgkin’s
lymphoma, AML
• Chronic myelomonocytic leukaemia

Monoblasts Myeloblast
• Activity: Why is monocytosis
Patient with AMML Patient with AML associated with chronic infections?
From: Lynch et al. (2016) Int J Lab Hem. 2018;40:107–114.
AML – acute myeloid leukaemia; AMML – chronic myelomonocytic leukaemia
 BIO3CBH –Lymphocytes
Lecturer
Helen Irving

B lymphocyte T lymphocyte

Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
Haemopoiesis

Source: Figure 1.2, Hoffbrand & Moss (2016) Hoffbrand’s essential Haematology 7th ed
 Lymphocyte maturation (lymphopoiesis)

Image source: Abbas et al. 2016 Basic immunology see Fig 1-10 in 5th edition
Classes of Lymphocytes
• Lymphocytes are characterised by
different CD markers on the cell’s
surface.
• B lymphocytes
• ~20% of blood lymphocyte
• secrete specific soluble antibodies
into plasma
• T lymphocytes
• 80% of blood lymphocyte
• CD4 Helper T cells more common
• Secrete cytokines
• Cytotoxic T cells carry CD8 marker
• Kill infected cells
• Regulatory T cells
• Restrict action of T cells and maintain
homeostasis of immune response

Image source: Abbas et al. 2016 Basic immunology see Fig 1-9 in 5th edition
 Phases of adaptive immune response

Antigen
independent

Image source: Abbas et al. 2016 Basic immunology see Fig 1-19 in 5th edition
 Lymphocyte disorders
Helen Irving

Reference text: Hoffbrand & Moss (2016) Hoffbrand’s Essential Haematology

Image source: Blausen.com staff (2014). "Medical gallery of Blausen Medical 2014". WikiJournal of Medicine 1 (2). DOI:10.15347/wjm/2014.010. ISSN 2002-4436.
 Lymphocytes
Lymphocytes (LYM): 20 - 40% WBC differentiation = 1.5-3.5 x 109/L

A few lymphocyte facts;: -------------

Normal lymphocyte
Large granular lymphocyte Reactive atypical lymphocytes

From Chris Bradley

http://www.histology.leeds.ac.uk/blood/blood_wbc.php Munoz, and Kuriakose Blood 2012;120:1543

Lymphocytes and Lymphocytosis
Lymphocytes (LYM): 20 - 40% WBC differentiation = 1.5-3.5 x 109/L

• Lymphocytosis is usually caused by

• acute or chronic infections Blasts - Immature lymphocytes indicative
of acute lymphoblastic leukaemia (ALL)
• lymphoid leukaemias or lymphomas
• thyrotoxicosis
• Lymphocytosis occurs in young children in
response to infections that typically induce
neutrophil reactions in adults.
• Lymphoadenopathy may be localised (local
infection or malignancy) or generalised due to
infection , non-infectious inflammatory
diseases, malignancy or drugs. Image source: https://webpath.med.utah.edu/HEMEHTML/HEME018.html
Causes of lymphoadenopathy

Source: Figure 9.13, Hoffbrand & Moss (2016) Hoffbrand’s essential Haematology 7th ed
Lymphocytes and Lymphocytosis
Lymphocytes (LYM): 20 - 40% WBC differentiation = 1.5-3.5 x 109/L
• Infectious mononucleosis is commonly caused by
Epstein Barr virus (EBV) which is the most common
cause of glandular fever associated with
Reactive atypical lymphocytes
lymphadenopathy, sore throat, fever and atypical
lymphocytes.
• Infectious mononucleosis is generally asymptomatic
but disease is characterised by lymphocytosis. From Chris Bradley

• Differential diagnosis includes testing for various

potential viral causes other than EBV (e.g.
cytomegalovirus (CMV), HIV or toxoplasmosis (a
parasite), etc.)
Lymphocytes and Immunodeficiencies
Lymphocytes (LYM): 20 - 40% WBC differentiation = 1.5-3.5 x 109/L

• Both congenital and acquired immunodeficiencies can reduce lymphocyte

numbers and lead to increased susceptibility to infections
• Congenital defects can occur in genes involved in regulating lymphocyte
maturation and also lymphocyte activation

• Immunodeficiency also occurs with tumours of the lymphoid system

• Chronic lymphocytic leukaemia
• Myeloma
Common causes of acquired immunodeficiencies
Condition
Human immunodeficiency virus (HIV)
infection
Irradiation and chemotherapy
treatments for cancer
Immunosuppression for graft rejection
or inflammatory diseases
Bone marrow cancers (metastases,
leukeamias)
Protein-calorie malnutrition
Spleen removal
Pathology
Depletion of CD4+ helper T cells
Decreased bone marrow precursors for
all leukocytes
Depletion or functional impairment of
lymphocytes
Reduced sites for leukocyte
development
Metabolic imbalance inhibits
lymphocyte development
Decreased phagocytosis of microbes
The end