1 Haematopoiesis

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Haematology and Transfusion Science(BMS 303)

Module Co-ordinator : Dr Declan McKenna

Module Overview

Week 1

Haematopoiesis

Week 2

The Red Blood Cell

Week 3

Haemoglobin

Week 4

Nutrition & Blood

Week 5

Haemostasis

Week 6

Disorders ofHaemostasis

Week 7

The White

Blood Cell

Week 8Haematological

malignancies

Week 9

Myeloid

Malignancies

Week 10

LymphoidMalignanciesWeek 11

Transfusion Science

Lectures, Thursdays

10.15 -13.15, LT16

Module Practicals

Practical 1

Manual Blood Count

Practical 2

Osmotic Fragility

Practical 3

Haemoglobin

Estimation

Practical 4

White Blood Cell

Differential Count

Practical 5

Coagulation

Practical 6

Blood Grouping

Thursdays, 14.00- 17.00, TL1Module Resources

All module resources located within BMS303 module

area on BBLearn

Module Handout

Lecture Notes (New notes released each week)

Coursework guidance

Practical Handbook Self Assessment Quizzes (one per week)

Reading List and electronic resources

Past Papers

Revision Materials (Final Week)

Recommended Textbook(s)

Essential HaematologyHoffbrand & Moss

6th Edition

HaematologyBlann, Knight & Moore

Dacie & Lewis PracticalHaematology

Lewis, Bain & Bates

10th Edition

Other Useful Resources

Journals

Blood

British Journal of Haematology

Websites

American Society of Haematology

http://www.hematology.org/

Heme Team Interactive Haematology

http://www.hemeteam.com/

Atlas of Haematology

http://www.hematologyatlas.com/

Apps

CellAtlas : Guide to Blood Cell Morphology
http://www.hematology.org/http://www.hemeteam.com/http://www.hematologyatlas.com/http://www.hematologyatlas.com/http://www.hemeteam.com/http://www.hematology.org/


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Whats so big about Blood?

Blood has immense significance in human culture

Religious symbolism

Bible, Quran, various religions

sprinkling blood was called bledsian = blessingInheritance: Blood relation, bloodline, blood is thicker thanwater, Royal blood, blue blood

Conflict : blood feud, bad blood, blood bath, blood money

Emotion : cold-blooded, hot-blooded, red-blooded,merriness (Sanguine)

Symbolic : Blood brothers, Blood sacrifice, blood oath,signed in blood, Blood on hands

Fiction

Vampires

I smell the blood of an Englishman!

Gore/violence/movies/videogames/death/censorship

Blood in MedicineEgyptians bathed in blood for health

Greeks : One of the four humors (body fluids)

Associated blood with springtime (sanguine)

Believed heart contained the soul

Believed circulatory system carried air

Romans : drank blood

In 2nd century AD, Galen developed a theory of theblood system that lasted until 1200s

Arabia

In 1242, Ibn al-Nafis realised blood passed from rightto left side of body via lungs, not heart

England

In 1600s, William Harvey published his seminalaccount of the circulatory system, with heart as pump

Blood ComponentsIn the 1600s, the invention of the microscope meant the discovery

of blood cells

Red Blood Cells (late 1600s)

White Blood Cells (1800s)

Platelets (1800s)

Plasma could be separated from cells (early 1900s)

Now, we know blood is a mix of cells (~45%) and plasma (~55%)

WHAT IS HAEMATOLOGY AND WHY IS IT

IMPORTANT?

VIDEOLINK :A Day in the Life of a Haematologist

http://www.youtube.com/watch?v=yIQz360XRZU

In the course of this module, we will focus on many of the

areas mentioned in this video

Red Blood Cells / White Blood Cells / Platelets

Immune system

Leukaemia / Lymphoma

Coagulation

Anaemia

Blood Transfusion

Blood Grouping

From the Greek haima, meaning blood

Week 1

HEMATOPOIESIS

Introduction

In Lecture 1 we introduce key concepts in haematology and

transfusion science, many of which will be expanded on in

later lectures.

We discuss how haematopoietic stem cells are the foundation

of the adult blood system, sustaining the lifelong production of

all types of blood cell.

We consider the problems caused by abnormal blood cell

production and describe how stem cell transplantation can be

a possible cure for haematological disease.

We also introduce some basic laboratory techniques that are

employed in the study of haematology
http://www.youtube.com/watch?v=yIQz360XRZUhttp://www.youtube.com/watch?v=yIQz360XRZU


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Content

Haematopoietic Stem Cells

Haematopoiesis

Abnormal Hematopoiesis

Haematopoietic Stem Cell Transplantation

Common Haematological Techniques

Conclusion

Learning Outcomes

After completing this lecture you should be able to:

Know the characteristics of Haematopoietic Stem Cells.

Summarise the process of haematopoiesis (blood cell

formation).

Give examples of disorders where haematopoiesis is

abnormal.

Describe how haematopoietic stem cell transplantation

works in the treatment of haematological disease.

List the various haematological tests than can be employed

in blood testing in the laboratory

Haematopoietic Stem Cells

Haematopoietic stem cells (HSCs) have two main characteristics:

1. Self renewal

They are able to proliferate and form more stem cells, a

process known as self-renewal. So although they are used to

continually produce new blood cells, the overall stem cell

numbers remain constant in a normal healthy individual.

2. Differentiation

The second characteristic feature of HSCs is that they are

pluripotent, meaning they are able to undergo differentiation to

produce highly specialised mature cell types. There is also

considerable amplification in the process with one stem cell

capable of producing a million mature blood cells after 20

rounds of cell division

Self renewal and differentiation of HSCs

As they mature, HSCs become increasingly differentiated

and lose the ability to self-renew

A single HSC can give rise to > 106 mature cells after several

rounds of division

Sites of Haemopoiesis

Foetus 0-2 months yolk sac2-7 months liver, spleen5-9 months bone marrow

Infants Bone Marrow (all bones)

Adults Bone Marrow(vertebrae, ribs, sternum, skull,sacrum, pelvis, ends of femurs)

Sites of Haemopoiesis

Foetus Adult

Yolk sac

birth

Central skeleton(vertebrae, ribs, sternum)

Distal bones

1 3 60504030201075 9Months Years

Liver&

SpleenHaematopoiesis


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Bone Marrow Environment

Haematopoietic stem cells are located within the bone marrow

The stem cell compartment.

Ideal environment for stem cells to grow and develop.

Bone marrow is composed of stromal cells and a

microvascular network

Stromal cells include:

Macrophages

Fibroblasts

endothelial cells

fat cells

reticulum cells

They secrete extracellular

molecules

collagen

fibronectin

haemonectin

laminin

proteoglycans (growth

factors and adhesion

molecules)

HSCs are stored in the bone marrow which provides a suitable

microenvironment for promoting both self-renewal and

differentiation where appropriate.

Bone Marrow Structure

Scanning electron microscope image ofbone marrow

Bone marrow smear

HaematopoiesisHaematopoiesis (also known as haemopoiesis) is the process wherebyHSCs develop into mature blood cells via a series of committed progenitor

cells that are restricted in their developmental potential.

The progeny of pluripotent stem cells become more irreversibly committed tospecific cell lineages with each cell division.

The further along the differentiation pathway a cell progresses the more

restricted the range of mature cells it can produce.

As they mature, fully differentiated blood cells are gradually released from

the bone marrow stromal environment into the bone marrow microcirculation

and eventually into the blood circulation.

Among the earliest steps in haematopoiesis is the commitment and

differentiation of the haematopoietic stem cell to become the precursor for

cells of either the myeloid lineage or the lymphoid


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Figure explanation (previous slide)

Representation of the pluripotent HSC and the

various progenitor and mature cells that arise from

it. The various progenitor cells can be identified invitro by culturing in semi-solid medium and

observing which type of colony they form (CFU).

Abbreviations: CFU, colony forming unit; BFU,

burst forming unit; Baso, basophil; E, erythroid; Eo,

eosinophil; GEMM, granulocyte, erythroid,

monocyte and megakaryocyte; GM, granulocyte,

monocyte; Meg, megakaryocyte; NK, natural killer.CFU-E

B o n e

M a r r o w

B l o o d

Myeloid

stem cell

BFU-E

CFU-E

Erythrocyte

Pluripotent

stem cell These two

pictures were

taken in a lab

in the

University of

Ulster and

show the

early stages

of erythroid

differentiation

using in this

case

embryonic

stem cells.

BFU-E

CFU-E

In vivo, immature cells in the various pathways are known and

identified as blasts, of which there are different types,

depending on which lineage they belong to.

For example :

Myeloblast : blast cell from myeloid pathway

Lymphoblast: blast cell of the lymphocyte pathwayErythroblast: blast cells in the red blood cell pathway.

These terms and others become important when studying

haematological disease, since an increase in blast numbers

and/or change in location from bone marrow can be indicative

of disease. This will be discussed further in Lectures 8 to 10.

BLASTS

Myeloid Lineage(CFU GEMM progenitor cell)

Lymphoid lineage(Common lymphoidprogenitor cell (CFUL))

Red cells (Erythrocytes) B-Lymphocytes

Platelets T-Lymphocytes

Monocytes NK Cells

Neutrophils

Eosinophils

Basophils

Mature blood cells produced via the myeloid and

lymphoid lineages

LYMPHOID

MYELOID

Regulation of Haematopoiesis

The cell type that a stem cell matures into is largely decided by theexternal signals it receives.

Haematopoietic Growth Factors (HGFs) play a critical role in

regulating the proliferation and differentiation into various matureblood cells

These can act synergistically or may induce the production of one

another by their action upon cells in the bone marrow environment.

The figure on next slide shows major growth factors involved in

haematopoiesis of myeloid lineages.

Abbreviations: SCF, stem cell factor; IL, interleukin; GM-CSF,

granulocyte-macrophage colony stimulating factor; M-CSF,macrophage colony stimulating factor; EPO, erythropoietin; TPO,

thrombopoietin; G-CSF, granulocyte colony stimulating factor.


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HGF EFFECT ON

MYELOID LINEAGESCharacteristics of Growth Factors

Glycoproteins that normally act at very low concentrations

Usually produced by many cell types e.g. stromal cells,monocytes, macrophages and lymphocytes

Usually affect more than one lineage

Usually show synergistic or additive interactions with other

factors

Multiple actions

Biological effects mediated via specific receptors.

Figure 6. Growth factors may

stimulate proliferation of earlyprogenitors, direct differentiation,

stimulate maturation, suppress

apoptosis of control function of

mature cells, as exemplified

above for the action of G-CSF

upon early progenitor cells and

mature neutrophil

Important Growth Factors

Many HGFs have been identified in the past decades and while thefunction and role of many are well characterised and understood,

the actions of some growth factors and how/when/why they act is

still unclear. Some of the better understood HGFs are listed below.

Stem cell factor (SCF)Interleukin-3 (IL-3)Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF)

Erythropoietin (EPO) Discussed further in Lecture 2Thrombopoietin (TPO) Discussed further in Lecture 5Macrophage-Colony Stimulating Factor (M-CSF)Granulocyte-Colony Stimulating Factor (G-CSF)

The link below has vast amounts of additional information about

HGFs and cytokines.

http://www.copewithcytokines.de/cope.cgi

Haematopoietic Growth Factor Receptor Signalling

The multiple actions of HGF on haematopoietic cells are

mediated through recognition of the growth factor by its specific

cell surface receptor.

Most haematopoietic cells have only a few hundred receptors

per cell for each growth factor and low levels of growth factor

binding to their specific receptor causes important biological

responses. This is achieved via a series of intracellular signalling

mechanisms.

Binding of a growth factor to its cell surface receptor leads to the

activation of associated kinases and the phosphorylation of the

receptor.

There are many varied and complicated signalling mechanisms

activated downstream of growth factor receptors. A few of the

main signalling pathways are shown on the next slide.

Figure 7. Binding of a

growth factor can activate

intra-cellular signalling

pathways which will

ultimately lead to the

transcriptional activation

of specific genes, which

promote cell growth.
http://www.copewithcytokines.de/cope.cgihttp://www.copewithcytokines.de/cope.cgi


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Mature Blood CellsSuccessful hematopoiesis results in the production of several types of

mature blood cell, each of which has a particular function within the

body (see Table on Next Page).

A healthy individual will have cell counts within the reference ranges

shown, although age, sex and general health must also be taken into

account in interpreting results.

Any deviation outside these values may be indicative of an underlying

illness or abnormality, which may range from a simple infection to a

more serious blood disorder.

We will discussRed Blood Cells (erythroblasts) and their function in Lectures 2 & 3Platelets and their function in Lecture 5 & 6White Blood Cells and their functions in Lecture 7

TYPE LINEAGE NAME PRODUCTIONPROCESS MAJOR FUNCTIONTYPICAL

NUMBERS

Red Blood

CellsErythrocytes Eryrthopoiesis

Carry oxygen (andCO2) round the body

(Haemoglobin)

5.0 0.5. x 1012 / L(Men)

4.3 0.5. x 1012 / L(Women)

Platelets Platelets Thrombopoiesis Blood Coagulation 280 130 x 109 / L

White

Blood

Cells

Monocytes Monopoeisis

Become tissue

macrophages whichphagocytose and digest

invading micro-organisms

and foreign bodies

0.2 - 1.0 x 109 / L

Basophils GranulopoiesisImmune response (canrelease histamine and

serotonin)

0.02 0.1 x 109 / L

Eosinophils Granulopoiesis

Modulate allergicinflammatory response

and destroy larger

parasites

0.02 0.5 x 109 / L

Neutrophils Granulopoiesis Phagocytose and destroyinvading bacteria

2.0 7.0 x 109 / L

B-Lymphocytes Lymphopoiesis Immune response (MakeAntibodies)

1.0 3.0 x 109 / L(total lymphocytes)

T-Lymphocytes LymphopoiesisKill virus-infected cells and

regulate activities of otherwhite blood cells

NK Cells Lymphopoiesis Kill virus-infected cells andsome tumour cells

VIDEO LINK. How Blood cells are formed

http://www.youtube.com/watch?v=tDTLC2swhlQ&feature=related

VIDEOLINK. What is blood?

http://www.youtube.com/watch?v=CRh_dAzXuoU&feature=related

Aplastic AnaemiaAbnormal Hematopoeisis can lead to a variety of disorders

Aplastic anaemia arises from

a reduction in the number of HSCs

a failure of remaining HSCs to divide and differentiate

sufficiently

exposure to radiation, chemicals, drugs and viruses.

Aplastic anaemia results in pancytopenia.

Pancytopenia is the reduction in the blood count of all the major

blood cell types; red, white and platelet.

This is characteristic of aplastic anaemias, although it can also

be observed in various leukaemias.

Because they lack all types of blood cell, aplastic anemia

sufferers may exhibit

Fatigue, pallor (due to lack of red blood cells)

Increased bruising / haemorrhage (due to lack of

platelets for coagulation)

Increased risk of infection (due to lack of white blood

cells)

Diagnosis involves a complete set of blood counts and tests,

in order to distinguish cause and severity.

Treatment and management may involve a number of

approaches, including stimulating immune system, steroids

and hormone therapy.

However, the only chance of permanent cure will be a stem

cell transplant. Hypoplastic Bone marrow in aplastic anaemia sufferer

Read more about aplastic anaemia here.

http://www.mayoclinic.com/health/aplastic-anemia/DS00322
http://www.youtube.com/watch?v=tDTLC2swhlQ&feature=relatedhttp://www.youtube.com/watch?v=CRh_dAzXuoU&feature=relatedhttp://www.mayoclinic.com/health/aplastic-anemia/DS00322http://www.mayoclinic.com/health/aplastic-anemia/DS00322http://www.mayoclinic.com/health/aplastic-anemia/DS00322http://www.mayoclinic.com/health/aplastic-anemia/DS00322http://www.youtube.com/watch?v=CRh_dAzXuoU&feature=relatedhttp://www.youtube.com/watch?v=tDTLC2swhlQ&feature=related


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HAEMATOPOIETIC STEM CELLTRANSPLANTATION

In many haematological disorders, a possible chance for a

complete cure comes from haematopoietic stem celltransplantation (HSCT).

This refers to the transplantation of blood stem cells into a

patient and is often referred to as a bone marrow transplant

(BMT), since the HSCs used for transplantation will have been

collected from the bone marrow.

However, it is becoming more common for HSCs to be collected

from the peripheral blood, in which case the process is referred

to as peripheral blood stem cell transplantation (PBSCT).

Sources of HSCs

The source of the HSCs can be :

Autologous (From patient)

Allogeneic (From matched donor).

Matching involves ABO blood typing and HLA

(tissue antigen) typing to minimise risk of transplant

rejection. A sibling is the preferred donor as the

likelihood of HLA-matching is higher

Syngeneic (From an identical twin) This would be

the ideal transplant.

The process of HSCT is

usually as follows :

1. HSCs collected, isolated

and stored from donor.

2. Patient is treated with high-

dose chemotherapy and/or

radiotherapy to eradicate

the patient's malignant cell

population and eliminatethe patient's bone marrow

stem cells

3. HSC transplant to replace

bone marrow stem cells

4. Blood system is restored by

the healthy, transplanted

HSCs

ALLOGENEIC HSCT

In autologous HSCT the

process is the same, except

that the source of the HSCs is

the patient themselves.

In this case, the HSCs are

harvested before the

chemotherapy.

AUTOLOGOUS HSCT

Autologous v Allogeneic HSCT

Graft versus Host disease (GVHD) is a complication that can occur after

a stem cell or bone marrow transplant in which the newly transplantedmaterial attacks patients body, even though the donor has been matched

to the patient to minimise this. It can be acute (within 3 months) or chronic

(may last lifetime).

Graft versus leukaemia (GVL) (AKA graft versus tumour) is a similareffect, but is beneficial, in which any white blood cells in transplant mount

an immune response against any residual leukaemic cells still present in

the patient.

Source of HSCs Benefits Limitations

Patient (Autologous) No rejection (GVHD)

HSCs may have potential

to turn cancerous again

No GVL

Donor (Allogeneic) GVLPossibility of rejection

(GVHD)

Sources of HSCs for transplant

Sources

Bone marrow

Peripheral blood

Umbilical cord blood


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Bone Marrow

stem cells are harvested directly

from crest of the ilium (pelvis)

Has large amount of red marrow

may also be taken from the

sternum

usually under general anaesthesia.

minimally invasive procedure with only minor

discomfort

However, if patient cancer was due to stem cell

defect, then using their own bone marrow may not

be useful as disease will likely develop againVIDEO LINK -BONE MARROW TRANSPLANT

http://www.youtube.com/watch?v=GIy2nMnuGGI

Harvested cells must be purified before storage for use.

Peripheral blood now the most common source of stem cells for HSCT.

PBSC yield boosted by injections of G-CSF

mobilizes HSCs from the donor's bone marrow into the

peripheral circulation.

Cells collected through a process known as apheresis.

Donor's blood is withdrawn through a needle in one arm &

passed through machine that removes WBC

The red blood cells are returned to the donor.

Umbilical cord bloodStem cells harvested from a newborn's umbilical cord and placentaafter birth.

Cord blood has a higher concentration of HSCs than is normally

found in adult blood.

Allogeneic cord blood is stored frozen at a cord blood bank because

it is only obtainable at the time of childbirth

Cord Blood banking (private and public) is increasingly big businessin the US and UK. http://www.nhsbt.nhs.uk/cordblood/

However, numbers of stem cells in cord blood are small need

ways to expand them in the lab.

VIDEO LINK: How to collect cord Blood

http://news.bbc.co.uk/1/hi/health/8556741.stm

Response to HSCT

Ideally, the patient will start to respond to treatment within a few

weeks, which will be monitored by blood cell counts.

Various supportive therapies may be administered before and

during this post-transplant period to improve the chances of a

successful outcome. These often include :

platelet transfusions to prevent bleeding/haemorrhage.

cyclosporin A therapy, which suppresses the patients

immune system and therefore minimises the risk of

transplant rejection.

Red blood cells to aid oxygen transport.

However, although advances have been made in the

administration of HSCT, the 100 day transplant related

mortality (TRM) is still between 10 and 15%, dependent on

patient age, donor type, disease status and stem cell source.

Even in a successful transplant, patients can often experience

many side-effects and it can take 1 -2 years for the treatment

process to be entirely completed and full health restored.

You can read some personal accounts of the experiences of

various non-Hodgkins Lymphoma patients who received

HSCT as treatment at the following website

http://www.nhlcyberfamily.org/stories.htm
http://www.youtube.com/watch?v=GIy2nMnuGGIhttp://www.nhsbt.nhs.uk/cordblood/http://news.bbc.co.uk/1/hi/health/8556741.stmhttp://www.nhlcyberfamily.org/stories.htmhttp://www.nhlcyberfamily.org/stories.htmhttp://news.bbc.co.uk/1/hi/health/8556741.stmhttp://www.nhsbt.nhs.uk/cordblood/http://www.youtube.com/watch?v=GIy2nMnuGGIhttp://en.wikipedia.org/wiki/File:Bone_marrow_biopsy.jpg


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Causes of death after SCT (2001-2006)

Complications of HSCT

Many complications of SCT exist, which

can be life-threatening if severe enough

Graft failure

Acute GVHD Chronic GVHD

Organ toxicity

Infection

Skin reactions Bleeding

Infertility

Nevertheless, at present it offers one ofthe best chances for a complete cure

from leukaemia / lymphoma and is

increasingly an option for treating these

diseases

Common haematologytechniques in the laboratory

Common haematological techniques

Blood Collection

Blood counts (Practical 1, Discussed in Lectures 2 & 7)

Blood films (Practical 4, Discussed throughout module)

Flow cytometry (Discussed throughout module)

Coagulation Screens (Practical 5, Discussed in Lectures 5 & 6)

Other Tests

A great number of tests and assays can be performed for various

micronutrients, components and characteristics of blood, as well as a

range of measurements that fall into the field of clinical biochemistry(e.g. hormones, metabolites, lipids).

A comprehensive overview of reference ranges for various tests can be

viewed here. http://pathcuric1.swmed.edu/pathdemo/nrrt.htm.Note that different labs often have slightly different ranges and these

will be constantly reviewed and updated where appropriate.

Blood collection Blood is collected by a trained phlebotomist usingvenupuncture (ie from a vein).

The blood is collected in tubes which have different

coloured tops, indicating the presence (or absence)

of various anti-coagulants to prevent the blood

clotting.

Without anti-coagulants, the blood will clot within 2 -5

minutes and can be centrifuged to separate theserum from the cells

For blood counts, EDTA is used to chelate thecalcium ions that are essential for clotting, thereby

preserving the integrity of the blood cells.

For coagulation tests, however, sodium citrate is

used to anticoagulate the plasma component.

The correct collection and storage of the collected

blood is crucial, since many tests become less

reliable if the blood is handled or stored wrongly

Blood CountsA full blood count is the most commonly performed haematological

blood test

Gives information on the numbers of each type of blood cell in a given

sample

Also measures haemoglobin concentration and calculates various Red

Cell indices will be calculated .

White cells differential count will indicate if the white blood cells are in

the correct proportion

In modern laboratories, this is usually carried out by automated

analyzers and if the values fall outside normal ranges, it will be flaggedup for further investigation.

However, counts can also be performed manually

Blood filmsAlthough blood counts are now usually automated, blood slides

will also be produced if necessary to allow examination and/or

count of the blood cells under a microscope.

For routine slide analysis, this involves staining with specific

dyes to highlight the distinctive features of each cell type.

Romanowsky stains are universally employed for routine

staining of blood films and when prepared properly gives very

satisfactory results.

Most modern haematology labs have machines that produce

slides automatically. However, they can also be prepared by

hand

http://www.youtube.com/watch?v=ZyU9iZ9d9QI&feature=related
http://pathcuric1.swmed.edu/pathdemo/nrrt.htmhttp://www.youtube.com/watch?v=ZyU9iZ9d9QI&feature=relatedhttp://www.youtube.com/watch?v=ZyU9iZ9d9QI&feature=relatedhttp://www.youtube.com/watch?v=ZyU9iZ9d9QI&feature=relatedhttp://pathcuric1.swmed.edu/pathdemo/nrrt.htm


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Cell StainsThese stains are able to show subtle differences in shades of staining

and stain granules differently due to the presence of two components

azure B (trimethylthionin) and

eosin Y (tetra-bromo-fluoroscein).

Azure B is a basic dye and binds anionic molecules. Therefore, ittargets acidic groupings of nucleic acids, proteins and primitive

cytoplasm

Eosin Y is an acidic dye and binds to cationic sites on proteins.

Therefore, it targets basic residues, such as those found onhaemoglobin

The combination of these two elements binding can therefore be used

to identify most cellular elements.Normal and/or abnormal cells can be thus identified by a trained

biomedical scientist or clinician

Different types of blood cell can be ident ified in a blood smear, based on physical

characteristics such as size, shape, colour, nuclei and granulation following

staining with Romanowsky solutions. E, erthyrocyte (many); P, platelet (small);

N, neutrophil; B, basophil; Eo, Eosinophil; M, monocyte; L, lymphocyte.

Flow cytometryImmunophenotyping (flow cytometry) is a method whereby the physical

and/or chemical characteristics of up to thousands of cells in a sample

can be analysed in seconds.

It is routinely used in the diagnosis of health disorders, especially blood

cancers.

HSCs, the various progenitor cells and mature blood cells all exhibit

different patterns of marker proteins on their surface, known as Cluster of

Differentiation (CD) markers.

The CD marker profile of these cells is now well established, allowing

them to be distinguished from each other.

We can use fluorescent antibodies targeted to specific CD markers to

label cells of interest.

This means that flow cytometry can then be used to calculate how many

cells displaying that marker (or a combination of markers) are present,

based on fluorescent signals from the antibodies.

The graph on

the left

demonstrates

how different

types of blood

cell can be

differentiated

between,

based on size

(x-axis) and

internal

complexity (y-

axis

Flow cytometry simple example

The two graphs above compare the lymphocyte profile in blood from a

normal person and a leukaemia patient. The leukaemia patient has an

abnormal profile, indicating presence of large (ie toward right of graph),but undifferentiated (ie toward bottom of graph) lymphocytes, suggestinga problem with the differentiation and development of the lymphoid

lineage. This information, along with other tests, helps make an informed

diagnosis of disease.

Other common tests

TEST FUNCTION

Erythrocyte

sedimentation rate

Assess inflammatory response to

injury/treatment

Plasma viscosityMeasures how thick or thin the plasma

component of blood has become

Haematinic assaysMeasures concentration of micronutrients and

plasma proteins. E.g. Iron, ferritin, vitamin B12

or folate. Discussed further in Lecture 4

Haemoglobin variant

detection

Separation methods like electrophoresis or

chromatography are used to identify diseases

linked to abnormal haemoglobin production .

Discussed further in Lecture 3.

Molecular techniquesAnalysis of genetic information has improved.These techniques will be discussed further

throughout the other lectures.

thick

thick

L3

PCR, microarry


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CONCLUSIONThis lecture has introduced several key concepts in

haematology and transfusion science.

We have looked at the concept of haematopoietic stem cells

and how they can give rise to progenitor cells which willultimately differentiate into mature blood cells.

We have seen how this process of haematopoiesis is

regulated by various growth factors, which help determine the

lineage along which a stem cell will differentiate.

Weve appreciated how haematological diseases can arise if

haematopoiesis is disrupted and seen how stem cell

transplantation may provide a cure for such diseases.

Finally, we have reviewed commonly used haematological

techniques used in blood testing.

Throughout the remainder of the module we will return to

these concepts and expand upon them in relation to various

aspects of haematology.

WEBSITES ABOUT HSCs

NIH Stem Cell Information

http://stemcells.nih.gov/info/scireport/chapter5.asp

National Cancer Institute Stem Cell Transplantation

http://www.cancer.gov/cancertopics/factsheet/Therapy/bone-

marrow-transplant

Stem Cell Database

http://stemcell.mssm.edu/v2/introduction.shtml
http://stemcells.nih.gov/info/scireport/chapter5.asphttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://stemcell.mssm.edu/v2/introduction.shtmlhttp://stemcell.mssm.edu/v2/introduction.shtmlhttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://www.cancer.gov/cancertopics/factsheet/Therapy/bone-marrow-transplanthttp://stemcells.nih.gov/info/scireport/chapter5.asp

Documents

1 Haematopoiesis