HEMATOLOGY-UNDERSTANDIN THE HEMATOPOIESIS

HEMATOLOGY

2. HEMATOPOIESIS
Continuous,regulatedprocessofrenewal,proliferation,differentiation,and
maturationofallbloodcelllines
Resultintheformation,development,andspecializationofallfunctionalbloodcells
thatarereleasedfromthebonemarrowintothecirculation
Maturebloodcellshavealimitedlifespan(120days+/-20daysforRBCs)andacell
populationcapableofself-renewalthatsustainsthesystem
Ahematopoieticstemcell(HSC)iscapableofself-renewalanddirecteddifferentiation
intoallrequiredcelllineages

3. STAGES OF HEMATOPOIESIS
Pre-natal/Primitive hematopoiesis
Mesoblasticphase
Hepatic phase
Medullary/Myeloid phase –continues to adult hematopoiesis
Adult/Definitive hematopoiesis

4. MESOBLASTIC PHASE
Also called as the yolk sac phase
Begins during the 19
th
-20
th
day of gestation in the blood islands of the yolk sac (mesodermal
extraembryonic layer yolk sac)
Hematopoietic activity is confined to erythropoiesis
Production of cells lasts until the: 8
th
to 12
th
week of gestation
Cells produced in the yolk sac: primitive erythroblasts and angioblasts(form blood vessels)
Primitive erythroblasts
important in early embryogenesis to produce embryonic hemoglobin (up to the 3
rd
month only)
Large, nucleated, and cannot extrude its nucleus
Migrates to the Aorta-Gonad Mesonephros(AGM)region to become definitive erythroblasts
(produces HSCs)
Hemoglobin produced (Embryonic hemoglobin):
Gower-I (2 epsilon and 2 zeta globin chains)
Gower-II (2 alpha and 2 epsilon globin chains)
Portland hemoglobin (2 zeta and 2 gamma globin chains)

5. HEPATIC PHASE
After3
rd
month,yolksacstopsproducingbloodcellsandisreplacedbythefetalliver
Startsatthe5
th
–7
th
weekofgestation(primarysiteuntilthe6
th
monthandcontinuetoproduce
until1
st
to2
nd
weekoflife)
Characterizedbyrecognizableclustersofdevelopingerythroblasts,granulocytes,andmonocytes
colonizingthefetalliver,thymus,spleen,placenta,andultimatelythebonemarrow
Developmentofspleen,thymus,kidneyandlymphnodescontributestothehematopoietic
process(extramedullaryhematopoiesis)
Thymus-firstorgantobedevelopedandproducesTcells(T-cellmaturation)
Kidneyandspleen–B-cellmaturation

6. HEPATIC PHASE
Bloodcellsformed:
Definitiveerythroblasts
Granulocytesandmegakaryocytes:3
rd
month
Lymphocytes:4
th
month
Monocytes:5
th
month
Hemoglobinformed:
Fetalhemoglobin(HgbF)2alphaand2gammaglobinchains
Adulthemoglobin(HgbA
1andA
2)detectablelevels
HgbA
12alphaand2betaglobinchains
HgbA
22alphaand2deltaglobinchains

7. MEDULLARY PHASE/MYELOID PHASE
Termedmedullaryhematopoiesisbecauseitoccursinthemedullaorinnerpartofthe
bonecavity
Primarysiteofhematopoiesisistheredbonemarrowstarting24weeksofgestation
(remainsthroughoutlife)
Beginsbetweenthe4
th
and5
th
monthoffetaldevelopment
HSCsandmesenchymalcellsmigrateintothecoreofthebone
Mesenchymalcells-typeofembryonictissuewhichdifferentiateintostructural
elementsthatsupportdevelopinghematopoieticelements
MyeloidactivityisapparentduringthisstagewithanormalM:Eratioof3:1–4:1

8. MEDULLARY PHASE/MYELOID PHASE
Detectablecytokines:
Erythropoietin (EPO)
Granulocyte colony-stimulating factor (G-CSF)
Granulocyte-macrophage colony-stimulating factor (GM-CSF)
Hemoglobin formed:
Fetal hemoglobin (HgbF)
Adult hemoglobin (HgbA
1and HgbA
2)

10. ADULT HEMATOPOIEITIC TISSUE
In adults, hematopoietic tissue is located in the:
Bone marrow -erythroid, myeloid, megakaryocytic, and lymphoid cells
Lymph nodes, Spleen, Liver, and Thymus (extramedullary hematopoiesis)
Lymphoid tissues
Primary lymphoid tissues where T and B cells are derived
Bone marrow and Thymus
Secondary lymphoid tissues lymphoid cells respond to foreign antigens
lymph nodes, mucosa-associated lymphoid tissue (MALT), gut-associated
lymphoid tissue (GALT)

11. BONE MARROW
Normal bone marrow contains two major components:
Red marrow hematopoieticallyactive marrow (blood cells and precursors)
Yellow marrow hematopoieticallyinactive marrow (adipocytes, with undifferentiated
mesenchymal cells and macrophages)
Infancy and early childhood
80-90%active marrow
all the bones in the body contain primarily red (active) marrow
5 to 7 years of age:
60%active marrow
adipocytes become more abundant and begin to occupy the spaces in the long bones
previously dominated by active marrow
Adults:
40%active marrow
Active marrow is restricted to the sternum, vertebrae, scapulae, pelvis, ribs, skull,and
proximal portion of the long bones

12. BONE MARROW
Retrogressionprocessinwhichtheredmarrowis
replacedbytheyellowmarrow
IMPORANTNOTE:
Yellowmarrowiscapableofrevertingbacktoactive
marrowincasesofincreaseddemandonthebone
marrow,suchasinexcessivebloodlossorhemolysis
Marrow cellularity -ratio of the red marrow to the yellow
marrow; usually decreases with age
Normocellular: marrow has 30-70%hematopoietic
cells
Hypocellular: marrow has <30% hematopoietic cells
Hypercellular: marrow has >70% hematopoietic cells
Aplastic: marrow has few or no hematopoietic cells

13. BONE MARROW
COLLECTIONPROCEDURES:
Bone marrow aspirate obtained by
bone marrow aspiration
Core biopsy obtained by Trephine
biopsy
COLLECTIONSITES:
Posteriorsuperioriliaccrest
Anteriorsuperioriliaccrest
Sternum
Anteriormedialsurfaceofthetibia
Spinousprocessofthevertebrae,ribs,
orotherred-marrowcontainingbones

14. BONE MARROW
EQUIPMENTS USED FOR SPECIMEN COLLECTION:
Surgical gloves
Shaving equipment
Antiseptic and alcohol pads
Drape material
Local anesthetic injection
No. 11 scalpel blade
Jamshidibiopsy needle or Westerman-Jensen
needle; Snarecoilbiopsy needle
14 to 18 gauge aspiration needle with obturator

15. BONE MARROW
TYPES OF BONE MARROW SPECIMEN:
Direct Aspirate Smears: wedge-shape smear; avoids crushing the spicules
Anticoagulated Aspirate Smears: K
3EDTA
Crush Smears: places additional glass slide directly over the specimen
Imprints: closely replicate aspirate morphology
Concentrate Smears: narrow-bore glass or a plastic tube such as Wintrobehematocrit
tube
Histologic Sections: 10% formalin, Zenkerglacial acetic acid, or B5 fixative
Marrow Smear Dyes: Wright or Wright-Giemsa dyes

17. BONE MARROW
NORMALMARROWCELLS:
HEMATOPOIETICCELLS–PHSC,Progenitorcells,precursorcells
STROMALCELLSoriginatefrommesenchymalcellsthatmigrateintothecentralcavityof
thebone
Endothelialcells-Regulatestheflowofparticlesenteringandleavinghematopoietic
space
Adipocytes-Secretessteroidsthatinfluenceserythropoiesisandmaintainsboneintegrity
Macrophages-Phagocyticcellandsecretescytokinesneededfortheregulationof
hematopoiesis
Osteoblasts-Boneformingcells;water-bugorcometappearance
Osteoclasts-Boneresorbingcellsordestroyingcells
Reticularcells/Fibroblasts-Supportsvascularsinusesanddevelopinghematopoieticcells

18. RED MARROW
Composedofhematopoieticcellsarrangedinextravascularcords
Cordsarelocatedinspacesbetweenvascularsinusesandare
supportedbytrabeculaeofspongybone
Thecordsareseparatedfromthelumenofthevascularsinusesby
endothelialandreticularadventitialcells
Hematopoieticcellsdevelopinspecificnicheswithinthecords
Erythroblasts-developinsmallclusters,andthemoremature
formsarelocatedadjacenttotheoutersurfacesofthe
vascularsinuses
Megakaryocytes-arelocatedadjacenttothewallsofthe
vascularsinuses,whichfacilitatesthereleaseofplateletsinto
thelumenofthesinus
Immature(myeloid)granulocyticcells-arelocateddeep
withinthecords
Nichesakahematopoieticmicroenvironment;playsan
importantroleinnurturingandprotectingHSCsandregulating
theirquiescence,selfrenewal,anddifferentiation

19. HEMATOPOIETIC STEM CELLS (HSC)
Also called has pluripotentialhematopoietic stem cell (PHSC)
can differentiate into progenitor cells committed to either lymphoidor myeloid lineages when stimulated
by cytokines
are capable of self-renewal, are pluripotent, apoptosis,and may give rise to differentiated progeny
TYPES:
Non-committed progenitor cells or undifferentiated HSCs
Committed progenitor cells / Lineage specific progenitors
Common myeloid progenitor/CFU-S/CFU-GEMM granulocytic, erythrocytic, monocytic, and
megakaryocytic lineages
Common lymphoid progenitor T, B, and natural killer lymphocyte and dendritic lineages
Lineage-specific progenitors give rise to morphologically recognizable, lineage-specific precursor cells
Precursor cells blast forms (ex. myeloblast, megakaryoblast, erythroblast)

23. STEM CELL SURFACE RECEPTORS
SURFACE RECEPTORS/MARKERS OF CELLS
CD 34All hematopoietic stem cells; early progenitor cells
CD 33
CD 38
Myeloid cells
CD 71 Erythroid differentiation
CD 10
CD 38
B-lymphocytes (cALLa)
CD 19
CD 20
Lymphoid differentiation (B-cells)
CD 7
CD 5
T-lymphocytes
CD 8T-cytotoxic/suppressor cells
CD 4T-helper cells
CD 16
CD 56
NK cells
Used for identification and
origin of HSCs can be
determined by
immunophenotypicanalysis
using flow cytometry

24. STEM CELL THEORY AND KINETICS
STEMCELLTHEORY-Describestheoriginofhematopoieticprogenitorcells
Monophyletictheory
suggeststhatallbloodcellscellsarederivedfromasingleprogenitorstemcellcalledapluripotent
hematopoieticstemcell(PHSC)
Polyphyletictheory
suggeststhateachofthebloodcelllineagesisderivedfromitsownuniquestemcell
STEMCELLCYCLEKINETICS
Bonemarrowiscapableofproducingthefollowingperkgbodyweight:
2.5billionerythrocytes
2.5billionplatelets
1billiongranulocytes
HSCs exist in the marrow in the ratio of 1 per 1000 nucleated blood cells
Mitotic index
percentage of cells in mitosis in relation to the total number of cells
normal value is 1-2%
Increased mitotic index implies increased proliferationexcept in megaloblastic anemia (prolonged mitosis)

25. CYTOKINES AND GROWTH FACTORS
regulatethe proliferation differentiation, and maturation of hematopoietic precursor cells
are responsible for stimulation or inhibition of production, differentiation, and trafficking
of mature blood cells and their precursors
are a diverse group of soluble proteins that have direct and indirect effects on
hematopoietic cells
EXAMPLES ARE: Interleukins (ILs), Lymphokines, Monokines, Interferons, Chemokines,
Colony-stimulating factors (CSFs)
Positive influence:
KIT ligand, FLT3 ligand, GM-CSF, IL-1, IL-3, IL-6, and IL-11
Negativeinfluence:
Transforming growth factor-b, tumor necrosis factor-a, and interferons

29. HEMATOPOIETIC GROWTH FACTORS/CYTOKINES
GM-CSF Hematopoietic stem cells
G-CSF Granulocytes
M-CSF Monocytes/macrophages
EPO Erythrocytes
TPO Platelets
IL-3 Hematopoietic stem cells (produced by T-
cells)
IL-5 Activates T-cytotoxic cells; eosinophils
IL-6 B-cells
IL-7 T and B cells
IL-9 T and B cells
IL-12 NK cells
SCF/Kit ligandHematopoietic stem cells
Flt-3 ligandSame as SCF

30. LINEAGE SPECIFIC HEMATOPOIESIS
Erythropoiesis
Leukopoiesis Myelopoiesis, Lymphopoiesis
Megakaryopoiesis

31. END