1. Geoffrey Brown
College of Medical
and Dental Sciences
Revising textbook accounts of Haematopoiesis

3. Products of Haematopoiesis
MARROW BLOOD TISSUES
Megakaryoblast platelet
Erythroblast erythrocyte
Mast cell precursor mast cell mucosal and connective tissue mast cells
basophil
Myeloblast eosinophil in sites of inflammation
neutrophil
Monoblast monocyte tissue / lymphoid tissue macrophages;
Kuppfer cell; osteoclast
Precursor? dendritic cell tissue dendritic cell / Langerhans’ cell;
veiled cell; interdigitating cell
Precursor? natural killer cell lymphoid tissues; sites of inflammation
Pro-B cell type 1 and type 2 B cells in blood and secondary lymphoid tissues;
(matures in marrow) plasma cells also in mucosal surfaces and bone marrow
T cell precursor α / β and γ / δ T cells; CD4+ve and CD8+ve T cells; in blood,
(matures in thymus) secondary lymphoid tissues and sites of inflammation

4. Products of Haematopoiesis
Granulocytes Monocyte Lymphoid cells
Megakaryocyte
(i) Neutrophil (i) B lymphocyte
(ii) T lymphocyte
Macrophage
(ii) Eosinophil
Erythrocyte
(iii) NK cell
(iii) Basophil Dendritic Cell

6. Progressive Stages in Haematopoiesis
STEM HAEMOPOIETIC MATURE
CELLS PROGENITORS CELLS
Immortal Lineage selection Functional
Expansion Self renewal for
Self renewal? lymphocytes

7. The ‘Conventional’ model of haematopoiesis - Weissman
Hematopoietic Stem Cell
Common Myeloid
Common Lymphoid
Progenitor
Progenitor
M/E Progenitor G/M Progenitor
B cell T cell NK cell
Platelet Granulocyte Macrophage
Erythrocyte
Myeloid lineages Lymphoid lineages

8. Haematopoiesis – Principles
(i)Two families of cells – myeloid and lymphoid
(ii)Preferred single route to each cell type
(iii)Immortality confined to stem cells
(iv)Growth factors permissive or instructive?

9. Two families of cells?

10. Colony forming assays reveal myeloid potentials
G-CFU M-CFU

11. CONVENTIONAL MODEL OF HAEMATOPOIESIS
Haematopoietic stem cell
Myeloid progenitor (GEMM-CFU) Lymphoid progenitor
BFU-E GM-CFC Eo-CFC Meg-CFC Mast-CFC Pro-B Pro-T
erythrocyte neutrophil platelets B lymphocyte
monocyte eosinophil mast cell T lymphocyte

12. A lymphoid/myeloid dichotomy
B and T lymphocytes More mundane
are exceptional myeloid cells
Dependent on
Dependent on
receptor for
unique antigen-
growth factor
specific receptor
Domain of Province of
immunologists hematologists

13. Haematopoiesis – Principles
(i)Are there two families of cells – myeloid and lymphoid?
(ii)Preferred single route to each cell type
(iii)Immortality confined to stem cells
(iv)Growth factors permissive?

14. Evidence that precursor cells of monocytes and
B lymphocytes are closely related
Wong, Bunce, Lord, Salt & Brown – Exp. Hematol. 1989
3D gels of phosphoproteins
IEF
SDS
HL60 cells restricted to Lines restricted to monocyte Pre-B cell lines
neutrophil differentiation differentiation (U937, ML-1, (Nalm 6 & SMSB)
(HL60Ast3) HL60M2 & 15-12)

15. There are bi-potent B lymphocyte/monocyte cells
Bi-potent 1982 – Macrophages from pre-B lymphoma
progenitor cells (5-AZT & transduced CSF1 R (1990))
1988/94 – HAFTL-1 and 702/3 cell lines
1992 – Foetal liver of mice
(Cumano et al.)
1995 – Tumours in IL-7 transgenic mice
(Fisher et al.)
2001 – Bone marrow of adult mice
(Montecino-Rodriguez et al.)
B cell Monocyte

16. Progenitors with lymphoid potentials and an incomplete set of
myeloid potentials
Pax5 (B cell factor)-/- Lymphoid-primed MultiPotent
Progenitors (Adolfsson et al., 2005)
Pro-B cell
LMPP
Meg/Ery Gran/Mon
potentials potentials
Myeloid NK cell T cell
Early Progenitors with Lymphoid
and Myeloid develop. potential
(Balciunaite et al., 2005)
Lin-veSca-1+veKit+ve Lin-veSca-1+veKit+ve
M-CSF IL-2 IL-7 IL-7 Flt3hiMpl+ve Flt3hiMpl-ve
trans. Notch
Notch
Myeloid NK cell T cell B cell Lymphoid
priming

17. There isn’t a strict myeloid/lymphoid dichotomy

18. Haematopoiesis – Principles
(i)Two families of cells – myeloid and lymphoid
(ii)Is there a preferred single route to each cell type?
(iii)Immortality confined to stem cells
(iv)Growth factors permissive?

19. Is there just a single route to each cell type?
Dendritic cell sub-sets are phenotypically and
transcriptionally identical (Ishikawa et al. 2007)
Mon B
NK
Neu CLP
T
CMP Eos
Bas
Ery Meg
There are at least two alternative routes to dendritic cells

20. Multiple routes to DCs, neutrophils and monocytes

21. A.
Platelet/Erythroid
Granulocyte/Monocyte
CMP
Granulocyte/Monocyte
Platelet/Erythroid
NK cell
HSC T cell
B cell C.
HSC
CLP
B.
CMP
Erythroid Myeloid MEP ?
?
MEP Erythroid
?
pro T
Platelet T cell
Platelet
HSC
B cell Myeloid B cell (NK?) T cell
Myeloid
Myeloid

22. More versatility - thymus progenitors have clandestine
myeloid potentials
DC
NK
B cell Myeloid
Thymus-Settling Early Thymocyte Double Negative-2
DN-3
Progenitors Progenitors (DN1) (DN2)
Loss of CD117 & CD44
CCR9+ve, CD135+ve, Loss of CCR9 & CD135 Gain of CD25 Gain of cytoplasmic CD3
CD117high, CD44+ve,
CD25-ve
X X
B cell NK DC Myeloid

23. Haematopoiesis – Principles
(i)Two families of cells – myeloid and lymphoid
(ii)Preferred single route to each cell type
(iii)Is immortality confined to stem cells?
(iv)Growth factors permissive?

24. Immortality extends to lineage-biased cells
STEM HAEMOPOIETIC MATURE
CELLS PROGENITORS CELLS
Myeloid-
biased
Lymphoid
-biased
Immortal Lineage selection Functional
Expansion Self renewal for
Self renewal? lymphocytes

25. Haematopoiesis – Revised Principles
(i)A single family of cells
(ii)Immortality extends to lineage-biased
cells
(iii)More than one route to some cell types
(iv)Progenitors have clandestine options
(v)HSC and progenitors are more versatile
than previously thought

26. The Sequential Determination Model (1985)
Mast Cell/
Basophil Neutrophil B lymphocyte
Platelet Eosinophil Monocyte T lymphocyte
Erythrocyte
HSC CLP
Neutrophil Monocyte
Meg Ery Bas Eos Neu Mon B T
Meg
sensitivity to
Ery macrophages from
DMSO pre-B lymphoma lines
Bas
Eos
Neu
G GM M
Mon
Ast4 Ast3 HL60 M2 Ast25 Ast1 B ?
Sp1 M4 17-6 T
15-12

27. Mast Cell/
Basophil Neutrophil B lymphocyte
Platelet Eosinophil Monocyte T lymphocyte
Erythrocyte
HSC CLP
(i)A single family of cells
(ii)Immortality extends to lineage-biased
cells
(iii)More than one route to some cell types
(iv)Progenitors have clandestine options
(v)HSC and progenitors are more versatile
than previously thought

28. A pair-wise relationship model

29. TRANSCRIPTION FACTORS AND PAIR-WISE RELATIONSHIPS
Knock-out mice Gene expression
GATA-2
megakaryocyte
c-myb GATA-1
erythrocyte EKLF
& decreasing
mast cell NF-E2 levels
PU.1
basophil
eosinophil
neutrophil C/EBPβ
monocyte
ikarus
B
lymphocyte
} EBF
T
lymphocyte
} GATA-3

30. C/EBPα PU.1
EDAG
Myb
high
Meg Ery Bas Eos Neu Mon DC B NK T
Notch
EKLF /CSL
Pax5
GATA-1 E proteins
MLLT3
FOG-1

31. IL-7
IL-5 IL-4
IL-21 IL-15
IL-9 IL-3 IL-33 GM-CSF IL-10
Meg Ery Bas Eos Neu Mon DC B NK T
Tpo Epo G-CSF
M-CSF

32. Haematopoiesis – Revised Principles
(i)A single family of cells
(ii)Immortality extends to lineage-biased
cells
(iii)More than one route to some cell types
(iv)Progenitors have clandestine options
(v)HSC and progenitors are more versatile
than previously thought

33. Lineage – where to draw the lines?
Lymphocytes Archetypes?
B cell T cell
NK cell
CD8 T cell
IFNγ, IL-2,
Ltα
CD4 T cell ?
(IL-10)
IL-4, IL-5
IL-13, IL-10
IL-25
IL-17a, IL-17f IL-4?
T helper 1 IL-21, IL-22 TGFβ, IL-35 IL-21
Accessorised?
IL-10 IL-10
T helper 2
T helper 17
T reg T follicular helper

34. Versatility – Are leukaemia stem cells as versatile?

35. Is decision-making growth factor driven?
A
Pluripotent Unipotent
stemcell progenitor
B

36. What is the way forward?
(i) High level cell indeterminacy Mature cells
Outcome
(ii) Boundary conditions &
Growth factors, cell-cell contacts Performance

37. Dr G Brown and Prof A Rot - University of
Birmingham;
Prof R Ceredig - National University of Ireland,
Galway;
Prof A Rolink - University of Basel;
Prof E Marcinkowska - University of Wroclaw;
Prof. G Studzinski - University of Medicine and
Dentistry, New Jersey;
Drs A Zelent and K Petrie - The Institute of Cancer
Research, London;
Prof A Kutner - The Pharmaceutical Research
Institute, Warsaw;
Dr S Elliman - Orbsen Therapeutics Ltd., Galway;
Prof N Barnes - Celentyx Ltd., Birmingham.
Martin Smith - High-Point Rendel Ltd, London
Prof Michael Danilenko, Ben Gurion University of
the Negev, Isreal
Dr Eustace Johnson, University of Aston
Prof Daniela Finke, University of Basel