Haematopoietic System Development Flashcards
Bone marrow function
Main site of haematopiesis
Thymus
Main site of T lymphocyte development
Spleen function
Blood depot and filter
Hameatopoietic system hierarchy
Highest - Haematopoietic
Can proliferate and differentiate give rise to downstream blood cells
CFU-S (splenic) (myeloid)
CFU-C (culture) (myeloid)
Common myeloid progenitor gives rise to myeloid cells
Common lymphoid progenitors give rise to lymphoid cells
HSC functional assay
Isolate bone marrow
Transplant into irradiated mouse (ablated all HSCs and other progenitors)
Then if HSC present after 3.5 months all haematipoetic cells in recipient will be derived from donor marrow
Reconstituted the haematopoietic system
CFU-S functional assay
Is more commuted progenitor than HSC
Transplantation into irradiated mouse will not reconstitute haematopoietic system long term
BUT
will produce colonies of myeloid cells in spleen
Each colony made from 1 transplanted cell (hence cfu)
Colony forms because cell survives but is corrupted in some way idk???
CFU-C functional assay
Colony Is also result of differentiation of one cell
C = culture
Plate cells from bone marrow into dish in semisolid medium with growth factors and they will produce colonies w diff morphologies of myeloid cells
Most committed from three discussed
Modes of HSC division
Asymmetric- maintenance of HSCs - homeostasis
Symmetric - expansion of HSCs (eg regeneration of haematopoietic system)
Symmetric division - expansion of progenitors
Mode gone with depends on conditions
Eg expansion in order to replenish blood cells
Foetal liver
Intermediate organ
Develops at early stage
Main haematopoietic organ prior to bone marrow development
Contains blood cells filling foetal liver parenchyma
CFU-S and yolk sac
Cfu-s only in yolk sac at E8
Then start to appear in F liver and circulation
Was believed that it was HSC source then (wrong !!!!)
Was thought then travel to liver after
First cfu-s seen at E10 there
The AGM
Aorta-gonad-mesonephros
Was shown that embryo had no CFU-S at E8
Yolk sac actually had v few CFU-S
Have blood cells at day 8 but no cfu-s
When testing whole embryo instead of just yolk sac
Note colonies came from body of embryo than the yolk sac
Appeared at similar time - E9 instead of 7/8
When are HSCs first detectsble
HSCs detectable by transplants only appear at E10.5-11.5
So how can the blood cells in yolk sac/embryo body be there on earlier days
Haematopoietic sites in embryo
Yolk sac
Chorion (becomes placenta)
Alontois
Foetal liver
AGM
How can blood cells in yolk sac appear in absence of HSCs?
Two alternative models
Model 1- two haematopoietic hierarchies exist
One transient and one permanent
Embryonic ones exist earlier but willl expire
And the definitive hierarchy coming from HSCs appear a bit later
Model 2- have common progenitor that gives rise to embryonic hierarchy and also to the definitive adult haematopoietic hierarchy
Model 1 or model 2?
One thing to note - yolk sac has large nucleated erythrocytes
V diff to adult ones
So adult peripheral erythrocytes are coming from HSCs while yolk sac ones come from yolk sac
MODEL 2 IS NO LONGER SEIOUSLY CONSIDERED - 1 IS PREFERABLE MODEL
Why is model 1 preferable? (2 separate haematopoietic systems - ephemeral embryonic and definitive adult)
Graft developing quail body onto chick embryo - replacing chick body
Discriminate chick v quail cells by nucleus structure
Before hatching shown that majority of cells in circulation were quail in origin (body)
Despite the fact that first blood cells come from yolk sac (chick tissue)
Supporting fact that 2 diff ones arise
Dye injected into xenopus blastocyst cells
Track origin of dorsal aorta
And the ventral blood island (yolk sac equivalent)
Can see even at early stage the origins of these tissues are v diff
Backs up 2 separate haematopoietic system model
Which hameatopoietic system comes from which region (at least in xenopus)
Transient embryonic one from yolk sac
Permanent definitive one comes from aorta gonad mesoneohros region AGM
Pros of mouse model (even tho zebra fish development is more visible)
Mammalian model
Inbred mines
Sequences genomes/good genetics
Accessible genome for manipulation (transgenesis, KO, ES cells)
Developed functional assays
Cons of mouse model over zfish
Develop inside mother. - harder to see - limited experimental access
More expensive to keep
Obtaining sufiicient numbers if embryos for certain experiments can be problematic
Quantitating on of HSCs assay
Limiting dilution analysis
Cells in H system migrate so looking at sections can be hard to find origin
Can count colonies on sleep
Or can do LDA
Can calculate no of HSCs in certain region
Prepare cell suspension
Then transplant into cohort of mice at certain dilution where some mice survive and some do not
One HSC is sufficient for rescue of HP system - dead mouse didn’t even receive one
Means that nice that survived statistically likely only got 1 HSC (or maybe a couple)
Can use the number of surviving mice to count the number of HSCs in the cell suspension transplanted and hence the region it was prepared from
Emergence of HSCs in embryonic tissues
Comparing AGM and yolk sac
No significant difference between them at E10.5
Increased LDA rescue of rats at same amount by E11.5
So is it AGM or YS producing the HSCs
Do transplantation assay to see when they are maturing (can’t rescue when immature ig)
Maturation from their embryonic precursors
Jumps from total of 4 in embryo at day 11 to 130 day 12
Way too fast for proliferation (cell cycle too slow)
So must be maturing from another cell type already pregnant
Where to HSCs mature from?
AGM region (and not the yolk sac) can autonomously generate definitive HSCs
3 waves of haematopoiesis in embryo
1 - maturation and gradually disappear
But recently shown that tissue macrophages (or circulating ones) arise from yolk sac??)
2- more potent erythroid progenitors appearing at day 8 (1 day later than w1)
Wave 3- HSCs appear at day 10.5-11.5
HSCs will take month to begin producing blood cells
Need some for embryo
So need to have the early blood cells for function in embryo