T cell development, Receptor repertoire selection and CD4/CD8 lineage commitment Flashcards
Where do T cells come from?
- Multipotent lymphoid.
Progenitors migrate from the bone marrow to the thymus through blood vessels
- In the thymus, the lymphoid progenitors differentiate to pre-T cells and are educated to differentiate self from non-self
- Positively selected T cells emigrate from the thymus to mediate and effect the cognate immune response
Cells in the thymus produce chemokines
Thymosin
Thymotaxin
Thymopetin
Thymic factors
Chemical attractants which secreted and transported via blood to bm, T cells recognise these and follow the gradient to the thymus
Thymus is divided into
cortex
medulla - where late phase of T cell development within thymus occurs
What cells are present in the thymus
Cortical epithelial cell
Medullar epithelial cell
Lymphocytes interspersed within the thymus
Other cell types play important role later in T cell development e.g macrophages, dendritic cells
Hassal’s corpuscle is where aggregation of mature lymphocytes occurs
Thymocytes at different developmental stages are found in distinct parts of the thymus
immature T cells come via the blood into the thymus they do not express functional TCR or core receptors CD4/8.
Proceed to have interactions with stromal cells and other cell types, double positive lymphocytes will express CD4 and CD8 , and mature CD3 or T cell receptor
Finally proceed towards the final stage- generation of a single positive mature CD4 or CD8 T lymphocyte (not double positive)
Then exit thymus
Key players
T cells:
- TCR
- CD4
- CD8
These are on surface of T cell as a T cell receptor complex, they can both bind (TCR and CD4/8) to MHC class molecules.
Non-T cells
- MHC I
- MHC II
On other cells, not T cells
CD = cluster of differentiation
MHC = major histocompatibility complex
What cells can you identify on a flow cytometry?
Can identify cells that are either only cd4 positive
Or CD8 - single positive
Neither – double negative
CD4 and CD8 – double positive
DN’s can be further subdivided into DN1 through to DN4
Where do double negative cells appear?
What cells are dominant in thymus?
Appear in the fetal thymus before double positive cells
Thoughout further maturation more cells appear DP.
DP in thymus are dominant population. Different from other secondary lymphoids like spleen where majority of cells committed to CD4/8, no DP.
Does T cell differentiate and TCR rearrangement occur separately?
T cell differentiation and t cell receptor rearrangement occurs simultaneously so that the double positive CD4/8 cells express mature t cell receptor ready for selection
Gamma delta T cells diverge from the main pathway of T cell development, at the stage of DN2.
These cells undergo little further changes before exiting thymus.
Alpha beta have different pathway.
What T cells are favoured during early fetal development?
Gamma delta T cells
Show up during early gestation, subside progressively, numbers go down in adults, only represent small proportion of all T lymphocytes in circulation
Alpha beta T cells other way around, progresisvely increase their numbers, present as main T cell type in adults
Antigen recognition by gamma delta T cells is different than alpha beta t cells
How are they different
Gamma delta T cells bearing specific receptors end up in skin (Vg5), gut (Vg2), uterus (Vg6) etc
Gamma delta T cells not MHC restricted, doesn’t recognise MHC presented peptides as conventional T cells do
Instead antigen is recognised directly, more like an antibody
Some cases ligands for the gamma delta TCR are self-proteins upregulated under stress conditions
In humans circulating gamma delta cells recognise a phospholipid antigen from mycobacterium tuberculosis – rather than peptide fragments
Play a role in cancer surveillance
What % of T cells are gamma delta
Gamma delta 10% of all generated T cells, alpha beta is 90%
Alpha beta T cells are characterised by expression of the alpha beta TCR, what is the process of assembly of alpha beta T cell?
Original germline chain of both alpha and beta are rearranged multiple times
Different fragments combined
Assembly of functional alpha beta t cell receptor is achieved
Progression through development correlates with rearrangement
- Maturation of T cell receptor takes place simultaneously with the transition of the T cell in terms of phenotype from DN to DP to SP cell
- Beta chain recombines during double negative stage
- Alpha chain recombines and rearranges later, during transition to double positive
Why do DP thymocytes need to progress to the SP stage?
What signalling supports transition to SP?
Needs functional TCRa chain rearrangement , a cell without a functional TCR can’t serve its purpose
Both CD4 and MHC II – needed to be a CD4+ cell
Or needs CD8, MHC I and TAP to be a CD8+ cell
Needs important molecules involved in cell signalling that support transition to SP
- ERK signalling
- Calcineurin signalling
What happens to cells that fail to complete thymocyte maturation (95%)?
95% cells fail either positive or negative selection
Therefore undergo death by neglect, by apoptosis
These cells are removed from tissue by macrophages, phagocytic cells that ingest them and recycle some components
Describe apoptosis process of cells
Non-select cells are removed from tissue
Programmed cell death
Controlled and supported by right processes within environment
Doesn’t cause damage to other cells or tissues
Acquires interaction between sFas (soluble Fas), present on surface of apoptotic cell
And Fas ligand, and other cell types that can initiate apoptotic cell death
Following the interaction, signalling pathway will be activated
Ends up with expression of active caspase 8, which will disintegrate DNA or cleave it, and kill the cell
DP cells have both CD4 and CD8 on surface
How do DP bind to MHC molecules?
What happens if binding is not strong enough?
DP cells will look for MHC molecules in thymic tissue
MHC molecules may be present on epithelial cells or APC, also present in thymic tissue.
Random event whether CD4 or CD8 will first find either MHC I or MHC II.
Whichever finds the target first will bind to the molecule, e.g DP binding to CD4 to MHC II on another cell, if binding is strong enough it will result in down regulation of the CD8.
Therefore DP cell will emerge as a single positive cd4 cell with very little or no CD8 expression at all.
Vice versa, CD8 on DP cells, first it finds targets or MHC I molecule on another cell, binds to it. If binding is strong enough CD4 is downregulated and DP cell emerges as single cd8 positive cell.
Binding is not strong enough, double positive cells will be killed by apoptosis or neglect. Eventually removed from tissue.
Most cells fail to bind to MHC I or II and will not be selected as CD4 or CD8 cells.
What is MHC in humans?
Human leukocyte antigens of MHC I and II type
MHC I = HLA- A, B, C
MHC II = HLA –DR, DQ, DP
HLA-A / MHC I expressed on thymic stromal cells and low level on APC (DC and macrophages)
HLA-DR / MHC II expressed on thymic medullary stromal cells and high level on APC
When does positive selection of T cells occur?
Positive selection occurs when DP T cell with a functional TCR binds to HLA molecules
If binding is strong enough, cell will be positively selected and allowed to continue development.
When binding is weak/no binding, the cell will be useless and be removed by apoptosis.
What does positive selection ensure?
Positive selection ensures that only T cells that are useful and can engage in recognition are selected. All other cells are removed
DP CD4/CD8 cells bind to MHC-I or MHC-II on thymic epithelial cells – it is a random event which one binds.
Following adequate binding of CD4:MHC-II, CD8 is downregulated and vice versa. Allows transition from DP to SP.
From here on, the SP CD4 or CD8 T cells are ready for negative selection.
Unselected cells die by apoptosis.
What is negative selection?
What problem do T cells have with this? how is it overcome?
Exclusion of self-reactive T cells as they would cause autoimmunity
Determined based on affinity of TCR for presented self-peptide: high – kill him, low – keep him
Eensures that remaining T cells are only reactive to foreign peptides
Self-reactive cells are not removed immediately, go through further TCR rearrangements (second chance) - before they are eventually removed if still self-reactive
Problem for T cells: thymus does not represent all self-antigens
But has a transcription activator gene which can induce expression of other tissue specific proteins (kidney, heart, etc)
Gene is called AIRE (autoimmune regulator): this allows negative selection against most bodily self-proteins
How does AIRE allow T cells to negatively select against self-antigens not present in thymus?
Transcription factor autoimmune regulator (AIRE) mediates ectopic gene expression in the thymic medullary stroma
other tissue specific genes, kidney, heart, liver, lungs, gut, apart from brain and testes – might be expressed if their transcription is activated by AIRE
Known as promiscuous gene expression – about 10% of all genes in the thymus are expressed this way
Remaining pool of self-reactive cells smaller
Without AIRE, immature thymocytes/t-cells negatively selected against a limited number of self proteins
Positive and negative selection summary altogether
Positive selection
- DP CD4/CD8 cells that bind either to MHC I or II
- Depending on strength of binding will have different fates
- Strong binding = useful, self restricted and progress towards SP CD4 or CD8
- Weak binding cells will be killed by apoptosis
SP cells can go to negative selection
- TCR binding to MHC peptide complex which contains self-peptides that can be found in the thymus
- Self-peptides presented by thymic cells, stromal cells and APCs like macrophages and dendritic cells
- APCs will also present some self-peptides found in other tissues due to action of AIRE
Antigen presentation and MHC expression control thymocyte slection
What cells present antigen?
- DC
- Medullary epithelial cells
- Cortical epithelial cells
- Endothelial cells
Certain cell types control different T cell developmental fates
- Positive selection
- Negative selection
- Receptor editing
How are regulatory T cells formed? What surface antigens do they express?
What is their main role?
Some SP T cells do not immediately exit thymus and go to other tissues and lymphoid organs.
They remain in the thymus for a short period of time, undergo stages of differentiation where they express CD25 and FoxP3 surface antigens
These cells do not proliferate in response to MHC self-peptide complexes
T regs accumulate in Hassall corpuscles and later migrate to different tissues
Main role: dampen T cell response, regulating it when further accumulation of T cells are no longer required
What happens once T cells pass both positive and negative selection?
Become conventional T cells
Migrate to secondary lymphoid organs looking for their target antigen
Immunological synapse
If they encounter specific antigen, they get activated, proliferate and become effector T cells
Some become memory T cells
If they don’t find the target they eventually die by apoptosis after period of circulation
