L4 - Lymphocyte development Flashcards
Where do all cells come from?
All cells come from bone marrow & become pluripotent haemopoietic stem cells
B cell development
Develop from haematopoietic stem cells in bone marrow that express PAX5 transcription factor
Continuous process: 3 x 1010 produced per day
Lots of cells don’t leave the bone marrow – most don’t survive when screened for receptors & self-reactive cells
Involves rearrangement & expression of Ig genes
Why do lots of cells not leave the bone marrow?
Most don’t survive when screened for receptors & self-reactive cells
What is CD45?
A key marker of lymphocytes
What is CD19?
A slightly later marker of B cells – not expressed in early B cells
Where do the final stages of development from immature B cells happen?
In peripheral lymphoid tissues (eg. spleen)
What does PAX5 do?
PAX5 turns on genes that are going to turn into a B cell – interacts with bone marrow stromal cells
What happens once PAX5 has turned on the right genes?
During the process the B cell will now start to make the randomly re-arranged cell receptor to put on its surface
First ones it makes are IgM & IgG isotypes
What happens if the receptors produced can recognise antigens in the bone marrow?
If receptor can recognise antigens, those B cells are given a signal to die
If they recognise nothing they exit the bone marrow & survive – has the ability to recognise something that isn’t expressed in that environment
THIS IS NEGATIVE SELECTION
Where do the B cells that survived negative selection go?
Go through the circulatory system to lymphoid organs/nodes
What happens if the receptors produced can recognise antigens in lymphoid organs/nodes?
If the receptor now recognises something, it is a potential antigen it needs to respond to
Cells with the right receptor proliferate – clonal expansion
Differentiate & become plasma cells to secrete the antibody instead of displaying it on the surface
THIS IS POSITIVE SELECTION
Why do not all B cells become plasma cells secreting antibody?
Some become memory cells & keep BCR on their surface – can remain for 10-20 years
Majority of cells won’t recognise an antigen as you’re not infected so will die after a few days
B cells die at the same rate they are made
Pre-B cells
H chain genes rearrange first
Ig-mu chain expressed with surrogate light chain – product of VpreB & lamda-5 genes
Why is a surrogate light chain used?
Tests whether heavy chain is going to be any good
Simple models to test binding of the heavy chain
If it fails it dies straight away
Where does the H chain bind the surrogate light chain?
H chain has to move to the surface to bind to the surrogate light chains
Associates with Ig-alpha & Ig-beta (invariant molecules) to do this
What happens if H chain binds the surrogate light chain?
Sends a signal to the B cell if it can bind to the surrogate light chain & can move to the next stage of development
Next: make proper light chains instead of surrogate light chains
Pre-B cell receptor pre-BCR
Signal from pre-BCR:
• Turns off RAG1 & RAG2 genes – don’t make another heavy chain
• 5-6 rounds of cell division
• Surrogate light chain expression stops
• RAG1 & RAG2 turned on again – allows light chains to be made
• L chain rearrangement starts
What are RAG genes needed for?
Gene rearrangement - are regulated in time
What types of L chain can B cells express?
Kappa or lamda
Each rearrangement has a 1 in 3 chance of being successful
As kappa chain rearranges first, more B cells express kappa than lambda
Immunoglobulin gene rearrangements
Error prone
Dies if it doesn’t productively re-arrange
Pre-B cells with non-productive rearrangements of light chain genes can be ‘rescued’ by up to 5 further rearrangements at the same locus – if after all these attempts still out of frame then lambda locus begins to rearrange
What happens to immature B cells that bind multivariant self-antigens?
Undergo either:
• Clonal deletion – cell dies by apoptosis
• Receptor editing – further L chain rearrangements of variable regions (another chance)
What happens to immature B cells that bind soluble self antigens?
Become unresponsive - anergic
Can escape death – can downregulate BCRs that are potentially autoreactive
T cell development
Initially similar to that of B cells – Originate from bone marrow stem cells – Re-arrange receptor genes – Express pre-T receptor – Elimination of self-reactive T cells by negative selection
How is T cell development different from B cell development?
Have 2 more robust selection methods – positive & negative selection
Undergo development/selection in the thymus
• Either rearrange alpha/beta TCR genes (CD4+ & CD8+) or gamma/delta TCR genes
T cells expressing alpha/beta TCR must bind with self MHC expressed in thymus
• Positive selection
TCR binds self-MHC
Cannot bind with too much affinity otherwise will die
Vast majority of T cells fail this test – 95% die
5% leave & go into blood & lymph nodes
Alpha/beta T cell development
Precursors produced in bone marrow migrate to thymus
Once in the thymus they develop into thymocytes:
- Re-arrange TCR genes (beta first) & express TCR
- Acquire other markers – eg. CD3, CD4 & CD8
- Undergo positive & negative selection
What is the thymus?
Bi-lobed organ in anterior mediastinum
Each lobe divides into many lobules
Each lobule has outer cortex & inner medulla
What cells are present in the thymus?
Lymphoid cells
Epithelial cells
Macrophages & dendritic cells
What happens in the thymus?
Where T cells are selected for their ability to recognise self-MHC & then if they survive they are killed if they recognise it too well
T cell maturation in thymus
Pro-thymocytes enter cortex via blood vessles from bone marrow
Re-arrange TCR genes
– Firstly re-arrange TCR-beta genes
– Expressed along with the pre-TCR-alpha (invariant): pre-T cell receptor
– Cells proliferate & then re-arrange TCR-alpha genes
Express TCR together with CD3
Also express CD4 & CD8
What does TCR expression require?
CD3 complex – essential for transmitting signals
– Delta, epsilon & gamma chains
– Zeta chain dimer
CD3 transmits signal to T cell nucleus following TCR recognition of p/MHC
Gamma/delta TCR
- Similar structure to alpha/beta receptor
- No CD4 or CD8 markers
- Less diversity than alpha/beta
- Expressed on separate T cell population
- Recognise different antigens
- Only go through positive selection
Lineage commitment to gamma/delta or alpha/beta depends upon which genes are first to rearrange successfully
T cells expressing a randomly rearranged alpha/beta TCRs may be able to recognise what MHC?
- Recognise self MHC plus peptide from ‘foreign’ Ag – immunity
- Recognise self MHC plus peptide from ‘self’ Ag – autoimmunity
- Not be able to recognise self-MHC – useless
Need to keep T cells with TCR 1, eliminate those with TCR 2 & 3 – positive & negative selection
Reality is a fine balance based in TCR/MHC affinity
Positive selection of T cells
Positive selection of cells which recognise self MHC (+ self-peptide)
Occurs when double-positive (DP, CD4+, CD8+) T cells recognise MHC on cortical epithelial cells in thymus – apoptosis if not recognised
Positively selected cells move to medulla
Proposed models for the role of CD4 & CD8 in thymic positive selection of DP thymocytes leading to SP T cells
2 possible theories:
- Instructive model
Interaction of 1 co-receptor, with MHC molecules on thymic stromal cells during MHC I or II mediated positive selection results in the down regulation of the other co-receptor - Stochastic (random) model
Down-regulation of the co-receptor occurs randomly in DP cells, prior to MHC-mediated positive selection
Those cells that have down-regulated CD4 will be selected on MHC I, & those that have down-regulated CD8 will be selected on MHC II
Negative selection of T cells
Negative selection of T cells which recognise self MHC (+ self-peptide) on thymic dendritic cells/macrophages with high affinity (as these T cells may induce autoimmunity if not removed)
TCR binding to MHC/self-peptide with high affinity causes T cell to die by apoptosis (clonal deletion)
TCR affinity for p/MHC is basis for selection
All T cells recognising self-MHC/self-peptide are positively selected
Those with the highest affinity TCR are then negatively selected
Ultimate goal = a population of T cells with low affinity for self-peptide + self MHC
These cells represent the safest/most useful to keep with the highest probability they will have a high affinity for self MHC when presenting peptides derived from pathogens
What happens to T cells that survive thymic selection?
< 5%
Express TCR capable of binding self MHC
Are depleted of self-reactive cells
Exit thymus as mature, single positive (SP) T cells
What are the types of SP T cells?
CD4+ SP T cells recognise Ag in association with MHC class II
CD8+ SP T cells recognise Ag in association with MHC class I
