T cell tolerance overview Flashcards
What is immunological tolerance?
specific unresponsiveness to an antigen that is induced by exposure of lymphocytes to that antigen
What is central tolerance?
tolerance induced through immature cells in primary lymphoid organs
What is peripheral tolerance?
tolerance induced through mature cells in secondary lymphoid tissue
What is central T cell tolerance?
deletion of T cells that strongly recognise self-antigen in the thymus or do not interact with MHC
What are thymocytes?
developing T cells
What are the 3 tolerance checkpoints in the thymus?
b-selection; positive selection; negative selection
What is the result of the independent development of the TCR repertroire?
some TCRs are self-MHC/p reactive and some are unable to bind to MHC/p
What is the function of b-repetoire selection?
is the new beta chain function
When does b-selection occur?
to pro-T cells wjho then go on to become pre-T cells
What happens if the new b-chain is function?
cell survival and division; TCRa gene rearrangement; differentation to double positive cell
How do the haematopoietic cells enter the thymus?
cortico-medullary junction
Where does positve selection take place?
cortex
What is the function of positive selection?
selection of thymocytes bearing receptors able to bind self-MHC molecules
What cells are involved in positive selection?
cortical thymic epithelial cells
What happens to cells that do not bind self-MHC?
death by neglect
What happens to cells without a function b chain?
death by apoptosis
What is hte function of negative selection?
eliminates thymocytes bearing high-affinity receptors for self-MHC molecules alone or self-antigen
Which cells are involved in negative selection?
medullary thymic epithelial cells and DCs
What do medullary thymic cells involved in negative selection express?
AIRE- autoimmune regulatory
What percentage of thymocytes survive selection?
5%
What is the name for double positive cells?
immature thymocyte
What happens after positive selection?
single positive T cell
What is the mutation in autoimmune polyglandular sydrome 1?
in AIRE gene
what is the result of a mutation in AIRE gene in APS-1?
defective negative selection in thymuc leading to the release of autoreactive T cells
Which organs does APS-1 affect?
endocrine glands
What other name is APS-1 given?
APECED- autoimmune polyendocrinopathy candidasis ectodermal dystrophy
What is anergy?
state of long-lasting partial/total unresponsiveness induced by partial activation
What are the 2 modes of inducing anergy?
antigen recognition and 1-absence of costimulation or 2-ligation of co-inhibitors
When is CTLA-4 expressed on T cells?
after activation
What is hte difference in affinity for CD80 and CD86 between CTLA4 and CD28?
CTLA4 has a much higher affinity for B7 molcules than CD28
Why is CTLA4 expression transient?
rapidly internalised
What is the function of CTLA4?
tolerance induction; limiting the immune repsonse
What are the 3 methods of peripheral tolerance?
anergy; deletion and Treg suppression
When does deletion of T cells take place?
chronic/persistent stimulation with self-antigen ; high/excessive dose of self-antigen
What are the mechanisms for deletion?
active- Fas/FasL or PD-1/PD-L1 interaction or passive- IL-2 deprivation
How was the function of Tregs discovered?
normal mouse with thymectomy 3-5 days after birth develops polyautoimmune disease which can be prevented by giving CD4+ CD25+ cells
What is CD25?
IL-2 receptor alpha chain
What are Tregs positive for?
CD4+ CD25hi; CD127lo; CTLA4 and FOXP3
What are the mechanisms of regulation by Tregs?
contact-dependent: inactivation of DCs or activated T cells; contact-independent: secretion of immunosuppressive cytokines; consumption of IL2
What immunosuppressive cytokines do Tregs secrete?
TGF-b; IL-10; IL-35
What is the difference between flow cytometry histogram and dot plot?
histogram looks at a single parameter whereas dot plot looks at 2
How are natural Tregs created?
in the thymus when recognise self-antigen
How are inducible Tregs created?
recognition of repeated self antigen in peripheral tissues
Where do natural Tregs live?
in peripheral tissues to prevent harmful reactions against self
How are Tregs induced in vitro?
stimulation of CD4 cells in presence of TGF-b and IL-2
What have mice models shown about the use of Tregs in disease?
mice with EAE improve after infusion with Tregs; the onset of DM in NOD mice is delayed by Treg infusion
How have Tregs been found to be related to human disease?
SLE and immune thrombocytopenia patietns have fewer Tregs ; IPEX have dysfunction of Tregs
What is the mutation in IPEX?
in FOXP3- transceiption factor for Tregs
What does IPEX stand for?
immune dysregulation; polyendocrinopathy; enteropathy and X-linked syndrome
What is antigen segregation?
physical barrier to self-antigen and happens at immunologically priviledged sites so cells have never been toleraised against hte auto-antigen
Why is PD1/PD-L1 signalling a fine balance?
if too low results in autoimmunity but if too high allows tumour growth
What is the name for cancer therapeutics utilising PD-1 signalling?
immune checkpoint inhibitors
What is foudn in the thymic stroma?
in young individuals this is a large network of epihtelia containing large numbers of dveloping T-cell precursors
What signalling initiates TCR rearrangement?
Notch signalling
What does the thymus differentiate from embryologically?
third pharyngeal pouch
What type of cells are found in the thymic cortex?
immature thymocytes and scattered macrophages
what cell types are found in the thymic medulla?
mature thymocytes; DCs; macropahges and some B cells
What mutations in humans and mice results in thymic hypoplasia?
DiGeorge- 22q11 deletions; mouse- nude mutation
What cells give the progentiro cells Notch1 signals?
thymic epithleial cells
What is the function of Notch signalling?
instructs the progenitor to commit to the T cell lineage rather than B cell, and initate T cell specific gene expression programming
Which transcription factors does Notch singalling induce the expression of?
T-cell factor-1 (TCF1) and GATA3
Give examples of genes that TCF1 and GATA3 induce?
those encoding components of CD3 complex and Rag1 (gene rearangements)
What is the third transcription needed to induce the entire program of T cell gene expression?
Bcl11b
What are apoptotic bodies?
residual condensed chromatin of apoptotic cells
What indicates that macropahges are responsible for removing apoptotic cells in the thymus?
apoptotic bodes are seen inside macrophages throughout the thymic cortex
When does the T cell gene expression programme begin?
in DN1 cells
What do DN1 cells express?
CD44 and Kit
What do DN2 cells express?
CD44 and CD25
At what stage are cells irreversibly committed to T cell lineage?
DN2
What happens during DN2?
begin to rearrange b-chain
What happens as the DN2 cell rearranges its b-chain?
become CD44lo and Kit lo forming DN3 cells
Why are DN3 cells arrested in development?
until prodtively rearragnet the b chain locus
What is formed during DN3?
b chain pairs with a surrogate chain pTa to form the pre-TCR
What does the presence of the pre-TCR on the cell surface trigger?
cessation of b-chain rearrangement and entry into the cell cyel and massive proliferation
What happens when the DN3 cells undergo massive proliferation?
lose CD25
What happens once the DN3 cell has lost CD25 expression?
known as DN4
What happens when DN4 cells stop proliferating?
cell expresses CD4 and CD8
What happens once the cell is double positive?
begin rearranging alpha locus
What transcription factor governs maturation into CD4 single positive cells?
ThPOK
What transcription factor governs maturation into CD8 single positive cells?
Runx3
What is the fist cell-surface molecule specific for T cells?
CD2 or Thy1 (in mice)
What other populations of cells are double negative in the thymus?
T cells expressing y:d TCRs and T cells bearing a:b TCRs of very limited diversity (iNKT)
What is Kit?
receptor for stromal-cell factor
What gene rearrangement takes place in DN2 cells?
DJ rearrangement
What gene rearrangement takes place in DN3 cells?
V to DJ rearrangement
What is the function of the transient expression of CD25 in developing thymocytes?
unclear- T cells in IL2 knockout mice develop normally
What is the function of Kit signalling?
mice lacking Kit have a much smaller number of DN T cells
What cells produce IL-7?
thymic stroma
What happens to mice knocked out of IL-7; IL-7 receptor or its signalling protein Jack3?
severe block in T cell development
How do the pre-TCRs dimerise?
pTa Ig domain associates with the Ig domain of the V subunit to form the pre-TCR itself then it binds to a V domain of another pre-TCR molecule
What does dimerization of the pre-TCR result in?
ligand-independent proliferation; stops b-chain rearrangement and souble positivity
What type of DP cell does the alpha positive locus begin to rearrange in?
small DP cell- once the large DPs have ceased to proliferate
How long is the time between entry of a T cell progenitor into the thymus and the export of its mature progeny in the mouse?
around 3 weeks
Which area of the thymus does most T-cell development take place in?
cortex
Where do DN3 cells reside?
subcapsular region of hte cortex
Where do cells migrate after entering through the cortico-medullary junction?
into the outer cortex then move back through as immature DPs to the medulla where only mature single-positive cells are
What cells make up the cortical stroma?
epithelial cells with long branching processes that express both MHC-I and -II
How are the thymic epithelial cells arranged around the thymocytes?
form a network surrounding them
When do cells move from hte cortex to the medulla?
after positive selection
Where are y:d T cells mainly found?
epithelial and mucosal sites
what are the first cells to develop in the fetal thymus?
y;d t cells
What happens to the first y:d T cells developed in the fetal thymus?
go to epidermis, w edged in the keratinocytes and take on a DC form—dendritic epidermal T cells –secrete keratinocyte growth factor and inflammatory chemokines and cytokines
What is the function of the second y:d T cells developed in the fetal thymus?
reside in the mucosal epithelia of tissues and dermis of the skin and produce IL-17
What makes hte y:d T cells produced during fetal development invariant?
composed of the same Vy and Vd regions
Why are there no N-nucleotides in the dETCs and Td;y-17 cells ?
fetal thymocytes do not express TdT
What is the function of the leader sequences in front of V segments?
encodes a signal sequence to direct chain to ER
Why does the pre-TCR at the surface stop further beta-chain development?
causes the phosphorylation and degradation of RAG-2
What does the pre-TCR signal constitutively via?
Lck
What happens to mice deficient in Lck?
T cell development is arrested before DP stage and no a gene rearrangements are carried out
Why are RAG-1 and RAG2 suppressed in DN4 prolfieration?
ensures that each cell which gives rise to a successful b chain gives rise to lots of CD4 and CD8 cells (no rearrangement occurs before proliferation has finished)- increases chance of one of them making a functional a chain
What is the function of ZAP-70 in T cell development?
promotes the development of single-positive thymocytes from the DPs
What alows many successive V to J rearrangements to take place?
the presence of multiple V gene segments and J segments which can allow successive rearrangement events to leapfrog over previous VJ segments, deleting any intervening ones
What is the difference between the cessation of gene rearrangement between B and T cells?
B cells- final assembly of immunglobulin stops it whereas T cells have to get signals from a self-pepdie:self MHC complex that positively selects the receptor
Why can T cells express 2 different alpha chains?
because expression of the TCR is not enough to halt rearrangement so allows different a chains to be produced
If a T cell expresses 2 different alpha chains does that not change the understanding taht a single functional specificity is expressed by each cell?
only one of the 2 receptors is likely to recognise peptide presented by a self MHC molecule so the other won’t have any function
What do the y:d subsets produced early in fetal development derive from?
HSCs from the fetal liver, not bone marrow
How long do DP cells last unless their TCR is engaged?
3-4days
What is the rescue of DP cells from programmed cell death and their maturation into SP cells called?
positive selection
How many T cells generated will be able to recognise self peptide; self MHC complexes?
10-30%
What gave the evidence that self-MHC complexes are required for the survival of immature T cells?
transgenic mice are given rearranged TCRs, and the cells only mature past the double-positive if they express the same MHC as the mouse from which teh rearranged TCR genes were originally developed
Is the specificty of the unselected repertoire of TCRs completely random?
no, if the specificity of unselected repertoire were completely random, only a very small proportion of thymocytes would be expected to recognise any Mhc molecule but there appears to be MHC-specificity encoded in the germline V gene segments
What creates the specificty of TCRs for MHC?
In the V segments, in the CDR1 and CDR2 regions which are highly variable have certain amino acids conserved
What is the positivity of thymocytes when they undergo positive selection?
double positive
Aside from selecting for TCRs that can bind MHC, what other function does positive selection have?
determines the cell-surface phenotype and function potential of the mature T cell by selecting the appropriate co-receptor for efficient entigen recognition
How was the role of ThPOK discovered?
through a naturally occurring mutation in mice which lacked CD4 development
How was the role of the thymic epithelium in positive selection discovered?
MHC-II gene was placed under a promoter restricted to thymic cortical epithelial cells which was then introduced as a transgene into MHC class II mutant mice and CD4 development was restored
What is expressed by thymic epithelial cells, which most cells do not express, and its absence results in severely impaired CD4 development?
cathepsin L- have high numbers of mhc-II on surface that are still associated with CLIP
What is expressed by thymic epithelial cells, not expressed by others, which if absent, prevents CD8 devleopment?
proteasome subunit b5T (since proteasome- peptide issue?)
When does negative selection take place in T cell development?
either when DP or SP- dependent on where the T cell encounters the antigen
What is the affinity hypothesis?
the choice between positive and negative selection is thought to hinge on the strength of the self peptide:self MHC binding by the TCR r
What is agonist selection?
the process of selection of positively selected cells receiving signals slightly weaker than those inducing negative selection differentiate into Tregs
What cytokine is required for Treg devleopment but not other T cell developmnet?
IL-2
What gives invariant NKT cells their name?
express NK1.1 receptor commonly foudn on NK cells
What receptor do iNKT cells recognise?
CD1
What do iNKT cells require to mature?
TCR interaction with CD1 and a signal through the adaptor protein SAP
Where do iNKT cells migrate to after leaving hte thymus?
peripheral lymphoid tissues and mucosal surfaces
What stimulates T cell emigration from the medulla
the S1P receptor is upregulated in SP cells and binds S1P which is found in high conc. in the blood and lymph
What do mature thymocytes express that facilitates localisation of naive T cells to peripheral lymphoid organs?
L-selectin which is a lymph node homing receptor
What other cell type aside from Tregs develop in response to agonist signalling?
iNKT cells
What stage of develpment does DJ b chain rearrangement take place?
DN2
What stage of development does V to DJ development take place?
DN3
What stage does functional b hcain rearrangement take place?
DN4
What stage is the pre-TCR first expressed?
DN3
What effect does ThPOK have on Runx3?
represses its expression
What explains MHC restriction of mature T cells?
CD4 and CD8 bind up intracellular Lck, it is necessary that the TCR binds is an MHC protein so it also bdins to CD4 or to CD8 and initiates isgnalling cascades
What antigenic quality correlates with self in the periphery?
reocnigition in the absence of danger signals produced by the innate immune system
What is functional deviation in the context of peripheral tolerance?
induction of T cell development instead of effector T cell development
Which cells express tissue-specific antigens in the thymus?
thymic epithlelial cells in the medulla and CD8a+ DCs
What are cells ignorant of self?
most circulating lymphocytes have a low affinity for self antigen but make no effector response to them
When can ignorant but self-reactive cells be recruited to AI responses?
if threshold for activation is exceeded- e.g by APC presenting that antigen and high levels of costimulation
When would unmethylated CpG sequeneces in DNA be found?
in apoptotic mammalian cells
How could TLR9 be responsible for autoreactive B cells?
if there is extensive cell death with inadequate clearance of apoptotic fragments then B cells can internalise unmethylated CpG fragments that then react with TLR9 activating the ignorant B cell
How is rheumatoid factor produced?
when immune complexes with IgG form during infection or immunisation, they are multivalent and is able to cross-linke IgG BCRs thereby creating anti-IgG antibody
How do immunologically priviledged sites prevent immune responses?
extracellular fluid from the site does not pass through conventional lymphatics; surrounded by tissue barriers to exclude naive lymphocytes; TGF-b is produced in the tissue; expression of Fas ligand at the site to induce apoptosis to lymphocytes entering the site
give examples of immunologically privileged sites?
brain; eye; testis; uterus (fetus)
What is regulatory tolerance?
tolerance due to Tregs who can suppress self-reactive lymphocytes that recognise antigens different from those recognised by the Treg cell
What is the key feature of regulatory tolerance?
Tregs can suppress autoreactive cells that recognise a variety of different self antigens, as long as the antigens are from the same tissue or are presented by the same APC
what is the function of natural Tregs?
when activated by the same antigen in the periphery, nTregs inhibit other self-reactive t cells that recognise antigens in the same tissue to prevent their differentiation into efefctor T cells or prevent their effector function- if encounter their antigen on APC, secrete cytokines which inhibit all surrounding auroreactive T cells
what is the function of FOXP3 negative regulatory cells?
produce IL-10 in the intestinal tissues
Why is discrimination between self and non-self imperfect?
a proper balance must be struck between preventing AI disease nad preserving immune competence
