Module 3: Tolerance
Generation of B and T cell receptor diversity by gene-rearrangement
1011 different unique BCR and 1016 unique TCR
B and T cells are not encoded by a single gene but are generated by random selection of one
V, one D and one J segment
Segment combining by recombination activating gene (RAG)
Fused VDJ DNA segment encodes antigen binding site
Deliberately inaccurately
o At the junctions nucleotides are deleted and added
More possibilities
On one cell all the receptors are the same
Tolerance
The immune system prevents autoimmunity due to these self-reactive cells at
two levels
The first levels is called the central tolerance, because it takes place in
the central lymphatic organs
o Will result in the removal of the majority of self-reactive
lymphocytes
Surviving lymphocytes enter blood lymph circulation
as mature but still naïve lymphocytes
Different mechanisms prevent that self-reactive lymphocytes that
have entered the periphery will be activated. This is referred to as
peripheral tolerance
T cell tolerance: Central tolerance of T cells (positive and negative selection)
Stem cells for B and T cells are formed in bone marrow. B cells mature in the bone marrow, while T
cell precursors move towards the thymus for their development into mature T cells
T cell precursor enters thymus in subcapsular region and moves to cortex
TCR rearrangement takes place
o If this results in a non-functional TCR the T cell dies by apoptosis
o Productive TCR rearrangement leads to a pre-T cell that expresses a TCR and the CD4
and CD8 co-receptor
The pre-T cells are subjected to positive selection
o in which those cells that are able to recognize self MHC molecules on thymic
epithelial cells, with sufficient affinity, will continue their development
Pre-T cells that bind too strongly to self-antigen-MHC complexes on thymic epithelia cells are
eliminated by apoptosis. This process is called negative selection
The mature, naïve T cells that remain are able to enter the circulation
o Positive/negative selection eliminates 90% of pre-T cells
The key factor in determining positive and negative selection is the strength of the antigen
recognition by the maturing T cell; low-avidity recognition leads to positive selection, and high-avidity
recognition induces negative selection
Selection of CD4+ or CD8+ T cells in the thymus
Progenitors of T cells initially express both CD4 as well as CD8 coreceptors
Positive selection takes place after low avidity recognition with MHC/(self)peptide complexes
o If this interaction between TCR is with MHC-I, then the CD8 co-receptor will interact
with the MHC-I molecule and is kept, while the CD4 co-receptor will be lost: the T cell
becomes a CD8+ T cell
1
, o if the TCR interacted with MHC-II the CD4 co-receptor will interact with the MHC-II
molecule and is kept, while the non-binding CD4 co-receptor is lost: the T cell
becomes a CD4+ T cell
Role of AIRE and TRAs
Tissue restricted antigens (TRAs): are antigens that are only expressed in certain specific tissues
If these antigens would not be expressed in the thymus then negative selection for these
antigen would not occur and auto-reactive T cells would leave the thymus
Thymus has a special mechanism for expressing these TRAs in medullary thymic epithelial cells,
allowing deletion of auto-reactive T cells specific for these autoantigens
Expression of these TRAs is under the control of the autoimmune regulator (AIRE) protein
o In the absence of AIRE, autoreactive cells would be escape deletion and be able to
respond to self-antigens in the periphery
o AIRE has been linked to the development of natural regulatory T cells that are
involved in maintaining tolerance in peripheral tissues
o AIRE has also been discovered in secondary lymphoid tissues, on stromal cells
Central tolerance of T cells (development of Tregs)
The CD4+ T cells that bind self-antigens in the thymus quite strong, but not strong enough to be
eliminated by negative selection are selected to become CD25+ regulatory T cells, or Tregs
These cells would otherwise be likely candidates to induce autoimmunity
Tregs prevent autoimmunity by secretion of suppressive cytokines (TGFβ and IL10) upon
recognition of their MHC/(self-)peptide complex
Peripheral tolerance of T cells: Anergy and Treg induction
If a T-cell, that does not recognize a self-antigen, slips through negative selection and binds the
antigen, anergy or unresponsiveness may occur. Because when a T cell recognizes an MHC/peptide
complex (signal 1) without co-stimulation, the T cell will not be activated
MHC/peptide recognition (signal 1) by naive T cells in the absence of co-stimulation (signal 2) can
also result in the induction of Tregs
There are several methods to create an anergic cell:
Create a signalling block, whereby the lymphocyte activation signal no longer reaches its
target inside the cell
Activation of inhibitory receptors CTLA-4 and PD-1 upon self-recognition
o CTLA-4 and PD-1 are found on the T lymphocytes and are able to bind to CD80/86
present on antigen presenting cells
CD80/86 receptors are essential in co-stimulation and activation of the T
lymphocytes
o By binding CTLA-4 to CD80/86, the T cell activation signal is blocked
o PD-1 can recognize the ligands PD-L1 and PD-L2. Binding of either ligand will result in
a signal block as well
two postulated mechanisms of action of CTLA4:
delivery of inhibitory signals that block TCR- and CD28-mediated signals
engagement of B7 molecules on APCs so they are inaccessible to CD28
Other mechanisms of peripheral tolerance
2
Generation of B and T cell receptor diversity by gene-rearrangement
1011 different unique BCR and 1016 unique TCR
B and T cells are not encoded by a single gene but are generated by random selection of one
V, one D and one J segment
Segment combining by recombination activating gene (RAG)
Fused VDJ DNA segment encodes antigen binding site
Deliberately inaccurately
o At the junctions nucleotides are deleted and added
More possibilities
On one cell all the receptors are the same
Tolerance
The immune system prevents autoimmunity due to these self-reactive cells at
two levels
The first levels is called the central tolerance, because it takes place in
the central lymphatic organs
o Will result in the removal of the majority of self-reactive
lymphocytes
Surviving lymphocytes enter blood lymph circulation
as mature but still naïve lymphocytes
Different mechanisms prevent that self-reactive lymphocytes that
have entered the periphery will be activated. This is referred to as
peripheral tolerance
T cell tolerance: Central tolerance of T cells (positive and negative selection)
Stem cells for B and T cells are formed in bone marrow. B cells mature in the bone marrow, while T
cell precursors move towards the thymus for their development into mature T cells
T cell precursor enters thymus in subcapsular region and moves to cortex
TCR rearrangement takes place
o If this results in a non-functional TCR the T cell dies by apoptosis
o Productive TCR rearrangement leads to a pre-T cell that expresses a TCR and the CD4
and CD8 co-receptor
The pre-T cells are subjected to positive selection
o in which those cells that are able to recognize self MHC molecules on thymic
epithelial cells, with sufficient affinity, will continue their development
Pre-T cells that bind too strongly to self-antigen-MHC complexes on thymic epithelia cells are
eliminated by apoptosis. This process is called negative selection
The mature, naïve T cells that remain are able to enter the circulation
o Positive/negative selection eliminates 90% of pre-T cells
The key factor in determining positive and negative selection is the strength of the antigen
recognition by the maturing T cell; low-avidity recognition leads to positive selection, and high-avidity
recognition induces negative selection
Selection of CD4+ or CD8+ T cells in the thymus
Progenitors of T cells initially express both CD4 as well as CD8 coreceptors
Positive selection takes place after low avidity recognition with MHC/(self)peptide complexes
o If this interaction between TCR is with MHC-I, then the CD8 co-receptor will interact
with the MHC-I molecule and is kept, while the CD4 co-receptor will be lost: the T cell
becomes a CD8+ T cell
1
, o if the TCR interacted with MHC-II the CD4 co-receptor will interact with the MHC-II
molecule and is kept, while the non-binding CD4 co-receptor is lost: the T cell
becomes a CD4+ T cell
Role of AIRE and TRAs
Tissue restricted antigens (TRAs): are antigens that are only expressed in certain specific tissues
If these antigens would not be expressed in the thymus then negative selection for these
antigen would not occur and auto-reactive T cells would leave the thymus
Thymus has a special mechanism for expressing these TRAs in medullary thymic epithelial cells,
allowing deletion of auto-reactive T cells specific for these autoantigens
Expression of these TRAs is under the control of the autoimmune regulator (AIRE) protein
o In the absence of AIRE, autoreactive cells would be escape deletion and be able to
respond to self-antigens in the periphery
o AIRE has been linked to the development of natural regulatory T cells that are
involved in maintaining tolerance in peripheral tissues
o AIRE has also been discovered in secondary lymphoid tissues, on stromal cells
Central tolerance of T cells (development of Tregs)
The CD4+ T cells that bind self-antigens in the thymus quite strong, but not strong enough to be
eliminated by negative selection are selected to become CD25+ regulatory T cells, or Tregs
These cells would otherwise be likely candidates to induce autoimmunity
Tregs prevent autoimmunity by secretion of suppressive cytokines (TGFβ and IL10) upon
recognition of their MHC/(self-)peptide complex
Peripheral tolerance of T cells: Anergy and Treg induction
If a T-cell, that does not recognize a self-antigen, slips through negative selection and binds the
antigen, anergy or unresponsiveness may occur. Because when a T cell recognizes an MHC/peptide
complex (signal 1) without co-stimulation, the T cell will not be activated
MHC/peptide recognition (signal 1) by naive T cells in the absence of co-stimulation (signal 2) can
also result in the induction of Tregs
There are several methods to create an anergic cell:
Create a signalling block, whereby the lymphocyte activation signal no longer reaches its
target inside the cell
Activation of inhibitory receptors CTLA-4 and PD-1 upon self-recognition
o CTLA-4 and PD-1 are found on the T lymphocytes and are able to bind to CD80/86
present on antigen presenting cells
CD80/86 receptors are essential in co-stimulation and activation of the T
lymphocytes
o By binding CTLA-4 to CD80/86, the T cell activation signal is blocked
o PD-1 can recognize the ligands PD-L1 and PD-L2. Binding of either ligand will result in
a signal block as well
two postulated mechanisms of action of CTLA4:
delivery of inhibitory signals that block TCR- and CD28-mediated signals
engagement of B7 molecules on APCs so they are inaccessible to CD28
Other mechanisms of peripheral tolerance
2
