Summary module 3 Immunotechnology (CBI-30806)

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

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, 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




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