Presented by

Somen Kumar Mistri

Department of Microbiology

University of Dhaka

Dhaka, Bangladesh

ImmunologicalTolerance

Central Tolerance

How does the immune system provide a high degree of sensitivity and specificity to the broad array of pathogens without attacking self?

How does a T-cell know whether to be CD4+ T-cell or CD8+ T-cell?

How does T-cells come in contact with all the self antigens in thymus?

What is the role of IgD in the development of B-cell tolerance?

Questions to consider

Tolerance

Central(Bone marrow , Thymus)

Peripheral(Spleen , Lymph node etc)

Central Tolerance

Thymic tolerance to self antigen

(In Thymus)

B-cell tolerance to self antigen

(In Bone Marrow)

Generation of T-cell and B-cell antigen specific repertoire involves the elimination of self-reactive

cells

Thymic tolerance to self antigens

Thymus - Site of T-cell Maturation

Cortex

Medulla

Overview of T-cell Development, Maturation, Migration and Activation

In a young mouse , the thymus contains around 108 to 2×108 T-cells.

Each day about 5×107 new T-cells are generated.

Only 106 to 2×106 (Roughly 2 - 4%) of these will leave the thymus each day as mature T-cells

So approximately 98% of T-cells that develop in thymus also die within the thymus.

Death is usually by apoptosis.

Some Interesting Facts About T-cell maturation

T-cells undergo two types of

selection process:

Positive Selection

Negative Selection

Recognition of Self -MHC

Avoid strong binding to self antigen

T-cells see antigen in context of MHC molecule. T-cell development requires TCR to recognize MHC class I or II

T-cell Development In Thymus

Thymocyte Selection Occurs In Thymic Architecture

T-cell Development In Thymus…. Positive

selection

1 2 3 4

Particular combinations of cell surface proteins can be used as markers for T-cells at different stages of differentiation.

Two distinct lineages of T-cell: α:β and γ:δ. When progenitor cells first enter the thymus from bone

marrow, they lack most of the surface molecules characteristic of mature T-cells. Their TCR genes are not rearranged at that time.

In fact, proteins such as RAG1 and RAG2 (Recombination Activating Gene ) are not expressed that are required for rearrangement.

At first T-cells do not contain :

CD3 : TCR complex CD4 or CD8

These cells are called DN thymocytes (Double negative)

T-cell Development In Thymus….

Once DN1 cells encounter the thymic environment , they begin to proliferate and express CD25.

During DN2 stage , rearrangement of the genes for TCR γ , δ , β chains begins. RAG expression gets turned on at this stage.

Cells destined to become γδ T-cells (˂5% of mature lymphocytes) diverge at the transition between DN2 and DN3.

After DN3 stage DN4 stage comes. RAG expression stops here and proliferation of DN4 cells occurs.

T-cell Development In Thymus….

Formation of pre-TCR activates a signal transduction pathway that has several consequences :

Indicates that a cell has made a productive TCR β-chain rearrangement and signals its further proliferation and maturation.

Suppresses further rearrangement of TCR β-chain genes, resulting in allelic exclusion.

Renders the cell permissive for rearrangement of the TCR α-chain.

Induces the developmental progression to the CD4+ 8+ double positive (DP) state.

T-cell Development In Thymus….

Newly formed DP cells reactivate RAG genes allowing rearrangement of the α-chain.

Pairing of α and β-chains allow T-cells to recognize a wide range of antigens (Both self and foreign).

At this point positive selection occurs.Cells that recognize MHC with moderate affinity on

cortical epithelial cells survive. Cells whose TCR interacts with MHC class I becomes

CD4+ T-cell (SP).Cells whose TCR interacts with MHC class II becomes

CD8+ T-cells(SP).Cells whose TCR fails to engage either a class I or class II

MHC molecule undergo programmed cell death.

T-cell Development In Thymus….

Thymocyte Selection Occurs In Thymic Architecture

Positive selection

Proliferation

Negative Selection•Positive selection involves recognition of self MHC at cortical epithelial cells.

Mechanism not really known.•Negative selection predominantly in medulla (Medullary epithelial cells)

Thymocytes Undergo Two Types of Selection Process In Thymus

Positive selection for thymocytes bearing receptors capable of binding self-MHC molecules, which results in MHC restriction. Cells that fail positive selection are eliminated within the thymus by apoptosis.

Negative selection that eliminates T-cells bearing high affinity receptors for self-MHC molecules alone or self antigen presented by self-MHC, which results in self-tolerance.

Binding of TCR and MHC (weak), results in positive selection

of T-cells

NEGATIVE SELECTION

APC

T-cell

MHC

TCR

POSITIVE SELECTION

Weak binding of

TCR and MHC-Peptide

complex :T-cell is positively selected

Strong binding of TCR and MHC-

peptide complex : T-cell is negatively

selected

APC

T-cell

MHC

TCR

Self antig

en

APC

T-cell

MHC

TCR

Positive and Negative Thymocyte Selection

Engagement of TCR by the MHC-peptide complex on some type of antigen presenting cell underlies both positive and negative selection. Positive and negative selection may occur at low and high degrees of TCR ligation, respectively.

Experiments show that the same peptide will induce positive selection at low concentration and negative selection at high concentration.

This has led to the avidity model, which postulates that a functionally low avidity interaction between T-cell and peptide-MHC involving a relatively low number of TCRs will positively select Double positive (DP) CD4+8+ thymocytes, while a high avidity interaction will lead to clonal deletion.

The overall avidity of the T-cell interaction is a function of :

Ligand Density × TCR Density × Affinity

Factors Affecting Positive and Negative Selection In The Thymus

Factors Affecting Positive and Negative Selection In The Thymus ….

It is postulated that a low avidity interaction between the T-cell and APC will give positive selection and that high avidity will give deletion.* Refers to affinity of peptide for the MHC or of the MHC-peptide complex for the TCR

The autoimmune regulator (AIRE) is a protein that in humans is encoded by the AIRE gene. AIRE is a transcription factor expressed in the medulla of the thymus and controls the mechanism that prevents the immune system from attacking the body itself.

AIRE allows endothelial cells in thymus as well as dendritic cells to express other proteins that they normally do not express.

Due to the presence of AIRE medullary epithelial cells as well as dendritic cells present self peptides in low amount to T-cells. This serves the negative selection purpose.

Mutation in AIRE results in Autoimmune Polyendocrinopathy–Candidiasis–Ectodermal Dystrophy (APECED).

Role of AIRE (Autoimmune Regulator)

B-cell tolerance to self antigens

Overview of B-cell Development, Maturation, Migration and Activation

B-c

ell

Deve

lopm

ent

B-cell development shows similar features to T-cell development, but takes place largely in the bone marrow.

Checkpoints in B-cell development include : Successful expression of Igα and Igβ in late pro-B cells. Successful rearrangement at the heavy chain locus in

pre B-cells. Successful rearrangement at the light chain locus and

receptor editing.Mature B-cells express IgD at higher levels than IgM.Self tolerance begins when IgM first appears at the surface

of the developing cell.

B-cells Pass Through Several Developmental Checkpoints

B-cell Development In Bone Marrow

/ Immature B

Expression of IgD is an important checkpoint in terms of eliminating self reactive B-cells.

Normally in a mature B-cell, when Antigen binds to mIg of that B-cell, it turns the B-cell on.

However, before IgD is expressed, if antigen (Self) binds to mIg then the B-cell gets turned off. Further development of that B-cell does not occur, but light chain rearrangement can continue.

Why IgD is Important To Make A B-cell Mature?

B-cells That Do Not Detect And Bind To Self Antigen Can leave The Bone Marrow

If in bone marrow, IgM binds to a non-cross linking self molecule it can not give strong signal to the B-cell to die, instead it generates a B-cell that may not be as responsive or clonally ignorant.

Autoreactive B-cells Die or Can undergo Light Chain Receptor Editing (Sequential VJ Joining)

Autoreactive B-cells Die or Can undergo Light Chain Receptor Editing (Sequential

VJ Joining)….

Allowing light chain rearrangement to continue among immature cells permits the B-cell to edit its receptor and rescue potentially autoreactive cells from inevitable death.

Thus by changing the light chain, a self reactive B-cell can be converted to a non-self reactive one.

After making a new light chain, the mIgM is again tested to see if this is also self reactive.

If the heavy chains association with antigen is very strong, then light chain rearrangement alone can not alter self reactivity. Ultimately that B-cell would not be allowed to leave the bone marrow.

Autoreactive B-cells Die or Can undergo Light Chain Receptor Editing (Sequential VJ Joining) …

Some B-cells Become Anergic

If cross linking occurs, but least sufficiently to produce strong signal, the B-cell does not die by apoptosis. It migrates to the periphery and actually become anergic B-cells. Even though they are self reactive they won’t make antibodies. Therefore they won’t cause any kind of autoimmune diseases.

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