The Thymus as The Primary Site of

T-cell Production
 Thymus Function

Jacques Miller (1961):

 Thymectomy (or congenital absence of Thymus in
Di George syndrome)
Fewer total lymphocytes
Reduced graft rejection
Reduced antibody responses

 Therefore T lymphocytes = cellular immune responses

– Miller (1961) Lancet ii: 748

 Thymic Atrophy Results In An Age-Related
Decrease In T-cell Production

Number of New T-cells produced per day

0 Age in years 70

In health, thymus atrophy is okay because T-cells are long lived

 The Thymus Consists Of Two Main Cellular Elements:

 HAEMOPOIETIC bone marrow/fetal liver derived

precursors which develop into T-cells

 Non-haemopoietic STROMAL CELLS, that drive the

development of T-cell precursors

Both elements are required for normal T-cell production

 Stromal Microenvironments In The Thymus

Subcapsular
Capsule Epithelium Dendritic
cell Macrophage

Medullary
Cortical Epithelium
Epithelium

Blood
vessel

Cortex Medulla
 T-cell Development Can Be Separated Into Distinct Stages
Lack expression
of CD4 and CD8
Termed CD4-CD8- or
‘double negative’

Dual expression Increasing

of CD4 and CD8 Maturity
Termed CD4+CD8+ or
‘double positive’

CD4 expression only CD8 expression only

‘CD4 single positive’ ‘CD8 single positive’
CD4+CD8- CD4-CD8+
 Stages Of T cell Development In The Thymus

Subcapsular
Capsule Epithelium Dendritic
cell Macrophage

DP
DP

Medullary
DN Cortical Epithelium
Epithelium

DN SP
Blood
vessel
DN
Cortex Medulla
 Two Main Checkpoints in T-cell Development

Checkpoint 1: Maturation from the CD4-8- to CD4+CD8+ Stage

Checkpoint 2: Maturation from the CD4+CD8+

to the CD4+ or CD8+ Stage
CD4+
CD8-

CD4- CD4+
CD8- CD8+

CD4-
CD8+
 Maturation from the CD4-CD8- to CD4+CD8+ Stage

Events include:

 Commitment to the T-cell lineage

 Cellular expansion (proliferation)

 Rearrangement of genes encoding the T-cell receptor

 Commitment to the T-cell Lineage

Thymic epithelial cells

give a Notch signal
Bone marrow/
Fetal liver
ab T-cells
precursor

gd T-cells Dendritic cells

Natural
Killer (NK) cells
Cellular Expansion of Early T-cell Precursors in Thymus

 Controlled by soluble growth factors (Interleukins)

CD4-CD8- Thymocyte Thymic Epithelial Cell

Express: Produce:

IL-2 Receptor
Interleukin-7 Receptor Interleukin-7
Interleukin-15 Receptor Interleukin-15
c-Kit (Stem Cell Factor receptor) Stem Cell Factor
 Cell Proliferation of Early T-cell Precursors in Thymus

 A role for Stem Cell Factor and IL-7: studies on mice

lacking IL-7 Receptor and c-Kit
Gene Knockout Mouse Thymus: Wildtype Mouse Thymus:
Genes encoding IL-7 Receptor
and c-Kit deleted

Less than 120 million

1 million thymocytes
thymocytes
 Rearrangement of T-cell Receptor Genes
T-cell Receptor b chain gene

Variable (V) regions Diversity (D) Joining (J) Constant (C)

regions regions regions

Fully rearranged gene

TCR b protein
 Selecting Cells Bearing Fully Rearranged
TCRb Chain Genes: A function of the Pre-T cell Receptor

Pre-T cell Receptor Complex

 Expressed only on CD4-CD8- thymocytes

Consists of TCRb protein, CD3 complex, and pre-Ta

Signalling through pre-TCR is essential for the generation

Of CD4+8+ thymocytes
 Selecting Cells Bearing Fully Rearranged
TCRb Chain Genes: A function of the Pre-T cell Receptor

Precursor
undergoing Cell Death
TCRb Unsuccessful
rearrangement rearrangement

Successful TCRb
rearrangement  Stops TCRb
Pre-Ta rearrangement
(Allelic exclusion)
CD3
 CD4, CD8 expression
Signalling
 TCRa rearrangement

 Cellular expansion
The Need For Positive and Negative Selection in Thymus

Unlike B-cells, T-cells can only recognise processed antigen

presented by self Major Histocompatibility Complex (MHC) molecules

Antigen
T-cell
Presenting
Cell
The Need For Positive and Negative Selection in Thymus

Millions of CD4+CD8+ thymocytes, each expressing an

abTCR complex of differing specificity

Because rearrangement of TCR genes is random,

we need to select on the basis of MHC recognition:

USELESS: Don’t recognise peptide/MHC at all

USEFUL: Recognise peptide/MHC of low affinity or avidity

HARMFUL: Recognise peptide/MHC at high affinity or avidity

Need to: Retain USEFUL specificities (POSITIVE SELECTION)

Remove HARMFUL specificities (NEGATIVE SELECTION)

 Models of Positive and Negative Selection

Affinity Model: Affinity refers to the ‘fit’ between a receptor

and its ligand, so
Low affinity TCR-peptide/MHC interactions give positive selection
High affinity TCR-peptide/MHC interactions give negative selection

Avidity Model: Avidity refers to the number of receptors engaged by

ligand, so
Low avidity TCR-peptide/MHC interactions give positive selection
High avidity TCR-peptide/MHC interactions give negative selection

In both models, Negative Selection occurs via APOPTOSIS

The Cortex: Positive Selection of Thymocytes

CD4+CD8+

CD4 CD8
TCR

MHC II MHC I

Cortical Epithelium
The Cortex: Positive Selection of CD4+ Thymocytes

TCR and CD4 binds CD4+CD8+

to MHC Class II
on thymic CD4 CD8
epithelium TCR
MHC II
MHC I

Loss of CD8
Expression Cortical Epithelium

CD4

CD4+8-
T-cell
The Cortex: Positive Selection of CD8+ Thymocytes

CD4+CD8+ TCR and CD8 binds

to MHC Class I
CD4 CD8
on thymic
epithelium
MHC II TCR
MHC I

Loss of CD4
Cortical Epithelium Expression

CD8

CD4-8+
T-cell
 The Medulla: Negative Selection of Thymocytes

CD4+CD8- CD4-CD8+

CD4 CD8
TCR TCR
MHC II
MHC II MHC I
MHC I

Medullary Epithelium Medullary Epithelium

High affinity/avidity: Negative selection

Low affinity/avidity: Survival
Conclusions

1. The thymus is essential for T-cell development

2. Stromal cells in the cortex and medulla support

T-cell development

3. The most immature thymocytes are CD4-CD8-. They

signal through the Pre-T-cell Receptor to develop to the
CD4+CD8+ stage.

4. CD4+CD8+ thymocytes express the abT-cell receptor.

These cells are SELECTED to produce CD4+ and CD8+ T-cells
that bind MHC in a non-autoreactive way.