Transcript No Slide Title

Lecture 9- T cell development
•Flow cytometry- how it works
•Thymic architecture and cells
•Thymic development I- generation of a TCRb chain
•Thymic development II- gd vs ab T cell development
•Positive selection of CD4+CD8+ ab T cells.
•Negative selection of high affinity autoreactive cells.
•Alloreactivity, recognition of the MHC molecules of others.
Fluorescence
Fluorescence 2
1-10 x 10-9 seconds
Fluorescence 3
Figure 2-13
Examples of commonly used fluorescent probes
Green fluorescent protein
Fluorescein
Couple to antibody
T cell development quality control- phase I
b-chain selection
• As with B cell development, T cells develop in an ordered
sequence.
• First, there is TCRb chain gene recombination, starting
with Db-to-Jb.
• Next, Vb-to-DJ rearrangement occurs. As rearrangement
is random and error-prone, b-chain rearrangement is
tested for functional protein using the surrogate chain
mechanism (preTa).
T cell development occurs in the thymus
Neonatal removal of the thymus, or congenital lack of the thymus
(DiGeorge Syndrome, nude mice) leads to T cell deficiency
Scanning electron micrograph
of thymus
Developing T cell
Thymic stroma
Thymocyte maturation
(approximate)
V(D)J
recombination
Positive
selection
Negative
selection
The first step in ab T cell development, TCRb rearrangement
and testing for protein.
Figure 5-6 part 1 of 3
Figure 5-6 part 2 of 3
b-chain+
T cell development quality control- phase II
• Cells with a good first chain then proliferate a bit, then
stop and redirect rearrangements at the second locus,
with Va-to-Ja recombination. At this time, preTa is no
longer expressed.
• In contrast to B cells, T cells that make an intact receptor
are screened for "positive selection" based on low affinity
for self-MHC/self-peptides.
• Cells that have no affinity for self-MHC/self-peptide
complexes are eventually lost by apoptosis.
Thymocyte maturation
(approximate)
V(D)J
recombination
Positive
selection
Negative
selection
The ability of CD8 T cells to recognize MHC class I bound
peptides and CD4 T cells to see MHC class II bound peptides
is not random, but is selected for during thymic education.
• Before selection, cells
expressing TCR a/b
coexpress both CD4 and
CD8 and are called
"double positive cells".
• These cells, which are the
major cell population in the
thymus, mature when their
TCRs see MHC (carrying
self-peptides) with a low
affinity.
• The coreceptor (CD8 or
CD4) engaged regulates
the fate decision to CD8 or
CD4 "single positive"
differentiation.
• Hence, CD8 cells have
TCRs that were selected
on class I, and CD4 cells
have TCRs that were
selected on MHC class II.
T cell populations during development
and in the lymphoid tissue
THYMUS
Fluorescence color 2
CD4
SPLEEN
Fluorescence color 1
CD8
T cells failing
positive
selection die
in the cortex
and are
immediately
engulfed by
macrophages.
Red stain
shows dead
cells, blue
stain,
macrophages.
Figure 5-5
Thymocyte maturation
(approximate)
V(D)J
recombination
Positive
selection
Negative
selection
T cell development quality control- phase III
Negative selection
•Cells that bind with too high an affinity for self-MHC/selfpeptide complexes are lost, probably by apoptosis.
U. of Cambridge
Figure 5-3 part 2 of 2
Figure 5-13
TCRa/b CD8+4+ double positive thymocytes
that fail to be positively selected can undergo
many rounds of TCRa recombination in an
attempt to generate a functional receptor.
Figure 5-19
T cell developmental stages
are identified by cell surface
markers and gene
rearrangement status.
A major checkpoint involves the
testing of functional TCRb for ability
to pair with the surrogate TCRa
chain, preTa.
Kuby et al.
W. H. Freeman & Co. and Sumanas, Inc.
Immunology, January, 1997
Molecular basis of direct
allogeneic MHC recognition
Concepts in a/b T cell development
• TCRa/b cells develop in the thymus from bone marrow
progenitors.
• TCRb is rearranged first and tested in association with
pTa.
• Cells that express TCRb protein then express CD4+CD8
and rearrange TCRa genes.
• Cells carrying appropriate a/b TCRs undergo positive
selection, leading to expression of just CD4 or CD8.
• Autoreactive T cells undergo negative selection in the
medulla.
• The result of selection is a skewing of the T cell repertoire
that promotes MHC restricted recognition.
• Thymic selection also leads to skewing toward
alloreactivity and high frequency reactivity to allografts.