International non- proprietary name: Cetuximab
Brand name: Erbitux
Target; Format: Erbitux EGFR; Chimeric IgG1
Indication first approved or reviewed: Colorectal cancer
First EU approval year: 2004
First US approved: 2004
Kuby Immunology EIGHTH EDITION
Lecture PowerPoint
CHAPTER 8
T-Cell Development
Copyright © 2019 by W. H. Freeman and Company
Punt • Stranford • Jones • Owen
Development of T cells in the thymus
Early T-cell precursor development occurs in the bone marrow
T-cell precursors begin their travel through the thymus at the cortex
T-cells that survive selection migrate into the medulla
Development of T cells in the thymus
Cells migrate to the thymus for further development
There, they go through a variety of different stages
Double negative (DN) cell has no CD4 or CD8 (CD4-CD8-)
Double positive cell (DP) is both CD4+CD8+
Positive/negative selection stages for a cell to become single positive CD4+ or CD8+
Development of T cells in the thymus
Final screening removes autoreactive cells
Release into the peripheral bloodstream
Recombination of TCR gene segments also occurs in the DN stages, yielding either an αβ or a γδ T cell
Early thymocyte development
When cells arrive at the thymus, they aren’t technically T cells
They can become NK cells, dendritic cells, B cells, and myeloid cells
A receptor known as Notch commits them to the T lineage
GATA-3 transcription factor becomes activated
Notch binding can commit cells to T lineage in vitro without the thymus being present
Early thymocyte development
Thymocytes progress through four DN stages
Each stage varies in expression of several key molecules
C-kit (CD117)—receptor for stem cell growth factor
CD44—an adhesion molecule
CD25—the α chain of the IL-2 receptor
TCR rearrangement begins in the cortex at the DN2 stage
Table 8-1, Double-negative thymocyte development, Page 296
Phenotype Location Description
DN1 c-Kit (CD117)++, CD44+, CD25– Bone marrow to thymus Migration to thymus
DN2 c-Kit (CD117)++, CD44+, CD25+ Subcapsular cortex TCR γ-, δ-, and β-chain rearrangement; T-cell lineage commitment
DN3 c-Kit (CD117)+, CD44–, CD25+ Subcapsular cortex Expression of pre-TCR; β-selection
DN4 c-Kit (CD117)low/–, CD44–, CD25– Subcapsular cortex to cortex Proliferation, allelic exclusion of β-chain locus; α-chain locus rearrangement begins; becomes DP thymocyte
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Early thymocyte development
Thymocytes can express either TCRαβ or TCRγδ receptors
TCRβ rearrangements are one of the first to take place and one of the most likely to be productive
Because of this, TCRαβ outcomes are more likely than TCRγδ
TCRγδ are more common in fetal development
Fetal developmental environment may provide different signal cues
As TCRγδ T cells are less common, the remainder of this chapter will focus on TCRαβ T cells
Early thymocyte development
DN thymocytes undergo β-selection, resulting in proliferation/differentiation
A successfully produced β chain is paired with the pre-Tα chain
A 33 kDa protein surrogate for real TCRα chain
Allows for formation of a pre-TCR complex (with CD3 proteins) and many early signaling events
Early thymocyte development
After β-selection has occurred, thymocytes are at the DP stage of development
Functional TCRα chain replaces surrogate pre-TCRα
The cell still expresses both CD4 and CD8, i.e., CD4+CD8+ (double positive)
Pos/neg selection occurs, yielding mature single positive T cell, CD4+ or CD8+
Positive and negative selection
CD4+CD8+ DP thymocytes make up 80% of thymic cells
These cells undergo thymic selection
Positive selection
Selects thymocytes bearing receptors capable of binding self-MHC molecules with low affinity, resulting in MHC restriction
Negative selection
Selects against thymocytes bearing high-affinity receptors for self-MHC/peptide complexes, resulting in self-tolerance
Most cells (95%) fail positive selection and fail to receive needed survival signals
Die by apoptosis
Positive and negative selection
Thymocytes “learn” MHC restriction in the thymus
A classic experiment in mice illustrates this principle
A strain (A × B) F1 animal’s immune system was purposely wiped out with radiation
Its thymus was then replaced with one from a parental B strain
Bone marrow from a sibling (A × B) F1 was used to re-form the immune system
Positive and negative selection
A classic experiment in mice (continued)
When challenged with strain A virally infected target cells, the cells weren’t destroyed
Inability to select T cells recognizing strain A cells
When challenged with strain B virally infected target cells, the cells were destroyed
The new thymus and bone marrow selected T cells that could recognize strain B cells
Positive and negative selection
T cells undergo positive and negative selection
Cortical thymic epithelial cells express high levels of MHC class I and II
Developing T cells can “browse” possible self-peptide/MHC complexes
These present self-peptides; three possible outcomes when T cells encounter these self-peptide/MHCs
TCRs can’t bind; cells die by neglect
TCRs bind too strongly; negative selection, apoptosis occurs
TCRs bind “just right”; positive selection to single-positive stage occurs
Positive and negative selection
Positive selection ensures MHC restriction
TCR that can bind MHC-peptide shifts T cell from DP to SP
If the TCR can bind to an MHC class II molecule, it also binds with the CD4 molecule, selecting the cell to the CD4+ subset
The opposite happens if the TCR binds to an MHC class I molecule, resulting in selection to the CD8+ subset
Positive and negative selection
Negative selection (central tolerance) ensures self-tolerance
Clonal deletion-induction of apoptosis in cells with too strong anti-self signaling/binding
Do we delete thymocytes reactive to tissue-specific antigens?
Not all tissue types are in the thymus
How does screening against these tissue antigens take place?
Autoimmune regulator (AIRE) protein induces expression of many tissue-specific proteins in medullary thymic epithelial cells
AIRE binds epigenetic marks on histones to recruit transcription factors
New T cells can be screened against these antigens safely in the thymus
Positive and negative selection
Negative selection (central tolerance) ensures self-tolerance
Other mechanisms of self-tolerance have been postulated and have some experimental support
Clonal arrest—autoreactive T cells are prevented from maturing further
Clonal anergy—autoreactive T cells are inactivated, not deleted
Clonal editing—second or third chances at rearranging a non–self-reactive TCR α gene
Clonal deletion remains the best proven and most common method of tolerance induction in the thymus
Positive and negative selection
The selection paradox: Why don’t we delete all cells we positively select?
Affinity model—strength of signal received is critical
Support found in the OT-I TR transgenic mouse system
All TCRs are of one type that can recognize one peptide
The MHC class I molecules on thymic epithelial cells (cTECs) have no, low, or high affinity for their peptide
Degree of selection for/against CD8+ SP T cells is determined
Positive and negative selection
An alternative model can explain the thymic selection paradox
The altered peptide model
Self-peptides produced by thymus epithelial cells are unique and distinct from peptides made by other cells
Thus, thymocytes positively selected by such interactions wouldn’t be negatively selected by later interactions
Still under investigation—some evidence that thymus cells process antigens differently from other cells
The two theories aren’t mutually exclusive—multiple mechanisms of selection may exist
Positive and negative selection
Do positive/negative selections occur at the same stage of development, or in sequence?
Most likely that negative selection can occur at various points in development
Positively selected cells must express CCR7 chemokine receptor to move to medulla for further development and selection/screening
The situation is likely complex, but the medullary region appears to be quite important in removing autoreactive T cells
Lineage commitment
Several models have been proposed to explain lineage commitment
Instructive model
TCR/CD4 and TCR/CD8 coengagement generate unique signals
The signals generated “instruct” the T cells which lineage to fully commit to
Lineage commitment
Several models have been proposed to explain lineage commitment
Stochastic model
Positively selected thymocytes randomly downregulate CD4 or CD8
Only those cells with the “correct” coreceptor receive signals to continue development
Strength of signal and duration of signal from TCR/coreceptor
Lineage commitment
Several models have been proposed to explain lineage commitment
These models may be too simplistic
Kinetic signaling model
Cells commit to the CD4 lineage if they receive a continuous signal
Cells commit to CD8 lineage if stimulation signal is interrupted
IL-7 promotes CD8 differentiation of interrupted thymocytes
Lineage commitment
DP thymocytes may commit to other types of lymphocytes
NKT cells
Express a TCR with an invariant TCRα chain
Interact with CD1 molecules presenting lipid antigens
Intraepithelial lymphocytes (IELs)
Usually CD8+, but also have features of innate immune cells
Regulatory T cells (TREG)
CD4+ subset that helps to quench adaptive immunity
Signaling cues for alternative development unclear at present
Exit from the thymus and final maturation
A cascade of events controls final maturation stages
Upregulation of Foxo1 transcription factor
Expression of Klf2, which upregulates sphingosine-1-phosphate (S1P) receptor
S1PR required to help T cells leave the thymus
Foxo1 also upregulates IL-7R (giving survival signals) and CCR7 (a chemokine receptor that helps cells exit and move to lymph nodes)
T cells that have just exited the thymus are recent thymic emigrants (RTEs)
They’re not as functionally mature (yet) as older cells—an active area of research
Other mechanisms that maintain self-tolerance
TREG cells negatively regulate immune responses
Belong to a subset of CD4 T cells characterized by expression of FoxP3 transcription factor
Developmental cues unclear
TREG cells function to:
Deplete the local area of stimulating cytokines
Produce inhibiting cytokines
Inhibit APC activity
Directly kill T cells
Other mechanisms that maintain self-tolerance
Peripheral mechanisms of tolerance also protect against autoreactive thymocytes
Some self-antigens are “hidden” because APCs lack the correct costimulatory molecules needed to initiate immune responses
Some self-antigens are presented by non-APCs, preventing initiation of autoimmunity
Strong self-antigen signaling through the TCR in the absence of co-stimulation may drive the T cells into anergy (nonresponsiveness)
Summary
Developing T cells (thymocytes) arise from multipotent CD4–CD8– precursors that migrate from the bone marrow to the thymus
Mature T lymphocytes have a diverse TCR repertoire that is tolerant to self-antigens yet restricted to self-MHC
The fate of a CD4+8+ thymocyte depends on the affinity of its TCR for self-peptide/MHC complexes encountered on stromal cells in the two major thymic microenvironments: the cortex and medulla
Mechanisms that remove autoreactive T cells during development, central tolerance, are reinforced in the periphery by a variety of mechanisms, including the activity of regulatory T cells
CD4+ and CD8+ thymocytes that survive positive and negative selection are allowed to migrate from the thymus into the bloodstream and complete their maturation in the periphery