Immunology - Chapter 8 T-Cell Development
What exactly are Tregs and what are the 4 ways they can function?
A fraction of thymocytes that experience high-affinity TCR interactions do not die by negative selection but develop into FoxP3+ regulatory T cells (TREG cells) that inhibit T-cell responses outside the thymus. These cells require help from cytokines, including IL-2 and IL-15, to complete their maturation. Regulatory T cells that develop in the thymus are called thymic TREG cells. Regulatory T cells that differentiate in the periphery are called peripheral TREG cells. The function and location of both TREG populations overlap, although they may have distinct responsibilities in different tissues. SO Tregs are DP thymocytes that express α and β chains but that are self reactive (so they react to our own MHCs pretty strongly). but they function to dampen the immune response when it is to strong. They do this in four different ways. 1.Cytokine deprivation: TREGs express relatively high levels of high-affinity IL-2 receptors and can compete for the cytokines that activated T cells need to survive and proliferate. - they can work through cytokine deprivation. Tregs express a receptor for IL-2 which is a pretty powerful cytokine. By having this receptor they are able to bind to IL-2 that is released by a t cell and decrease the amount transmitted to other cells deceasing its effect. And since the T cell is now not able to signal to the other T cell it thus inhibited 2.Cytokine inhibition: TREGs secrete several cytokines, including IL-10 and TGF-β, which bind receptors on activated T cells and reduce signaling activity. - Tregs also secrete IL-10 (which can bind and basically shut down pathways) and TGF-β which are both inhibitory cytokines which shut down signals. Ex stops the T cell from secreting more TNF α for instance. 3.Inhibition of antigen-presenting cells: TREGs can interact directly with MHC class II-expressing antigen-presenting cells and inhibit their maturation, leaving them less able to activate T cells. - inhibit of antigen-presenting cells by binding to APCs by binding to an MHC peptide and sending a signal through CD3 to inhibit the APCs 4.Cytotoxicity: TREGs can also display cytotoxic function and kill cells by secreting perforin and granzyme. - they can also kill T cells by secreting s ome of their granules so basically Tregs are there to stop the immune response and without them we would be more susceptible to autoimmune disorders
Importantly, when cells arrive at the thymus, they aren't technically T cells: they can become NK cells, dendritic cells, B cells, and myeloid cells. So what precipitates their development towards T cells? In other words, what is the first signal these cells received when they arrive at the thymus?
A receptor known as Notch commits arriving progenitors to the T lineage. Activation of Notch triggers activation of GATA-3 transcription factor. SO when cells arrive at the thymus depending on if they have received a signal from NOTCH or not determines if they are a T or B cell SO THE FIRST SIGNAL THESE CELLS RECEIVE WHEN THEY ARRIVE AT THE THYMUS is NOTCH
what is the kinetic model?
Because the two previous models may be too simplistic, another model, the kinetic model, was introduced. It has received the strongest experimental backing to date. The key feature is the following: thymocytes whose TCR preferentially interacts with (is restricted to) MHC class II generate a continuous signal that initiates a CD4+ helper T-cell developmental program. We now know that all CD4+CD8+ thymocytes down-regulate surface levels of CD8 in response to positive selection. Given this response, only MHC class II-restricted T cells will maintain continuous TCR/CD4/MHC class II interaction, and therefore develop to the CD4 lineage. However, with the loss of CD8 expression, MHC class I-restricted T cells will lose the ability to maintain TCR/CD8/MHC class I interactions. Singer and colleagues provide evidence that these thymocytes, interrupted from their TCR/coreceptor engagement, are subsequently rescued by IL-7, a key cytokine that promotes differentiation into the CD8+ lineage. •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 the disruption of signaling triggers a release of IL-7 which leads to the expression of CD8 TCR But the cell can only recognize the IL-7 when there is a signal disruption and only CH4-CD8+ cells have receptors for IL-7 IL-7 triggers the down-regulation of CD4 and the up-regulation of CD8 allowing the T cell to become a cytotoxic t cell
What are CD4+CD8+ cells? what is positive selection? Negative selection?
CD4+CD8+ (DP) thymocytes are small, nonproliferating, and dominate the thymic cortex, comprising more than 80% of cells in the thymus as a whole. Significantly, they are the first subpopulation of thymocytes that express a fully mature surface TCR-αβ/CD3 complex and are therefore the primary targets of 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. This eliminates 2-5% of cells. •Most cells (95%) fail positive selection and fail to receive needed survival signals: they will die by apoptosis. This process is known as death by neglect.
what occurs during the four DN stages of development? What molecule plays an important role in the DN stages?
CD4−CD8− (DN) thymocytes progress through four stages of development (DN1-DN4) defined by CD44, CD25, and c-Kit expression. During these stages they proliferate and rearrange the T-cell receptor (TCR) β, δ, and γ genes. c-Kit: receptor tyrosine kinase, CD25: the alpha chain of IL-2 receptor through C-Kit these cells are receiving survival signals that keep them in a stem state (a state where they remain DN). This C-Kit expression decreases from DN1 --> DN4 meaning now these cells are enabled to interpret signals from the c-kit ligand
After β-selection has occurred, thymocytes are at the?
DP stage of development: 1.Functional TCRα chain replaces surrogate pre-TCRα 2.The cell still expresses both CD4 and CD8 3.Negative then positive selections occur, yielding mature single positive T cell, CD4+ or CD8+ SO pretty much up until this point the DP stage we have just been generating T cells that have a functional α and β chain. we have not yet told them what they are supposed to recognize (which is our MHC population)
Does a DP thymocyte always become at SP thymocyte? if not, what else can a DP thymocyte become?
DP thymocytes may commit to other types of lymphocytes: •NKT cells. They express a TCR with an invariant TCRα chain. They interact with CD1 molecules presenting lipid antigens. These are not the same cells as NK cells. - CD1 is the molecule that looks like a MHC but presents glycolipids instead of peptides. SO NKT cells bind to CD1 •Intraepithelial lymphocytes (IELs). They are usually CD8+, but also have features of innate immune cells and patrol barrier tissues - so these cells are able to go into different tissues. They are tissue specific T cells. •Regulatory T cells (TREG cells). They are a CD4+ subset that helps to quench adaptive immunity. More on these later... - These are the most important. These cells arise from those DP thymocytes that were binding with farily high to the cTECs but instead of being deleted were selected because there binding was high but not too high All three of these cell types can develop from DP thymocytes in response to autoreactive, high-affinity TCR interactions: the same interactions, in fact, that mediate negative selection. What determines whether a thymocyte undergoes negative selection or an alternative developmental pathway remains a topic of much interest.
What happens at the DN2 stage of development?
During the DN2 stage of development, the genes for the TCR γ, δ, and β chains begin to rearrange (at the same time). The TCR α-chain locus, however, remains inaccessible to the recombinase machinery and does not yet rearrange. At the late DN2 stage, T-cell precursors fully commit to the T-cell lineage and reduce expression of both c-Kit and CD44. occurs in the DN stages, yielding either an αβ or a γδ T cell. To a large extent, the choice to become a γδ or αβ T cell is dictated by how quickly the genes that encode each of the four receptor chains successfully rearrange. Most thymocytes become TCR-αβ T cells (rearrangement of a β chain is more likely). However, TCR-γδ cells are the first T cells to mature during fetal development and populate the periphery in several early waves.
How do T cells exit the thymus?
It takes less than 3 days for a newly formed DP thymocyte to mature to the SP stage, if positively selected. SP cells spend a longer period (from 4 to 12 days) in the medulla, browsing the surface of epithelial and dendritic cells for self antigen before being given permission to leave the thymus. The exit of SP thymocytes from the thymus depends on expression of the sphingosine 1-phosphate receptor 1 (S1P receptor 1). We now know that mature thymocytes up-regulate S1P receptor 1 as part of the developmental program initiated by positive selection. The S1P receptor 1 interacts with its ligand, S1P, at the corticomedullary junction, facilitating thymocyte egress into the perivascular space and bloodstream. Mature thymocytes that have just left the thymus are referred to as recent thymic emigrants. They are not yet optimally functional and undergo a post-thymic phase of maturation in secondary lymphoid tissue that fully licenses them as naïve T cells. Common lymphoid progenitors arise in the bone marrow then they travel as immature cells to the thymus where they receive a pulse of NOTCH which allows them to now commit to the T cell route. They become DN and progress to DP and are selected and are ready. But how do they exit the thus once they are ready so they can begin sampling antigens? When the cells are mature enough and they are ready to exit the thymus they start expressing the S1P receptor which allows the cell to start becoming sensitive to levels of S1P. S1P looks like a chemokine but it is NOT it is a glycolipid and it is expressed in high amounts in the blood steam. So once the cell starts becoming sensitive to levels of S1P it will sense that levels of S1P are higher near the corticomedullary junction and therefore will start to migrate towards the corticomedullary junction where they can enter the blood stream where the signal is the highest. what is neat is that the S1P receptor is also involved in B cell development so they exit the thymus into the blood steam and travel to a lymph node where they can become mature and do their job.
Negative selection (central tolerance) ensures? what is clonal deletion?
Negative selection (central tolerance) ensures self-tolerance. Most negative selection occurs via a process known as clonal deletion, where high-affinity TCR interactions directly induce apoptosis. How do we delete thymocytes reactive to tissue-specific antigens? After all, the thymus does not express proteins from all tissues (e.g. myelin-basic protein). The transcription factor autoimmune regulator (AIRE) protein induces expression of many tissue-specific proteins in medullary thymic epithelial cells (mTECs). AIRE binds epigenetic marks on histones to recruit transcription factors. New T cells can be screened against these antigens safely in the thymus SO T cells after selection progress towards the single positive stage. They then migrate towards the medulla. Once in the medulla, they become associated with mTECS which express a transcription factor called AIRE. This TF induces the expression of proteins that are not usually found in the thymus. This helps the T cells learn about antigens not normally in the thymus
what does positive and negative selection help ensure? what are Cortical thymic epithelial cells (cTECS)? what are the three possible outcomes that can occur when our T cells encounter cTECs?
Positive and negative selection ensure that these mature T cells express TCRs with low affinity for self-peptide/self-MHC complexes, as long as they were presented in the thymus. On the other hand, there is no guarantee that these mature T cells express receptors that will be useful during any given infection: our immune system is "gambling" that one of the TCR will react to a MCH/peptide combination. Cortical thymic epithelial cells (cTECS) 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 selection, mediated by cortical thymic epithelial cells (cTECs), ensures?
Positive selection, mediated by cortical thymic epithelial cells (cTECs), ensures MHC restriction (the ability to recognize self MHC molecules). Low/intermediate-affinity signals between DP thymocytes and MHC/self-peptides on cTECs triggers a maturation program that sends cells to the medulla and initiates their commitment to CD4+ or CD8+ single-positive lineages. SO those that bind high but not to high are also selected to become Tregs which are T cells that are involved in the regulation of the immune system
What is lineage commitment? what does it require? are there different ways to explain lineage commitment? if so what are they?
Positively selected DP thymocytes must decide whether to become helper CD4+ T cells or cytotoxic CD8+ T cells. This process, called is called lineage commitment. It requires changes in genomic organization and gene expression that result in: 1.silencing of one coreceptor gene (CD4 or CD8) 2.expression of genes associated with a specific lineage function There are several theories to explain lineage commitment: 1.the instructive model 2.the stochastic model 3.the kinetic signaling model
where does T cell development occur? What type of organ is this?
T-cell development occurs in the thymus, the primary lymphoid organ in our chest cavity. Thymocytes are produced throughout our lifetime, although after puberty the thymus shrinks (involutes) and produces fewer and fewer T cells over time. Immature lymphocyte precursors migrating from the bone marrow, through the blood, and into the thymus. Here they proliferate, differentiate, and undergo selection processes that reduce their numbers by 95% (!), yet generate a diverse population of naïve CD4+ and CD8+ T cells. - immature lymphocyte precursors arrive at the corticomedullary junction. they then migrate through the cortex and then into the medulla and back into the corticomedullary junction to reenter the blood T-cell precursors begin their travel through the thymus at the cortex. T-cells that survive selection migrate into the medulla. They mature further in the thymic cortex, and then finalize their maturation in the thymic medulla; they exit as mature cells where they entered, via the corticomedullary junction. B cells go through all of there maturation in the bone marrow but T cells are a little different in that they mature in the thymus. You have progenitors from the bone marrow that enter the bloodstream and travel to the thymus. in the thymus is where we mature into a nieve t cell
what is the altered peptide model?
The altered peptide model is an alternative model can explain the thymic selection paradox. In this model, cortical thymic epithelial cells, which induce positive selection, make peptides that are unique and distinct from peptides made by thymic cells that induce negative selection. Thus, those thymocytes selected on these unique peptide/MHC complexes would not automatically be negatively selected when they browse the medulla and other negatively selecting cells. The model is still under investigation, but there is some evidence that thymus cells process antigens differently from other cells. The two theories aren't mutually exclusive...: multiple mechanisms of selection may exist
what is the instructive model?
The instructive model: •TCR/CD4 and TCR/CD8 coengagement generate unique signals The signals generated "instruct" the T cells which lineage to fully commit to so if a receptor binds to a peptide presented by an MHC class I we will down-regulate CD4 and up-regulate CD8 becoming CD8 AND if the receptor binds to a peptide presented by MHC class II we will down-regulate CD8 and up-regulate CD4 becoming CD4
So, by now, you may have started to appreciate the following paradox: if positive selection allows only thymocytes reactive with self-MHC molecules to survive, and negative selection eliminates self-MHC-reactive thymocytes, then how do any T cells escape selection and mature? What is the affinity model of selection? what happens if you don't have TAP?
The most straightforward model advanced to explain the paradox of MHC-dependent positive and negative selection is the affinity model of selection. This model asserts that differences in the strength of TCR-mediated signals received by thymocytes undergoing positive and negative selection determine the outcome of the interaction. Experiments conducted in the OT-I TR transgenic mouse system support the model: •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 SHOWs the affinity of a molecule is linked to the survival of the cell and the death of cells can occur via death by neglect if no interaction or chromal deletion if the T cell reacts to well
What is a stochastic model?
The 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 it says we randomly down-regulate CD4 or CD8 creating a stronger interaction in the end
What happens to double-negative (DN) thymocytes that have successfully rearranged their TCR β chains? What is β selection?
They are valuable and are identified and expanded via a process known as β-selection. This results in the proliferation/differentiation of cells after the formation of a pre-TCr signaling unit. SO we select the T cells in the DN3 and DN4 stage that is expressing a fully functional β chain. THEN we have four events 1. TCR α chain locus rearrangement 2. stimulates expression of CD4 and CD8 coreceptors (so the cell we become eventually DP) 3. simulates proliferation 4. stops additional TCR β chain locus rearrangements (allelic exclusion) (so we can have only this one β chain) also at the DN3 stage, we get a pre-TCR
Thymocytes progress through __________ double-negative stages of development.
Thymocytes progress through four double-negative stages of development. The various stages of development take place in distinct microenvironments within the thymus; these microenvironments provide both membrane-bound and soluble signals that regulate maturation. During the time it takes cells to develop to maturity in the thymus (1 to 3 weeks), thymocytes pass through a series of stages: 1.Double negative (DN) cell has no CD4 or CD8 (CD4-CD8-) 2.Double positive cell (DP) is both CD4+CD8+ 3.Positive/negative selection stages for a cell to become single positive CD4+ or CD8+
How do we generate such a group of T cells that are both tolerant to self and restricted to self-MHC?
We now appreciate the fundamental role the thymus plays in the development of immature T cells or thymocytes. So, in this lecture, we will investigate how to generate a large population of T cells that express a diverse array of receptor specificities that interact with self-MHC/peptide complexes but do not react against them. 1.Early thymocyte development 2.Positive and negative selection 3.Lineage commitment 4.Exit from the thymus and final maturation Other mechanisms that maintain self-tolerance
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
