T cell Differentiation

Thymic Selection

thymic selection ensures that thymocytes with only very specific characteristics are selected to develop further. The thymocytes that survive thymic selection - a very small percentage of the initial double positive population - differentiate further and leave the thymus to form the peripheral populations of mature CD4+ and CD8+ T cells. 1. Positive Selection - In the first stage, positive selection, the TCR of the double positive cell interacts with MHC class I and II molecules (and peptides) expressed by epithelial cells in the thymic cortex. This interaction results in survival and differentiation of the thymocyte into a mature T cell; the majority of double positive cells do not make this critical interaction and thus are not selected - they die by apoptosis. A further consequence of the interaction between the double positive cell and the thymic epithelial cell is that the thymocyte undergoes "lineage choice" - committing to developing into either the CD4+ or the CD8+ T cell lineage. The precise mechanism by which this occurs is still not completely understood, but it results in the downregulation of either CD4 or CD8. Thus, at this stage, the thymocyte becomes a single positive cell, expressing either CD4 or CD8. Another critical feature of this selection process is that the developing αbT cell that survives positive selection becomes "educated" to the MHC molecules expressed by the thymic cortical epithelial cells. This means that for the rest of the life of the T cell, even as a mature cell when it leaves the thymus, it will respond to antigen only when the antigen is bound to the MHC molecules that the developing T cell encountered in the thymus, either MHC class I or MHC class II. A CD8+ T cell will respond only to antigen plus one MHC class I molecule expressed in the thymus, and a CD4+ T cell will respond only to antigen plus one MHC class II molecule expressed in the thymus. For this reason, the MHC molecules expressed in a person's thymus and that educate his or her developing T cells are referred to as self-MHC; for that person, all other types of MHC molecules are "non-self". This is the origin of the phenomenon we referred to as the MHC restriction of T cell responses; more specifically, it is self-MHC restriction of T cell responses. 2. Negative Selection - As we described above, a thymocyte needs to have some affinity for self-MHC to be positively selected. However, allowing T cells with too high a reactivity to self-MHC to leave the thymus could result in undesirable autoimmune responses in tissues. To prevent this from occurring, thymocytes undergo negative selection, in which developing T cells with too high a reactivity to self-MHC are removed. In negative selection the key interactions are between the TCR and coreceptor, either CD4 or CD8, expressed on the thymocyte and MHC class I and II molecules (and associated peptides) expressed on dendritic cells and medullary epithelial cells. A thymocyte expressing a TCR that reacts with too high an affinity to the combination of MHC and peptide is deleted by apoptosis. Thus, negative selection removes T cells expressing T cell receptors with high (or strong) reactivity to self components and is important in establishing selftolerance. To summarize the selection process, we can say that double positive cells with affinity that is either too low or too high for self-MHC do not survive thymic selection. Only double positive cells with some intermediate affinity for self-MHC survive thymic selection. **In some rare cases, negative selection in the thymus does not take place and an autoimmune condition - autoimmune polyendocrinopathy-candidiasisectodermal dystrophy syndrome (APECED) - develops in which multiple endocrine organsparticularly the adrenals, parathyroids and thyroid, are damaged. We also know that expression of these self-antigens by thymic medullary epithelial cells is controlled at least in part by the gene autoimmune regulator (AIRE), which codes for a transcription factor - the rare individuals who lack an AIRE gene product are defective in negative selection

Double Positive T cells

After successful rearrangement of the α-chain and b-chain genes in the thymic cortex, developing T cells begin to express both CD4 and CD8 coreceptor molecules at their cell surface. These thymocytes are αβTCR+ CD3+ CD4+ CD8+ and are referred to as CD4+ CD8+ or double positive cells; they comprise the majority of thymocytes in the young mammalian thymus. Only time when T cells express both coreceptors. Double positive cells then undergo a multi-step process known as thymic selection

T Cell Differentiation

All blood cells, including T cells, are derived from pluripotent (or "multipotential") stem cells in the bone marrow. The thymus is the primary lymphoid organ for T cells, in which the developing T cell acquires a TCR. The development of mature T cells absolutely requires the thymus. Children with a congenital abnormality in which the thymus does not develop (DiGeorge Syndrome) do not produce mature T cells. T cell development occurs throughout the life of an individual but diminishes significantly after puberty. In fact, the size of the thymus itself decreases with the onset of puberty. Loss of the thymus after childhood does not pose a problem to the immune system because a long-lived mature T cell population has already been established. Immature T cells developing in the thymus are called thymocytes. Throughout T cell maturation, from precursor cell to mature T cell, thymocytes interact with a network of non-lymphoid, structural cells (stromal cells) of the thymus. Thymocytes trickle through this network, starting at the peripheral area (thymic cortex) and moving towards the central portion of the organ (thymic medulla). The thymic stromal cells provide critical cell surface interactions and produce cytokines that are essential for the development and maturation of T lymphocytes. The two main types of stromal cells encountered by developing thymocytes are epithelial cells and dendritic cells; both of these cell types express MHC Class II molecules (as well as MHC class I). The thymus is a site of intense proliferation of developing T cells. However, only a small percent of the T cells that are produced in the thymus actually leave — more than 95% of the developing T cells die in the thymus by apoptosis.

Functions of the Coreceptors, CD4 and CD8

CD4 and CD8 do not bind antigen but they enhance the ability of antigen to activate T cells through the TCR and so promote the interaction between a T cell and another host cell. CD4 and CD8 have two major functions: 1. They are specific adhesion molecules - CD4 binds specifically to MHC class II and so CD4 tightens the connection between a CD4+ T cell and a cell expressing MHC class II. MHC class II is expressed on antigen presenting cells (APC), in particular dendritic cells, macrophages, and activated B cells (thymic non-lymphoid cells). CD8 binds specifically to MHC class I so CD8 tightens the interaction between a CD8+ T cell and a cell expressing MHC class I. MHC class is expressed on all nucleated host cells. . To be more specific, CD4+ T cells are restricted by MHC class II molecules and CD8+ T cells are restricted by MHC class I molecules. 2. They act as signal transduction molecules - CD4 and CD8, like CD3 and ζ, relay signals to the inside of the T cell after the TCR has bound antigen. CD4 and CD8 bind to the invariant portion of MHC molecules, that is, the same in every person, and outside the peptide-binding groove of the MHC molecule. In humans CD4 and CD8 are expressed on more cell types than T cells and so their expression is not absolutely specific for T cells. CD4 is also expressed on macrophages and dendritic cells. **Clinical Correlate: The virus HIV binds to CD4. This allows the virus to enter and thus infect CD4+ T cells, and macrophages and dendritic cells, which also express CD4. This infection eventually results in acquired immunodeficiency syndrome (AIDS), discussed in a subsequent lecture.

TCR Structure

Like B cells, T cells express an antigen-specific receptor that is clonally distributed; that is, every clone of T cells expresses a TCR with a unique sequence. The TCR is a transmembrane molecule composed of two chains connected by a disulfide (S-S) bond. On the major subset of human T cells the TCR comprises one α chain and one β chain. Thus this subset of T cells is known as αβ T cells. A minor subset of T cells has a TCR that contains one γ chain and one δ chain - γδT cells - but less is known about the function of the γδ subset than about αβ T cells. The extracellular portions of the α and β chains have both variable (V) and constant (C) regions. The antigen binding site of the TCR is formed by a combination of the Vα region and the Vβ region (Similarly the antigen binding site of γδ T cells is formed by the Vγ region and the Vδ region). Have small cytoplasmic domain but outer domains stick out for antigen to bind to them. The TCR has only one antigen binding site, in contrast to the BCR (membrane Ig) which has two. The TCR is expressed in non-covalent association with CD3 and ζ ("zeta") polypeptides on the surface of T cells as a multimolecular complex - the TCR complex. CD3 and ζ do not bind antigen but function as signal transduction molecules, transmitting signals from the cell surface to the nucleus of the T cell after the TCR binds antigen. These signaling pathways lead to changes in the T cell's pattern of gene expression. CD3 is invariant, i.e. the same on all T cells (αβ and γδ), and because it is expressed only on T cells, its expression can be used to identify T cells and separate them from all other cell types. Zeta is not specific for T cells though. Molecule is constant on T cell once it is placed there, does not change during antigen stimulation. No class switch with TCR. Mechanisms that generate diversity for TCR is the same. **Many more alpha beta T cells than gamma delta T cells**

CD ("cluster of differentiation") nomenclature

Molecules expressed on the surface of lymphocytes and many other cell types are given a CD number, from CD1 upwards; currently over 300 different molecules have been assigned. This nomenclature standardizes the names of molecules on the cell surface among scientists all over the world and helps to emphasize the similarities between the "same" CD molecules in different species, e.g. CD4 is very similar in humans, chimpanzees, mice, etc.

Related Diseases

Myocardial Infarction (MI): Atherosclerosis that develops in coronary arteries results in decreased blood flow to the heart and can lead to an MI or other coronary heart diseases. Some studies suggest that atherosclerosis may in part be a response to a self-molecule (auto-antigen) Diabetes Mellitus: This disease develops as the result of a response to self-molecules expressed in the pancreas. HIV/AIDS: HIV binds to CD4, which in humans (but not every mammalian species) is expressed on macrophages and dendritic cells as well as CD4+ T cells. Thus, all these cells can be infected with HIV. In an HIV+ person a decrease in numbers of circulating CD4+ T cells below a threshold (200 cells per μL) characterizes AIDS Rheumatoid Arthritis: This inflammatory condition results from the response to selfmolecules, particularly in, but not limited to, the joints.

Differentiation of other cells in the Thymus

Other sets of cells also develop in the thymus: 1. γδ T cells - develop early in ontogeny, but are soon swamped by the development of αβ+ T cells. gδ T cells are generally found at much lower numbers than ab T cells in the circulation of normal adult humans, but they are found predominantly at mucosal epithelial sites such as the skin, gut, and lung, and are thought to form a first line of defense against pathogens. Generally, γδ T cells lack the CD4 and CD8 coreceptor molecules found on αβ‑expressing T cells, but γδ T cells found in the intestine express CD8. γδ T cells rapidly produce cytokines in response to pathogens such as mycobacteria, and have also been described to have cytotoxic function. They also respond to heat‑shock proteins (proteins that form in cells when they are heated or stressed in different ways). γδ T cells do not interact with the peptide-MHC complexes recognized by αβ T cells; they respond to phospholipids and other small non-protein molecules, known as phosphoantigens. 2. NK cells - are involved in the early phase of the immune response and are considered part of the innate immune defenses, kill virus-infected and tumor cells. Most NK cells develop in the bone marrow, but some develop in the thymus from the lymphoid precursors that give rise to the T-cell lineage. In the thymus, the NK cell development pathway splits off from the T-cell lineage at an early stage of double negative cells before TCR genes start to rearrange. Thus, NK cells do not express a TCR. 3. Treg cells - A subset of CD4+ T cells (approximately 10% of peripheral CD4+ T cells) that inhibit the actions of other sets of T cells. Treg cells that develop in the thymus are autoreactive; that is, they are T cells that recognize combinations of self-peptide and self-MHC but have survived negative selection in the thymus. Some Treg cells also develop outside the thymus. Treg cells play a role in inhibiting responses to both self and foreign antigens; in this way, they help to maintain and regulate self-tolerance and limit potentially damaging host responses to pathogens in tissues.

Pre-T Cells

Pre-T cells are cells in the thymic cortex that have productively rearranged a TCR β gene and express the TCRβ chain and a second molecule, pre-Tα, on their surface. Soon after successful β-chain rearrangement, α-chain genes also rearrange. The pre-Tα associated with the TCRβ chain on the surface of pre-T cells is non-rearranging and invariant. The combination of b chain and pre-Tα (together with CD3) constitutes the pre-T cell receptor (pre-TCR), analogous to pre-B cells and the pre-B-cell receptor. αβT-cells

Double-Negative Cells

Precursor cells (the common lymphoid progenitor) leave the bone marrow and enter the thymus to undergo maturation. TCR γ, δ, and β-chain genes then start to rearrange more or less simultaneously in the thymic cortex; these cells do not express either the CD4 or CD8 coreceptor (do express CD3 though) and so are referred to as double negative cells. Cells expressing γ and δ as their TCR split off from cells that will express α and β as their receptor early in intrathymic development. Cells expressing γ and δ as their TCR exit the thymus into the periphery, and move predominantly to mucosal epithelial sites such as the skin, gut, and lung. In adult humans gδ T cells are a small proportion of total T cells.

Leaving the Thymus

Single positive T cells exit from the medullary region of the thymus and circulate through secondary lymphoid organs (lymph nodes, spleen, and mucosal associated lymphoid tissue [MALT], including Peyer's patches, tonsils, and the appendix) where they form the pool of mature T cells in the periphery that may encounter and respond to antigen. In summary, thymic differentiation of T cells generates: 1. A huge repertoire of mature CD4+ T cells and CD8+ T cells that use ab as their TCR, which the individual uses to respond to the universe of non‑self (foreign) antigens 2. Mature CD4+ and CD8+ T cells that are tolerant to self

TCR Antigen Recognition

T cell responses are directed against antigens that get into cells; in particular, they are focused on pathogens that penetrate, infect, and live inside cells of the body. In order to deal with these infections, T cells must be able to distinguish between cells that have become infected and normal, non-infected cells. This is achieved by a T cell responding to a host cell that displays a linear section of a protein - a peptide - derived from the "foreign" antigen on its surface; specifically, the TCR interacts with a peptide fragment of a protein antigen bound to a major histocompatibility complex (MHC) molecule expressed at the surface of the host cell. The TCR makes contact with both the peptide and parts of the MHC molecule. This requirement for MHC expression in T cell responses is referred to as the MHC restriction of T cell responses. **Ig molecules can undergo somatic hypermutation and class switch recombination. These pathways do not occur in T cells - the TCR does not change over the lifetime of the T cell. TCR α (and γ) chains are constructed from V and J gene segments, like Ig light chains, whereas TCR β (and δ) chains are constructed from V, D, and J gene segments, like Ig heavy chains. **Antigen recognition by alpha beta T cells involves 3 structures. MHC molecule, peptide, and TCR

Coreceptors CD4 and CD8

The αβ TCR is also associated at the cell surface with the molecules CD4 and CD8, which are known as coreceptors. CD4 and CD8 are close to but not physically associated with the TCR. An individual mature T cell expresses either CD4 or CD8 but not both. Thus, mature T cells are either CD4+ CD8- or CD4- CD8+ , dividing T cells into two major functional subsets: CD4+ T cells and CD8+ T cells. More T cells (2 to 1) have CD4. AIDS patients have lower ratio. CD4 and CD8 have different function. the major function of CD4+ T cells is to synthesize cytokines that activate many cell types, and the major function of CD8+ T cells is to kill infected host cells

Humoral and Cell-Mediated Immunity

t B cells and their secreted products, antibodies, deal with antigens outside cells (within the plasma, lymph, or interstitial fluids). That arm of the immune response is known as humoral immunity. The B cell receptor for antigen expressed on the cell surface, and its secreted form (antibody), are immunoglobulin (Ig) molecules. Ig interacts with antigen, which can be any molecular type — protein, carbohydrate, lipid, or nucleic acid. On the other hand, T cell responses deal with pathogens inside cells of the host. Thus, T cells play a crucial role in the response to pathogens such as bacteria, virus, and parasites that infect and live in host cells. Because T cells interact with other cells, many T cell responses are frequently referred to as cell-mediated immunity. T cells are also involved in the response to many other "harmless", non-infectious antigens - for example, the proteins in a vaccine - that are taken up into host cells such as macrophages and dendritic cells. In contrast to B cell responses, most T cell responses focus on proteins, and not other chemical structures. Because T cells are restricted to interacting only with other cells, the pathways used by T cells to recognize antigens differ from the pathways used by B cells to recognize antigen.