Week 1: Fusion Session | T Lymphocyte Development PDF

1/17/24, 1:11 PM  Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024                       Fusion sessions are online learning activities followed by a live session where you will translate the content into practice. These require completing learning content in Canvas before attending live sessions. To encourage preparation, attendance, and participation, recordings of fusion sessions will not be posted; prepare accordingly so that you can fully participate. Remember: Learning from written materials is a critical professional and personal skill that RUSM is helping you develop through these sessions. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 1/50 Nerd Notes Ninja Thymus overlyes : · the heart in media stimen primary lymphoid organ allows T-cells to mature primarily · Immune : T-cell development · w/ it age during functions atrophys + filled w/ chemotactic younger ages of life fibrous tissue gif And ↓ -i J 7 Y W > - 00000 · 6 .. agents m ·...L [ ⑮ ⑧ - 8 T-cells ~ Thy mosin ⑳ produced ⑳ UW mus int Thymotaxin ⑭ Thymopoetnors - Tser E ou · spleen D 20 S . . ! ⑥ & S - ⑳ mus & ⑧R - · : lymph node 20 can also , go to tonsils mucosa eatic tissue , and many other places T-cells go deep part of the cortex of to · prevent formed due to · It TED4 M M & MHCs I T-cytotoxic CD25 cell cells of immune and T-helper + T- Cytotoxic cells diseases interleukin 2 on the cells that can cat O G & &s simniate T-cell -As activity anto &S CD8 1 nO T-regulatory the M the node 9 · to the ⑧ regulate M - O · O O . O 8 o sheaths pulp (white & D cells Positive I periarteriolar lymphatic # I ⑧6 lymph T-cells · & T-cel Warmundsnoodst - regulate the activity concentrateda (chemokines) to draw in T-cells /process called chemotaxis) heme O CD8 [D4 TCR + BCR none - ⑧ & "ono signaling. : no CD4 , CD8 d ↓ ⑨ 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 1. Work through the content on this page. 2. Contact faculty via email or office hours if you have questions about content to ensure you are prepared for the session. 3. Take the quiz. (You have three attempts.) 4. Attend and participate in the live session. 5. Take the quiz again. 6. Study missed content. 7. Take the quiz for the final attempt.  T lymphocytes are the cornerstone upon which all adaptive immune responses rest, including B lymphocyte activation and antibody production. In this section of the Immunology content, we will focus on the life cycle of T lymphocytes, with special emphasis on their early development, as well as the role of T cell early development in establishing the body’s capacity to discriminate between self and non-self. This section of content bears particular importance in establishing the bases of many immunodeficiencies, especially severe combined immunodeficiencies (SCID), and immunopathologies such as autoimmune disease and hypersensitivities. Once we have covered the essentials of innate immunity, cytokines, and inflammation, we will further develop the roles of T lymphocytes in immune responses (mainly in the Antigen Presentation & T Lymphocyte Biology lecture & the Immune Response to Infection: Prototypes class activity). By the end of this session, you will be able to meet the following learning objectives: 1. Compare and contrast TCRs and BCRs, particularly in terms of antigen recognition and MHC-restriction. Topcomplex- (MHC-) restriction and demonstrate the 2. Define major histocompatibility requirements for successful MHC-T cell receptor (TCR) engagement. 3. List the antigen-presenting cells bearing MHC-I molecules and the antigen-presenting cells bearing MHC-II molecules (i.e., professional antigen-presenting cells). https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 2/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 4. Compare and contrast the cells that respond to MHC-restricted antigen presentation in terms of: a. Identity (CD4 or CD8 lymphocyte). b. MHC type requirement (MHC-I or MHC-II). c. Antigen-presenting cells involved in their (1) development, (2) activation, costimulation, and differentiation, and (3) effector functions. d. Functions. 5. Differentiate between a:b TCRs and g:d TCRs (i.e., α:ß T lymphocytes & γ:δ T lymphocytes), as well as the development and functions of the cells bearing these different TCRs. 6. Describe the roles of the bone marrow, thymus, and secondary lymphoid tissue in the life cycle of α:ß T lymphocytes, and discuss the events occurring at the various stages of their development, including the surface markers that are acquired or expressed, their function(s), as well as the signaling events (cytokines & membrane-bound molecules) required at each stage. 7. Compare and contrast positive and negative selection: a. Describe the processes involved in positive and negative selection. b. Discuss the outcomes of positive and negative selection. 8. Define central tolerance and describe the process that leads to central tolerance. 9. Describe the basics of antigen presentation and T cell activation (these will be developed in detail in the Antigen Presentation and T Lymphocyte Biology lecture). 10. Discuss the consequences of incomplete T lymphocyte stimulation: a. Define peripheral tolerance. b. Describe the process leading to peripheral tolerance. 11. Compare and contrast central tolerance and peripheral tolerance and discuss their role and importance. 12. Discriminate effector T lymphocyte subsets (TH1, TH2, TH17, CTLs, TFH & Treg) and outline their respective functions (these will be developed in detail in the Antigen Presentation and T Lymphocyte Biology lecture). 13. List the cytokines responsible for the differentiation of each type of effector T lymphocyte subset (these will be developed in detail in the Antigen Presentation and T Lymphocyte Biology lecture). 14. List the characteristics of innate-like lymphocytes (ILLs) (e.g., NK cells, NKT cells, & IELs) and describe their main functions. Top of memory T lymphocytes. 15. Discuss the role and importance 16. Discuss the population dynamics of T lymphocytes throughout a person’s life. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 3/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Click the tab below to learn more about the relevant USMLE content. Topics covered in this session are highlighted. Development of cells of the adaptive immune response, including positive and negative selection during immune development Structure, production, and function granulocytes, natural killer cells, macrophages, mast cells, dendritic cells, cell receptors (e.g., complement receptors and Toll-like receptors), cytokines, chemokines T lymphocytes, including T-lymphocyte receptors, accessory molecules (e.g., CD3, CD4, CD8, B7), cell activation and proliferation, cytotoxic T lymphocytes, and memory T lymphocytes B lymphocytes and plasma cells, including B-lymphocyte receptors, immunoglobulins, cell activation and proliferation, including development of antibodies and memory B lymphocytes host defense mechanisms, host barriers to infection, mucosal immunity (e.g., gutassociated lymphoid tissue and bronchus-associated lymphoid tissue), anatomical locations of T and B lymphocytes Cellular basis of the immune response and immunologic mediators antigen processing and presentation in the context of MHC I and MHC II molecules (e.g., TAP, beta-2 microglobulin), intracellular pathways, mechanisms by which MHC is expressed on the surface, including distribution of MHC I and MHC II on different cells, mechanisms of MHC I and MHC II deficiencies, and the genetics of MHC regulation of the adaptive immune response (e.g., peripheral tolerance, anergy, regulatory T lymphocytes, termination of immune response, and B-T lymphocyte interactions) activation, function, and molecular biology of complement (e.g., anaphylatoxins) functional and molecular biology of cytokines (e.g., IL 1-15) Basis of immunologic diagnostics (e.g., antigen-antibody reactions used for diagnostic purposes, ELISA, immunoblotting, antigen-antibody changes over time, ABO typing) Principles of immunologic protection vaccine production and mechanisms of vaccine action biologically active antibodies (e.g., monoclonal antibodies, polyclonal antibodies Top including IVIG, VZIG, rabies immunoglobulin) Effect of age on the function of components of the immune system https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 4/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Abbas, A. K., Lichtman, A. H., & Pillai, S. (2022). Cellular and molecular immunology. Elsevier. Abbas, A. K., Lichtman, A. H., & Pillai, S. (2018). Cellular and molecular immunology. Mtm. Delves, P. J., Martin, S. J., Burton, D. R., & Roitt, I. M. (2011). Roitt’s essential immunology. Oxford Wiley-Blackwell. Murphy, K., Travers, P., & Walport, M. (2008). Janeway’s immunobiology. Buch. Garland. Paul, W. E. (2008). Fundamental immunology. Wolters Kluwer Health.  Thymus-dependent lymphocytes, or T lymphocytes, play a major role in the defense against pathogens such as viruses, intracellular protozoa, fungi, and bacteria, as well as extracellular pathogens (bacteria, protozoan, fungi, & helminths, i.e., worms), by (1) providing help for the regulation of pathogen-tailored adaptive immune responses and the antibody response or (2) directly killing infected host cells. Additionally, it has been shown that T cells play an important role in immune surveillance, as well as the recognition and killing of tumor cells. Although this section of the course deals with T lymphocyte development, we do need to introduce a few concepts related to antigenic receptors and antigen (Ag) presentation, as these are central to the development of T cells.  T cells are like B cells in that they Top express a surface receptor that recognizes a specific antigen. T lymphocyte and B lymphocyte Ag recognition are mediated by Ag receptor complexes composed of two main components: an Ag binding part and a signaling part. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 5/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Feature or function Forms of antigens recognized Diversity Antibody (immunoglobulin) T cell receptor (TCR) Macromolecules (proteins, polysaccharides, lipids, nucleic acids), small chemicals Top Mainly peptides displayed by MHC molecules on APCs Each clone has a unique specificity; potential* for ~1011 distinct specificities Each clone has a unique specificity; potential for ~1016 distinct specificities Linear epitopes https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 6/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Antigen recognition is mediated by: Variable (V) regions of heavy and light chains of membrane Ig Variable (V) regions of α and ß chains of the TCR Signaling functions are mediated by: Proteins (Igα and Igß) associated with membrane Ig Proteins (CD3 and ζ) associated with the TCR Effector functions are mediated by: Constant (C) regions of secreted Ig TCR does not perform effector functions The B cell receptor (BCR) complex is comprised of: 1. a membrane-bound immunoglobulin (antibody) molecule responsible for binding Ag, and 2. a host of other molecules (Igα & Igß, CD19, CD21, & CD81) involved in the transduction of the signal through the cell’s plasma membrane to yield a physiological response. Therefore: the immunoglobulin portion is responsible for Ag specificity, whereas the other molecules of the BCR complex trigger a signaling cascade culminating in a physiological response, i.e., antibody production in the case of B cells (an antibody is the secreted, soluble counterpart of the BCR immunoglobulin moiety, meaning that the secreted antibodies possess the same Ag specificity as the BCR of the cell secreting them). Finally, and this is important in terms of differentiating BCRs and TCRs, the BCR recognizes free Ag (conformational, i.e., 3D structure, & linear epitopes of proteins, polysaccharides, lipids, nucleic acids, even some metals, etc.), as opposed to the TCR which recognizes Ag linked to major histocompatibility complex (MHC) molecules displayed by Ag-presenting cells (below). Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 7/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Major histocompatibility complex-restricted antigen presentation requires adequate interactions of the TCR with both antigenic peptide and MHC molecule. Like the BCR, the T cell receptor (TCR) complex is composed of two parts: 1. the TCR chains per se, which are responsible for Ag recognition and, therefore, antigen specificity and 2. a set of other molecules (CD3 as well as CD4 or CD8, depending on the T lymphocyte subset) responsible for the signal transduction leading to a set of physiological responses. Top However, unlike the BCR, which recognizes free Ag, the TCR recognizes (1) a linear peptide bound to (presented by) MHC molecules exhibited by Ag-presenting cells (APCs); the requirement for the TCR to interact with both the antigenic peptide and the APC’s MHC (i.e., not free Ag but Ag bound https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 8/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 to an MHC molecule) is known as MHC-restriction. The successful engagement of the TCR requires compatibility with both peptide Ag and MHC. Contrary to BCRs, which bind free antigens (either in linear conformation or three-dimensional conformation), TCRs bind linear peptide antigens in association with MHC molecules. Note: The outcomes of TCR engagement will be further developed in the Antigen Presentation & T Lymphocyte Biology section of this course. Complete the activity below. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 9/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Drag the receptor to the statement to which it best ﬁts. Speciﬁcity conferred by membrane-bound α- & ß- chains BCR Binds Ag associated with an MHC molecule TCR Binds free Ag TCR Binds both linear & conformational Ag BCR Binds linear Ag BCR Signaling conferred by CD3 complex BCR Recognition mainly limited to peptide Ag TCR Speciﬁcity conferred by membrane-bound antibody TCR Signaling conferred by Igα/Igβ BCR Associated with CD4 or CD8 Recognition of protein, carbohydrate, lipid, nucleic acid Ag TCR Associated with CD19 & CD21 BCR TCR  Check TCR BCR Binds Ag associated with an MHC molecule Binds free Ag Binds linear Ag Binds both linear & conformational Ag Specificity conferred by membranebound α- & β-chains Specificity conferred by membranebound antibody Signaling conferred by CD3 complex Signaling conferred by Igα/Igβ Recognition mainly limited to peptide Ag Recognition of protein, carbohydrate, lipid, nucleic acid Ag Associated with CD4 or CD8 Top Associated with CD19 & CD21 https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 10/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024  Note: Ag presentation is further addressed in the Antigen Presentation & T Lymphocyte Biology section of this course. This section only introduces the necessary prerequisites needed to cover the development of T lymphocytes. As aforementioned, T lymphocytes respond to antigenic stimulation in the context of MHC molecules displayed by APCs. There are essentially two main subsets of T lymphocytes: CD4 T lymphocytes and CD8 T lymphocytes, and each subset of T cells reacts to one of two MHC types, MHC-I or MHC-II. Likewise, there are two main categories of APCs depending on MHC expression: all nucleated cells (i.e. basically all cells of the body except for erythrocytes) express MHC-I, whereas MHC-II expression is mostly restricted to so-called professional APCs (pAPCs), i.e., dendritic cells (DCs), macrophages, thymic epithelial cells (TECs), and B lymphocytes (because MHC-II-expressing cells are nucleated, they also express MHC-I; consequently, pAPCs express both MHC-I and MHC-II – I mention this because medical students often think that pAPCs only express MHC-II & not MHC.). Major histocompatibility complex I molecules engage the TCR on the surface of CD8 T lymphocytes which are primarily involved in the killing infected cells and tumor cells. CD4 cells, whose main function is to regulate adaptive immune responses, respond MHC-II-restricted Ag presentation. Complete the activity below. Drag MHC-I and MHC-II to the statements that best correspond. Expressed by all nucleated cells MHC-I Involved in the regulation of adaptive immune responses MHC-II Mainly restricted to pAPCs MHC-II Used by APCs to engage CD8 T lymphocytes Involved in the killing of infected/tumor cells Used by pAPCs to engage CD4 T lymphocytes Top MHC-I MHC-II MHC-I  Check https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 11/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 MHC - I MHC - II Expressed by all nucleated cells Mainly restricted to pAPCs Used by APCs to engage CD8 T lymphocytes Used by pAPCs to engage CD4 T lymphocytes Involved in the killing of infected/tumor cells Involved in the regulation of adaptive immune responses  Although TCR engagement is key to all adaptive immune responses, the generation of the TCR is only one part of a sequential process that occurs during T lymphocyte development and maturation. T cell development involves several distinct stages. This process starts in the bone marrow with the generation of T cell progenitors (common lymphoid progenitor cells), which then enter the thymus and undergo their development and education. Once a mature T cell exits the thymus, it traffics from secondary lymphoid tissues to secondary lymphoid tissues (e.g., from lymph node (LN) to LN using lymphatic vessels) in search of its specific antigen. Upon encounter with said specific antigen, it undergoes a program of activation, proliferation, and differentiation into an effector cell to address the infection or injury or a memory cell for future exposure to the same Ag. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 12/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Stem cells originating in the bone marrow enter the thymus and develop into mature naïve T lymphocytes prior to exiting the thymus and circulating through secondary lymphoid tissue; important developmental milestones are presented. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 13/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Where do lymphocyte progenitor cells arise?  Spleen  Lymph node  Liver  Bone marrow  Thymus  Check Where do lymphocyte progenitor cells arise? Bone marrow (Correct answer) Liver Lymph node Spleen Thymus  T cells develop into mature functional T cells after passage through the thymus. T cell progenitors first arise from pluripotent hematopoietic stem cells in the bone marrow (red marrow of flat bones – mostly in adults/epiphyseal ends of long bones – mostly in children). Some of these pluripotent hematopoietic stem cells further differentiate into common lymphoid progenitor cells that give rise to lymphocytes (B cells & T cells). Some common lymphoid progenitor cells then enter the bone marrow vasculature to enter the general circulation and eventually reach the thymus. Studies with athymic animals clearly demonstrate the importance of the thymus. The nude mouse, nu/nu, is Top congenitally athymic and is exquisitely incapable of handling many types of infections. It is unable to stimulate antibody production, cannot mount delayed-type reactions, and cannot even reject xenografts. Immune functions can be restored by the implantation of a thymus under the kidney capsule. In humans, patients with complete DiGeorge syndrome fail to make T lymphocytes as a https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 14/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 result of the absence of a thymus; consequently, these individuals possess, in effect, no adaptive immunity. Normal T cell development is dependent upon both direct contact with thymic stroma as well as interaction with secreted molecules. Thymic subcapsular and medullary epithelial cells produce a variety of polypeptides that influence the phenotypic maturation of progenitor cells from the bone marrow and the modulation of the functions of mature T cells. These thymic peptides include thymosin-α, thymosin-ß, thymulin, thymopoietin, and thymic humoral factor. T cell progenitors enter the thymus at the junction between the cortex and the medulla. At this point, they are referred to as thymocytes. Cell migration is in the direction of cortex to subcapsular zone to cortex to medulla, and cells acquire developmental markers as they progress through Top maturation. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 15/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Where do lymphocytes develop into mature naïve T lymphocytes?  Thymus  Liver  Lymph node  Spleen  Bone marrow  Check Where do lymphocytes develop into mature naïve T lymphocytes? Lymph node Thymus (Correct answer) Bone marrow Spleen Liver Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 16/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Localization of thymocyte development stages in the thymus Capsule Subcapsular Region Cortex Click here to begin Corticomedullary Junction Medulla Progenitor cells leaving the bone marrow possess the stem cell marker CD34 and lack the characteristic cell surface glycoproteins of mature T cells, and their receptor genes are in the germline configuration. T cell receptor gene rearrangements do not occur until the cells enter the thymus. T cell progenitors enter the thymus at the corticomedullary junction and migrate towards the cortex, where most of the maturation takes place. Upon interaction with cortical thymic stromal cells (cTECs or cortical thymic epithelial cells), thymocytes receive signals to proliferate Top and commit to the T cell lineage: 1. cTEC-secreted IL-7 engages thymocyte interleukin 7 receptors (CD127, also called IL-7R; dependent on functional JAK3-mediated signaling – individuals lacking either common IL2Rγ functional chain or the tyrosine kinase it is associated with (JAK3) or a functional IL-7R https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 17/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 α-chain are severely immunocompromised due to lack of further thymocyte development, i.e., they have little or no T cells, & this will be further developed in the (1) Cytokines, Chemokines & Signaling & (2) Immunodeficiencies II learning activities 2. direct signaling provided by Notch1 ligand on the surface of cTECs to stimulate Notch1 on the surface of thymocytes. Within one week, they are committed to the T cell lineage (pre-T cells) and start to express the CD2 adhesion molecule; at this stage, they lack expression of the CD4 and CD8 co-receptor molecules (referred to as CD4- & CD8-) and are therefore termed double-negative (DN) thymocytes. What cytokine is key in the early development of T lymphocytes?  IL - 2  TFN - α  IL - 4  IL - 7  IL - 1  Check What cytokine is key in the early development of T lymphocytes? IL-1 IL-2 IL-4 IL-7 (Correct answer) TNF-α Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 18/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 A critical step in T cell development is the acquisition of the TCR; this takes place at the edge of the thymic cortex (subcapsular region). As for the BCR, rearrangement of the TCR involves the recombination activating genes (RAG), RAG-1 and RAG-2, and terminal deoxyribonucleotidyl transferase (TdT). These genes are only expressed by thymocytes during the rearrangement of the T cell receptor genes (i.e., during T cell development in the thymus). The TCR is a heterodimer composed of two glycosylated polypeptide chains belonging to the immunoglobulin superfamily, and there are two types of TCRs: α:β TCRs and γ:δ TCRs. The α:β TCR is composed of one αchain and one β-chain, whereas the γ:δ TCR is made up of one γ-chain and one δ-chain. The α:β TCR is the type found on the majority of T lymphocytes, and it binds MHC/antigenic peptide complexes (MHC-I or MHC-II). The γ:δ TCR is found on less than 5% of T lymphocytes and recognizes antigens that are MHC-restricted or not (e.g. CD1:glycolipid antigens; refer to MHC & Antigen Presentation handout material and in a later section of this handout). The TCR β- and δchain loci contain V (variable), D (diversity), and J (joining) segments and, similarly to the BCR, recombination enzymes first join a D segment to a J segment to yield a DJ segment, which is then rearranged to join a V segment resulting in a VDJ segment. The TCR α- and γ-chain loci on the other hand only contain V and J segments so that rearrangement is limited to a single VJ segment; this rearrangement is performed by the same recombination enzymes (i.e., RAG-1, RAG-2, and TdT). Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 19/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Just like the pre-BCR and BCR, the pre-TCR and TCR are inherently incapable of signaling and must therefore be associated with other polypeptide chains able to assist in signal transduction; six such subunits, collectively referred to as CD3, cluster with the TCR to provide phosphorylation sites (ζ[zeta]-chains) on the cytoplasmic side of the plasma membrane for the recruitment the next kinase in the signaling cascade. Thymocytes start to rearrange their β-, δ-, and γ-chain genes at the same time in a competitive manner. By contrast, in B cells, immunoglobulin gene rearrangement is orderly and sequential. The appearance of one set of receptors signals the cessation of rearrangement of the competing set of Top receptors. If a functional γ:δ receptor is produced before the elaboration of a functional βchain, then a γ:δ TCR-expressing T cell develops (differences between α:ß and γ:δ T cell populations will be discussed below) and exits the thymus without undergoing any selection process. In thymocytes that express a functional b-chain first (the majority), the ß-chain https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 20/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 associates with an invariant pre-T α-chain (a surrogate α-chain expressed until the formation of a bone fide α-chain) and the CD3 molecule (see below for a description of CD3), forming a pre-TCR complex. Expression of the pre-TCR signals double-negative thymocytes to stop rearrangements of the b-chain gene and undergo proliferation. The cells are also signaled to initiate expression of CD4 and CD8, giving rise to double-positive (DP) thymocytes (CD4+CD8+). CD4 is a molecule composed of a single polypeptide chain of the immunoglobulin superfamily. CD8, on the other hand, is composed of a heterodimer made up of an a-chain and a b-chain; both polypeptide chains also belong to the immunoglobulin superfamily. Once they complete proliferation, rearrangement of the TCR a-chain ensues. While this is occurring, γ - and δ-chain rearrangements are ongoing and, again, if a functional γ:δ TCR is produced first, the cell commits to the γ:δ lineage and exits the thymus; if on the other hand, a functional α:ß TCR is produced first, the cell commits to the α:ß lineage and further thymic development will continue. The expression of the TCR signals the end of all gene rearrangements and stimulates the expression of other T cell markers (see below). These cells have a lifespan of about 3-4 days and die of apoptosis unless they are rescued by engagement of their TCR (positive selection – see below). Only 10-30% of TCRs generated (i.e., 10-30% of DP cells) will interact with self-peptide:self-MHC complexes. Cells that fail to express a TCR die by apoptosis and are phagocytosed by macrophages in the thymic cortex. Which of the following enzymes is involved in the rearrangement of the TCR?  Autoimmune regulator (AIRE)  Stem cell factor (SCF)  Recombination-activating genes 1 & 2 (RAG1/RAG2)  Activation-induced cytidine deaminase (AID)  Adenosine deaminase (ADA)  Check Top Which of the following enzymes is involved in the rearrangement of the TCR? Adenosine deaminase (ADA) Activation-induced cytidine deaminase (AID) https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 21/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Autoimmune regulator (AIRE) Recombination-activating genes 1 & 2 (RAG1/RAG2) (Correct answer) Stem cell factor (SCF) Top The TCR complex (TCR/CD3 complex) is invariably associated with a co-receptor (CD4 or CD8) that (1) binds the MCH, and (2) possesses, associated with its cytoplasmic tail, the tyrosine kinase (primarily a TK called lymphocyte-specific protein tyrosine kinase, or lck) responsible for the phosphorylation of CD3 ζ-chains and the next tyrosine kinase in the pathway (i.e., ZAP-70) https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 22/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 (zeta-chain-associated protein kinase; this will be further developed in the Antigen Presentation and T Lymphocyte Biology activity). Like B cell receptors, T cell receptors must undergo a selection process. T cell receptors must be tested to (1) ensure the TCR can interact with self-MHC (positive selection) and (2) that the TCR does not respond to self-antigen (negative selection), i.e., is autoreactive with the potential to cause autoimmune disease. There are two positive/negative selection pathways: one is sequential, and the other is nonconsecutive. Click each tab below to learn more about the different pathways. Sequential Pathway Nonconsecutive pathway In the sequential pathway, thymocytes whose TCR binds self-MHC:self-peptide with moderate affinity are given survival signals and are thereby positively selected. Those that fail to sufficiently interact with self-MHC die by neglect and are thereby negatively selected. Positive selection occurs deep in the thymic cortex, and the antigen-presenting cells involved in the process of positive selection are cTECs; these cells present tissue-restricted peptides in the context of both MHC class I and MHC class II. Thymocytes positively selected for MHCreactivity in this way must then be tested for self-reactivity, which mostly occurs in the medulla: TCRs that bind self-MHC:self-peptide too strongly are signaled to die, whereas the others survive, mature, and exit the thymus. Positive selection leads to the maturation of cells that can sufficiently react with the person’s MHC allotypes and also determines if a double-positive cell becomes a CD4+ cell or a CD8+ cell. Double-positive cells interact with self-peptide: selfMHC complexes. When a TCR preferentially interacts with a class I MHC molecule, the cell commits to the CD8+ cell lineage; likewise, when a TCR preferentially interacts with class II MHC, the cell commits to the CD4+ lineage. The CD4 and CD8 co-receptor molecules then become part of the TCR complex. Cells that remain CD4+CD8+ are generally negatively selected – in other words, if the TCR fails to interact with either MHC class I or MHC class II, the cell dies. Thymocytes that have been positively selected move on to the thymic medulla, where they are presented self-peptides by medullary thymic epithelial cells (mTECs), bone marrow-derived dendritic cells (bmDCs), and bone marrow-derived macrophages; these cells express tissue-specific antigens (syn. tissue-restricted antigens), as a result of so-called promiscuous gene expression, that would otherwise never be expressed in the thymus (e.g., insulin and hundreds of tissue-specific proteins) and present these tissue-specific peptides to thymocytes in an effort to eliminate thymocytes reactive to these tissue-specific proteins. This Top is possible due to a singular transcription factor these cells express, an autoimmune regulator, or AIRE, which activates these tissue-specific genes. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 23/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Watch this video: AIRE: From promiscuous molecular partnerships to promiscuous gene expression (https://www.youtube.com/watch? v=lUYeMQmGmO4) One key factor in the process of negative selection is the ability of these cells to provide costimulation (CD80/CD86 – see below in T Lymphocyte Signaling); thymocytes specific for selfMHC: self-peptide complexes that receive co-stimulation in the thymic medulla are signaled to die, thereby eliminating those positively selected thymocytes that are self-specific and could potentially lead to autoimmune disease. Note that, as we will see later, once in the periphery, antigen presentation with co-stimulation yields activation, clonal expansion, and differentiation, not death. After the negative selection process, thymocytes that have escaped negative selection achieve maturation and leave the thymus as mature, naïve (i.e., have never been presented with foreign Ag), resting α:ß T lymphocytes. Overall, T lymphocyte development takes place in about 3 weeks, and at the end of the process, only about 2% of DP cells survive the selection process and are exported to the periphery. Negative selection is the main driving force behind the development of central tolerance (i.e., the process by which one’s immune system tolerates one’s antigens by eliminating the most self-reactive T lymphocytes in the thymus). Tolerance, as well as the breakdown of tolerance, is an important concept when considering autoimmune disease, hypersensitivities, and tissue transplant. In the nonconsecutive pathway, thymocytes with high TCR affinity are signaled to die and deleted (central tolerance), whereas those with moderate TCR affinity go on to become mature, naïve T lymphocytes; thymocytes with intermediate TCR affinity develop into thymic regulatory T lymphocytes (tTreg) that act as suppressive cells in the periphery to repress immune reactivity to self-antigen previously seen in the thymus and thus contribute to peripheral tolerance. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 24/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What process leads to the elimination of self-reacting (self-speciﬁc) lymphocytes?  Single-positive lineage commitment  Positive selection  Peripheral tolerance  Clonal expansion  Negative selection Top  Check https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development  25/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Question 1 What process leads to the elimination of self-reacting (self-specific) lymphocytes? Positive selection Peripheral tolerance Clonal expansion Negative selection (Correct answer) Single-positive lineage commitment Question 2 Which of the following describes the elimination of self-reacting T lymphocytes in the thymus? Positive selection Peripheral tolerance Central tolerance (Correct answer) Immune senescence Clonal expansion Mature T cells exit the thymus and enter the pool of recirculating lymphocytes. Mature, naïve T lymphocytes that encounter large, constant amounts of antigen shortly after exiting the thymus and in specific locations (e.g., self-antigen for those few T cells that have eluded negative selection, food, normal microbiota, dust, pollen etc.) are either eliminated by activation (activation-induced cell death), become anergic due to lack of co-stimulation (pAPCs presenting these antigens usually do so in absence of co-stimulation – many of these cells can survive in an anergic), or are suppressed by anti-inflammatory cytokine stimulation expressed by Treg lymphocytes. Eliminating or rendering T lymphocytes unresponsive to ‘ubiquitous’ antigens once they have left the thymus and reached the periphery is called peripheral tolerance. Mature, naïve, resting T lymphocytes are longer-lived than B cells and, in the absence of antigen stimulation, may live for years. In the presence of antigens, they may become activated and release cytokines that mediate effector functions (these cells have a dramatically shortened lifespan – days to weeks) or become memory cells responsible for long-term immunity (with a lifespan of months to years – see below). Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 26/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Which of the following describes the regulation of self-reacting T lymphocytes in the periphery?  Central tolerance  Negative selection  Immune senescence  Peripheral tolerance  Positive selection  Check Which of the following describes the regulation of self-reacting T lymphocytes in the periphery? Negative selection Positive selection Peripheral tolerance (Correct answer) Central tolerance Immune senescence  To perform their function(s), T lymphocytes require three separate signals: (1) activation per se (i.e., TCR complex engagement), (2) co-stimulation (i.e., CD28 engagement to reinforce signal 1), and (3) differentiation (i.e., cytokine receptor(s) engagement); signal 2 is required to reinforce signal 1 and make the activation complete (survival and clonal expansion of the lymphocytes), whereas signal 3 is the signal that actually determines the subset of T lymphocyte the cell will differentiate into (e.g., T lymphocyte required for an immune response to intracellular microbes, as opposed to an immune response against extracellular microbes, each of which requiring different Top strategies). https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 27/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 To perform their function(s), T lymphocytes require three separate signals: (1) activation (i.e., TCR complex engagement), (2) co-stimulation (i.e., CD28 engagement to reinforce signal 1), and (3) differentiation (i.e., cytokine receptor(s) engagement). Recognition of a specific peptide: MHC complex by a T lymphocyte triggers signal transduction events mediated by the TCR complex. As previously mentioned, the TCR complex is composed of two parts: (1) the TCR chains, which are responsible for Ag recognition and, therefore, antigen specificity, and (2) a set of transmembrane molecules (CD3 as well as CD4 or CD8 depending on the T lymphocyte subset) responsible for the signal transduction leading to physiological responses. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 28/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 The TCR α- and β-chain extracellular domains provide the specificity of the TCR complex, whereas CD3 provides the signaling capacity of the TCR complex. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 29/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What part of the TCR complex allows it to transduce antigen recognition into an intracellular messaging response?  CD28  CD3  CD80/CD86  CD1  TCR α- and β-chains  Check What part of the TCR complex allows it to transduce antigen recognition into an intracellular messaging response? CD1 CD3 (Correct answer) CD28 CD80/CD86 TCR α- and β-chains However, T lymphocyte activation by MHC-restricted antigen presentation alone is not sufficient to achieve a physiological response; survival and differentiation signals are also required. The survival signal is delivered by pAPCs in the form of the surface co-stimulatory molecules CD80 and/or CD86, also called B7.1 and B7.2, respectively (or B7 in the old nomenclature); the T cell surface receptor for co-stimulatory molecules is CD28. However, co-stimulation has different consequences depending on the location where Ag presentation occurs. During negative selection in the thymus, Ag presentation and co-stimulation leads to T cell death and, therefore, is the process driving central tolerance. In peripheral (secondary) lymphoid tissue, activation of mature T lymphocytes also requires differentiation signals in the form of cytokines, and these cytokines effectively dictate the type of effector T lymphocyte subset (TH1, TH2, TH17, TFH, Treg) that results from antigen presentation. So, in summary, Ag presentation and CD80/CD86 co-stimulation in the Top is responsible for central tolerance, whereas Ag presentation thymus results in T cell apoptosis and in peripheral lymphoid tissue requires survival and proliferation signals (CD80/CD86) as well as cytokine signaling to differentiate T cells according to the immune functions required. Finally, incomplete T lymphocyte activation in secondary lymphoid tissue, as previously mentioned, leads https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 30/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 to T cell anergy and peripheral tolerance. Note: T lymphocyte signaling will be further developed, in detail, in the Antigen Presentation & T Lymphocyte Biology lecture. Top T cells failing to receive co-stimulation during antigen presentation either become (1) anergic (non-responsive to antigenic stimulation) or (2) become tolerant to said antigen. https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 31/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 What is the T lymphocyte receptor for costimulatory molecules?  CD80/CD86  CD28  CD1  CD3  TCR α- and β-chains  Check What is the T lymphocyte receptor for costimulatory molecules? CD1 CD3 CD28 (Correct answer) CD80/CD86 TCR α- and β-chains As aforementioned, the differentiation of T lymphocytes into effector cells in peripheral lymphoid tissue is driven by cytokines. Generally, antigen presentation achieved in the presence of IL-12 and IFN-γ yields TH1 cells; TH1 cells are involved in adaptive immune responses to intracellular pathogens (viruses and intracellular bacteria, fungi, & protozoans). Antigen presentation achieved in the presence of IL-4 (with or without combinations of IL-5, IL-10, & IL-13) generates TH2 cells; TH2 cells are involved in adaptive immune responses to helminths (worms). Antigen presentation achieved in the presence of IL-1ß, IL-6, IL-23, and TGF-ß yields TH17 cells; TH17 cells are involved in adaptive immune responses to microscopic extracellular pathogens (some bacteria, fungi, & protozoans) but also in inflammatory and autoimmune diseases; TH17 cells activated in the presence of IL-6 and TGF-β yield so-called inflammatory TH17 cells, also called regTH17 cells Top extracellular microbes), whereas TH17 cells activated in the (these are the cells useful to eliminate presence of IL-1β and IL-23 yield so-called pathological TH17 cells (those involved in inflammatory and autoimmune diseases). Antigen presentation achieved in the presence of TGF-ß generates https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 32/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Treg cells. CD8 T lymphocytes generally differentiate into CTLs (cells that are specialized in the killing of infected host cells and transformed cells. Ag presentation is a part of several stages of the T cell’s life cycle, and at each stage, Ag presentation serves a different function. Click through this interactive to learn more about Ag presentation. Ag Presentation in the Life Cycle of a T Cell START Top Process Site https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 33/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 Antigen presentation for the purpose of T lymphocyte development (Positive and negative selection) Antigen presentation to double-positive thymocytes by thymic cortical epithelial cells (positive selection) and bone marrow-derived macrophages and dendritic cells (negative selection - i.e., central tolerance) Thymus - Development (Primary lymphoid organ) Antigen presentation for the purpose of T lymphocyte development (Response to infection) Antigen presentation to naïve T lymphocytes by professional antigenpresenting cells (mostly dendritic cells & macrophages) Lymph nodes, spleen - Activation (Secondary lymphoid organs) Requires co-stimulatory signals (CD80/CD86) Antigen presentation for the purpose of effector responses (B lymphocyte help, macrophage activation, etc.) Site of infection & secondary follicules HELP (Affected tissue) Antigen presentation to T helper (TH) lymphocytes (activated T cells) at site of infection so that TH cells can deliver further signals to heighten the effector responses of the cells presenting antigen (killing of microbes, antibody production, etc.) No requirement for co-stimulation Antigen presentation for the purpose of reactivation of memory T lymphocytes Top Antigen presentation to memory T lymphocytes upon re-exposure to infection to activate the anamnestic response https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 34/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 We will not go into future detail on this topic.  Depending on the cytokine(s) providing the differentiation signal(s) during antigen presentation and co-stimulation, T lymphocytes differentiate into subsets specialized into responding to specific types of infections. Top As mentioned above, there are two forms of TCRs: α:β TCRs present on the majority of T cells, and γ:δ TCRs present on some T cell subsets. T lymphocytes that express α:ß TCRs recognize peptides bound to MHC (refer to MHC & Antigen Presentation handout material). As described https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 35/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 above, α:ß TCRs that preferentially interact with peptides presented by MHC class I also express the CD8 co-receptor, while those that preferentially interact with peptides presented by MHC class II express the CD4 co-receptor. The activation of CD8+ T lymphocytes generally yields cytotoxic T lymphocytes (C TLs) and memory CTLs. The activation of CD4+ T lymphocytes yields distinct T lymphocyte effector and memory cells: T helper 1 (TH1), T helper 2 (TH2), T helper 17 (TH17, also called regulatory TH17 cells or Treg17), T follicular helper (TFH), regulatory T (Treg) cells (discussed in the next section), and many other T lymphocyte subsets we will not discuss. γ:δ TCRexpressing T cells comprise a small population of the T cell pool (<5%). γ:δ T cells can be CD4+, CD8+, or CD4-CD8- and recognize peptide and non-peptide antigens (e.g., bacterial cell wall phospholipids or bacterial glycolipids) in the presence or absence of MHC (refer to MHC & Antigen Presentation handout material and in a later section). They appear to recognize commonly occurring microbial components and contribute to the body’s first line of defense (i.e., act as part of the innate system; hence they are referred to as innate-like lymphocytes (ILLs), which include intraepithelial lymphocytes (IELs), natural killer cells (NK cells), and natural killer T lymphocytes (NKT lymphocytes); as opposed to IELs and NKT lymphocytes, NK cells do not possess a TCR however. Intraepithelial lymphocytes (IELs) are a heterogeneous group of cells that are found in the epithelium (about one IEL for 10-15 epithelial cells); 90% of these cells are T lymphocytes, and 80% of them carry CD8 and although they can have either α:α, α:ß or γ:δ TCRs, most of them express γ:δ TCRs – so we generalize that intraepithelial cells are γ:δ CD8 T lymphocytes; in contrast to the CD8 T lymphocytes you are accustomed to, these IELs do not require prior MHC-Irestricted activation by pAPCs to kill their cellular target and, therefore, are considered to be part of the innate compartment; also, they are unusual in that they kill infected cells either through MHC-Irestricted Ag presentation, just like CTLs, or through MHC-I-like molecule stimulation expressed by stressed or infected cells, just like NK cells; in a nutshell, you could think of IELs as a kind of innate CTL-NK cell hybrid! Intraepithelial cells fall into two classes: Type a IELs and Type b IELs. Type a IELs are IELs composed of T lymphocytes that follow the conventional development of T lymphocytes in the thymus (i.e., they go through both positive and negative selection), and the vast majority of them are CD8+ T lymphocytes. These possess a conventional α:ßTCR, as well as a conventional α:ß heterodimer CD8 co-receptor. They are activated in Top MALTs and the skin. Since these cells possess a conventional α:ßTCR and a conventional a:b heterodimer CD8 co-receptor, they kill cells in an MHC-I-restricted manner the way CTLs do. Also, since they undergo negative selection in the thymus, few self-reactive IELs leave the thymus. Type a IELs differ from CTLs in that they express high levels of NKG2D https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 36/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 homodimers (also found on the surface of NK cells), a receptor that activates killing by binding to non-classical MHC-I molecules (e.g., MICA & MICB). Type b IELs, on the other hand, are unconventional CD8+ T lymphocytes in that they possess an α:αCD8 homodimer co-receptor instead of the conventional α:β heterodimer CD8 co-receptor. This α:α CD8 homodimer co-receptor is paired with either a conventional α:ßTCR or α γ:δTCR. However, Type ß IEL α:ßTCRs or γ:δTCRs do not bind conventional peptide: MHC-I ligands; instead, they bind non-classical MHC-I molecules and other ligands (e.g., PAMPs). Since these cells do not interact with conventional peptide: MHC-I ligands and they possess an unconventional α:αCD8 homodimer co-receptor, there is little potential for autoimmunity; unconventional α:αCD8 homodimer co-receptors have very low affinity for conventional peptide: MHC-I ligands. Type ß IELs, like Type α IELs, possess high levels of NKG2D homodimers, which can also activate killing. The development of Type ß IELs is poorly understood. Unconventional CD8+ T lymphocytes may arise from late DN/early DP thymocytes that partially undergo positive selection in the thymus but escape negative selection due to the low-affinity α:α CD8 homodimer co-receptor. They then exit the thymus to finalize their maturation in mucosal epithelia or the skin, a process that seems to also require IL-15. Top https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 37/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 1. The virus invades a cell in the mucosal epithelium. 2. The cell that has been infected presents viral peptides to CD8 IEL through MHC class 1 molecules. 3. The activated IEL eliminates the infected epithelial cell through perforin/granzyme and the Fas-dependent pathways. 4. Infection, damage, or exposure to toxic peptides induces stress in epithelial cells, leading to expression of MIC-A and MIC-B. 5. The NKG2D receptors on IELs engage with MIC-A,B, triggering IEL activation. Simultaneously, CD8 homodimers interact with TL molecules. 6. The stressed cell is targeted for destruction by the activated IEL through the perforin/granzyme pathway. Intraepithelial cells are innate-like lymphocytes found in mucous membranes and can innately recognize and kill infected cells the same way NK cells and cytotoxic CD8 lymphocytes do. Regulatory T lymphocytes (Treg) express the α:ß-TCR, as well as the CD4 co-receptor and high levels of CD25 (a component of the receptor for IL-2, the IL-2 R α-chain) and CD152 (CTLA-4 – a negative regulator of T cell activation). Despite positive and negative selection, a few self-reactive T cells are maintained and allowed to exit to the periphery. Indeed, in order to be able to deal with microbial antigens that may closely resemble human antigens, some T cells with potential selfreactivity must be allowed to escape negative selection. Treg cells exist to suppress the response of self-reactive CD4 T cells; they are referred to as natural regulatory T lymphocytes and are important in exerting peripheral tolerance. They are characterized by the expression of CD25 and CD152 on their cell surface and by the activation of the transcriptional repressor protein FoxP3. The exact mechanism by which these cells exert their effects is not completely understood. It is clear, though, that these cells play an important role in regulating normal immune cell functions; individuals (males, as FoxP3 is encoded on the X chromosome) with FoxP3 deficiency develop fatal autoimmune disorders characterized by autoimmunity against a variety of tissues. Other T lymphocyte subsets exist but will not be developed further (this being said, some of the following subsets are going to be tested on NBME/USMLE tests at some point in the near future, but I will keep track and determine when to further develop these.). These include (1) TH3 lymphocytes, mostly another type of regulatory T lymphocyte, and function to control immune responses at the mucosa level, (2) TH22 (CD4+ IL-22 secreting cells) and cytotoxic (3) Tc22 lymphocytes (cytotoxic CD8+ cells that also secrete IL-22) cells that are involved in inflammation and wound healing, (4) TH9 lymphocytes Top (IL-9-secreting cells) involved in antitumor immunity as well as both inflammatory and regulatory processes – and involved in autoimmune disease and allergy, (5) CD8+ inducible Treg cells, (6) pathogenic TH17 cells, (7) T H1/H17, and others. Other lymphoid cells bearing similarities with T lymphocytes include natural killer cells (NK cells) and natural killer T lymphocytes (NKT cells). These cells respond to MHC-I-like proteins (refer to https://rossmed.instructure.com/courses/3318/pages/week-1-fusion-session-%7C-workshop-t-lymphocyte-development 38/50 1/17/24, 1:11 PM Week 1: Fusion Session | Workshop: T Lymphocyte Development: Hemtlgy Onclgy Infectn Imm - January 2024 the MHC & Antigen Presentation handout). Such proteins include HLA-E (human leukocyte antigen E), MICA (MHC-I chain-related gene A), and MICB (MHC-I chain-related gene B), which are induced by cellular stress to signal cytotoxic cells that stressed cells need to be eliminated. For example, binding of NKG2D homodimers, a KAR (killer activating receptor) of NK cells, to MICA or MICB on the surface of a stressed or infected cell leads to the destruction of the cell by NK cells, provided that the stressed cell expresses little or no MHC-I. Natural killer cells will be dealt with in the Innate Immunity lecture and will no longer be developed here. The NKG2D ligands consist of MHC-like molecules, MIC-A, MIC-B, RAET1 family members, and their expression is provoked by cellular stress. Infected and transformed cells express ligands (MIC-A & MIC-B (MHC class I polypeptide–related sequencesTop A & B, & UL16 binding protein 2 (ULBP2)) that engage natural killer cell killer-activation receptors (KARs, i.e. NKG2D); s

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