Intro to Immunology - Lymphocytes PDF 2024

INTRO TO IMMUNOLOGY - LYMPHOCYTES Block: Foundations Block Director: James Proffitt, PhD Session Date: Wednesday, July 31, 2024 Time: 4:00 - 5:00 PM Instructor: Lonnie Lybarger, PhD Department: Cellular & Molecular Medicine Email:...

INTRO TO IMMUNOLOGY - LYMPHOCYTES Block: Foundations Block Director: James Proffitt, PhD Session Date: Wednesday, July 31, 2024 Time: 4:00 - 5:00 PM Instructor: Lonnie Lybarger, PhD Department: Cellular & Molecular Medicine Email: [email protected] INSTRUCTIONAL METHODS Primary Method: IM13: Lecture ☐ Flipped Session ☐ Clinical Correlations Resource Types: RE18: Written or Visual Media (or Digital Equivalent) INSTRUCTIONS Please read the session objectives and notes prior to session. The notes give additional depth and can be used to supplement the lecture, if needed. Likewise, the recommended readings indicated below can be used as a supplement. READINGS RECOMMENDED Reading: Additional information on these topics can be found primarily in chapters 3-8 of Basic Immunology: Functions and Disorders of the Immune System, 7th ed. (Abbas, Lichtman, and Pillai), 2024. This textbook is available electronically through the Health Sciences library. LEARNING OBJECTIVES: 1. Describe the structure and features of antigen receptors on B and T cells, including the different protein chains and domains. 2. Explain the molecular process by which antigen receptor diversity is generated for B and T cells (VDJ recombination). 3. Explain the molecular process of isotype switching (class switching) of immunoglobulins. 4. Describe the properties and functions of each antibody isotype. 5. Identify the effector functions of Helper, Cytotoxic, and Regulatory T cells, including the major sub-lineages of Helper T cells. 6. Describe the cell types and steps involved in T cell activation, including the roles of APCs and costimulation. 7. Describe how antigenic peptides for MHC Class I and Class II are generated, and what this means for immune surveillance. 8. Explain the structural differences between MHC Class I and MHC Class II. Block: Foundations | LYBARGER [1 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES CURRICULAR CONNECTIONS Below are the competencies, educational program objectives (EPOs), block goals, disciplines and threads that most accurately describe the connection of this session to the curriculum. Related Related Competency\EPO Disciplines Threads COs LOs CO-01 LO #1 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #2 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #3 MK-02: The normal structure Immunology N/A and function of the body as a Molecular whole and of each of the major biology organ systems CO-01 LO #4 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #5 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #6 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #7 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CO-01 LO #8 MK-02: The normal structure Immunology N/A and function of the body as a whole and of each of the major organ systems CONTEXT: In this session we will explore some of the key general features of lymphocytes. This includes the generation of antigen receptors by lymphocytes to create unique lymphocyte clones, each with the potential to recognize different antigens – both foreign and self-antigens. These concepts provide an essential foundation for understanding the role of the adaptive immune system in health and disease. Block: Foundations | LYBARGER [2 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES We will then examine the respective roles of B and T lymphocytes (B and T cells) in immunity. B cells are producers of antibodies – key molecules of immunity that are present in the circulation and within tissues. Further, antibodies are widely used in medicine and biotechnology for a huge number of applications, highlighting the need to understand their properties. You will encounter antibodies again and again throughout your medical education and career. T cells exert their functions by interaction with other cell types using an important receptor/ligand combination that allows T cells to recognize cells that have encountered pathogens. T cells orchestrate adaptive immune responses and represent important targets for boosting immunity. They also can be targets for blocking unwanted immune reactions in the clinic. In the Immunity and Infection Block in Year 2, you will revisit these concepts and go into much greater depth. The intention of the immunology sessions in Foundations is to provide a basic understanding of some key concepts that you will encounter in all blocks of the curriculum. LECTURE NOTES: I. DISTINGUISHING FEATURES OF THE ADAPTIVE IMMUNE RESPONSE Specificity: - Response is directed only against the stimulating antigen Adaptability: - Responses can be made against an immense variety of antigens, even those that are not naturally occurring Self/non-self discrimination: - Responses are made against foreign (‘non-self’) antigens and not usually against ‘self’ antigens Memory: - Ability to recall previous contact with a foreign antigen and to respond in a ‘learned’ way by initiating a rapid and vigorous response following re- exposure to the antigen - Invoked during the secondary immune response, but established after the primary immune response ***permits vaccines to work! Block: Foundations | LYBARGER [3 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES II. LYMPHOCYTES - KEY PLAYERS OF THE ADAPTIVE RESPONSE B lymphocytes: make immunoglobulins (antibodies) that act as antigen receptors. Upon activation they differentiate into antibody-secreting cells known as plasma cells. T lymphocytes: express T cell receptors (TCRs) that bind antigen (in the form of peptides) and can be subdivided into: i. T helper (Th) cells (CD4+) - secrete numerous cytokines, activate dendritic cells and macrophages, and “help” B lymphocytes in producing antibodies. ii. T cytotoxic (Tc) (CD8+) cells - destroy virus-infected cells and tumor cells. iii. T regulatory (Treg) cells - regulate immune responses by controlling the activities of other immune cells including T cells – typically suppressive. Natural Killer cells: arise from the lymphocyte lineage and can kill virus- infected cells. They do NOT have antigen receptors like T and B cells. NK cells will be discussed later, in the Immune Response lecture…. Block: Foundations | LYBARGER [4 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES III. ANTIGEN RECEPTORS The Clonal Selection model, covered in the prior Introduction to Immunology session, explains key features of the adaptive immune response. The essential elements are summarized here: 1. Every individual possesses numerous clonally-derived naïve lymphocytes (T and B) with distinct antigenic specificities that exist PRIOR to contact with antigens. Specificities arise by a random DNA rearrangement of antigen receptor genes. 2. Each lymphocyte clone has many copies of an antigen receptor of a ≈single specificity on its membrane. 3. Antigen ‘selects’ and binds to the lymphocyte with a receptor with specificity for that antigen. 4. Binding to specific antigen stimulates that lymphocyte clone to proliferate. In the case of B lymphocytes, they differentiate into plasma cells that secrete antibodies that will bind that antigen. In the case of T cells, they differentiate into effector (helper or cytotoxic) cells. Some of these cells are set aside as long- lived “memory” cells in anticipation of re-exposure to the same antigen. 5. Cells capable of responding to self-antigens (auto-reactive) are destroyed following contact with self-antigen (during development) or are rendered unresponsive (in adults). B LYMPHOCYTES produce antibody molecules. Resting/naïve B cells (those that have never encountered their cognate antigen) have a cell surface form of the antibody (antibody = immunoglobulin; Ig). When antigen binds to the cell surface antibody, this initiates a series of signals (along with “help” from T cells) that leads to secretion of the same antibody type into the circulation. The antibody response is sometimes called the humoral response, because antibodies are present in plasma (and elsewhere) Antibody molecules serve a variety of functions within the immune system: Block: Foundations | LYBARGER [5 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES Opsonization = facilitates uptake/ingestion by phagocytes ANTIBODY STRUCTURE - Two copies each of a heavy-chain (MW ≈ 50,000) and light-chain (MW ≈ 25,000) - Each antibody molecule is divalent (two antigen-binding sites) - Variable (V) region binds antigen (combination of heavy- and light- chains) - Constant region (also called the Fc fragment) mediates effector functions: fix/activate complement, bind Fc Receptors on APC Block: Foundations | LYBARGER [6 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES T LYMPHOCYTES – A key feature of these cells is the T Cell Receptor (TCR). - Each T cell clone expresses one receptor type (specificity for one antigen) - Unlike B cells, T cells react with peptides derived from the breakdown of antigenic proteins. These peptides are ‘presented to T cells while bound to a Major Histocompatibility molecule (MHC; see below). T cells do not react with free antigen. The TCR has a variable region that contacts antigen, and the variable region is different in protein sequence between each T cell clone. T CELL RECEPTOR STRUCTURE: - Clonally distributed on T cells – each T cell expresses a TCR with specificity for a ≈single antigen - Two classes of TCR -  T cells are most abundant (>90%). The other class is the  T cells - Contain variable and constant regions, somewhat analogous to antibodies – diversity generated in similar way - always membrane-bound - are NOT secreted - simultaneously bind to an antigenic peptide + MHC molecule Block: Foundations | LYBARGER [7 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES Comparison of T cell receptors (TCR) and Antibody molecules Block: Foundations | LYBARGER [8 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES IV. GENERATION OF ANTIGEN RECEPTOR DIVERSITY - There cannot be a single gene for each antigen receptor - too many needed - Multiple DNA gene segments encode the variable region - segments are randomly recombined to create new combinations in developing lymphocytes (B cells in bone marrow, T cells in the thymus) - one segment of each type is used to create a functional gene (for heavy-chain: 1 x V, 1 x D, and 1 x J are randomly selected) This process is called VDJ recombination - similar mechanism for BCR and TCR Example - Immunoglobulin genes: - For Ig Heavy-Chain locus, there are about 100 V segments, one of which can be recombined with one of the 27 D segments and one of the several J segments. This creates a large number of different combinations (one combination per developing B cell). The recombining segments will encode the variable region of the antibody. - The immunoglobulin molecule also contains two identical copies of a light-chain (either  or ), and a similar mechanism is used to create diversity in the  or  genes. Ig heavy-chain locus example: In each developing B cell clone, one V, one D, and one J gene segment is selected at random to be joined together. This region of the DNA encodes the variable, antigen-binding part of the Ig heavy-chain. A similar process occurs on the light-chain gene. When the heavy- and light-chains pair, it creates a unique molecule with respect to the antigen it can bind. Block: Foundations | LYBARGER [9 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES V. ANTIBODY CLASSES (ISOTYPES) AND THEIR FUNCTIONS - different heavy-chain constant fragments (isotypes) can be appended to the same variable region - results in loss intervening regions - antibody produced maintains same specificity - developmentally controlled (IgM and IgD on naïve B cells, for example) - CD4 T cells required for switching (CD40/CD40 ligand interactions) - each isotype has distinct functional properties One rearranged antibody heavy-chain V-region can be linked to different Fc portions: In a B cell undergoing class-switching, the same Variable region is connected (through DNA splicing) to a different Constant region. The new antibody molecule that will be produced by that B cell will bind to the same antigen as before. However, because it has a different Constant region, the antibody can perform different functions, as described below. Note: this is a different process than VDJ recombination. Block: Foundations | LYBARGER [10 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES IgM Exists in serum as a pentamer (10 total binding sites); mostly confined to bloodstream due to large size Expressed on surface of naïve B cells as monomer, where it acts as antigen receptor Does NOT cross placenta. (Note: elevated levels of IgM in fetus is indicative of perinatal infection) Predominant antibody during first week of infection Strong activator of complement IgG Principal Ig of serum (≈ 80%). Consists of 4 subclasses in humans Longest half-life of all Igs (>20 days in serum) Only antibody that crosses placenta (esp. IgG1 & 3) Provides immunity to fetus for up to 1 year after birth Has opsonic activity - facilitates uptake of pathogens by macrophages and neutrophils Activator of complement Neutralization of toxins and viruses IgA Most abundant Ig in the body (when you include mucosal surfaces) May exist as monomer (in serum) or dimer (secretory IgA) Predominant form of Ig in sero-mucus secretions such as breast milk, colostrum, tears, saliva Secretory IgA protects exposed mucosal regions from pathogenic organisms IgA in breast milk is transferred to gut of newborn infant where it provides protection against newly encountered bacteria. Functions mainly as a neutralizing antibody IgE Found in extremely low concentration in serum Important in allergic reactions (binds to Fc Receptors on mast cells and basophils). When antigen binds to the IgE on the mast cell surface, it triggers degranulation of the mast cell, releasing inflammatory mediators. Induced in response to parasitic infections, such as worms. Block: Foundations | LYBARGER [11 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES VI. T CELL SUBSETS CD4+ = T-helper (TH): - TCR reacts with antigenic peptides presented by MHC class II - peptides derived from exogenous sources (viruses, bacteria, etc.) - “help” B cells make antibody (cytokines and cell-cell interactions) - “help” macrophages become active (via cytokines) - secrete factors that influence nature of immune response - CD4 T cells, when activated, can differentiate into several distinct subtypes (sub- lineages). Two major subtypes of CD4 T cells are the TH1 and TH2 cells. TH1 cells are important for macrophage activation to deal with intracellular bacteria. TH2 cells direct anti-parasite responses. TH2 cells are also important in some allergic reactions such asthma. Other CD4 subsets exist and are an area of intensive research. CD8+ = T-Cytotoxic (T c): - TCR reacts with antigenic peptides presented by MHC class I - peptides derived from endogenous sources in most cases (proteins synthesized within the cell. During virus infection, for example, virus-encoded proteins are synthesized within the cell.) - cytolytic activity toward virus-infected and tumor cells Block: Foundations | LYBARGER [12 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES (T regulatory cells = Treg): - CD4+ CD25+, T cells whose T cell receptors react to peptide presented on MHC class II (MHC class II-‘restricted’) - often inhibitory and are critical to prevent autoimmunity VII. ANTIGEN-PRESENTING CELLS AND THEIR INTERACTION WITH T CELLS - Naïve (antigen-inexperienced) T lymphocytes become activated and exhibit the effector functions mentioned above after contact with peptides displayed on antigen-presenting cells (APC; i.e., dendritic cells, macrophages, Langerhans cells, B cells). Signals from the APC influence the differentiation of the T cells (T H1 versus TH2 differentiation of CD4 T cells, for example). - Antigen-presenting cells ingest, process, and ‘present’ antigen in the form of peptides that are recognizable by T cells (bound to MHC molecules). - As with B lymphocytes, naïve T cells proliferate and differentiate into effector (helper or cytotoxic) cells upon stimulation. Some of these cells are set aside as ‘memory’ cells in anticipation of re-exposure to the same antigen. - APCs express costimulatory molecules that are also necessary to activate T cells - In addition to peptide + MHC, APCs provide co-stimulation in the form of cell surface receptor/ligand interactions (like B7/CD28) and cytokines that are required for activation of naïve T cells. Without co-stimulation, T cells are not activated by antigen. - Among the different types of APCs, dendritic cells are the most potent at activating T cells. However, macrophages and B cells can function as APCs in specific and important contexts, as will be described in the “Immune Response” and “Hypersensitivity Reactions” lectures. Block: Foundations | LYBARGER [13 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES Regulation of APC function during an immune response - In response to infection, APCs mature (become activated), making them much better at stimulating naïve T cells - Variety of stimuli can activate APC: - Pathogen-associated molecule patterns (TLR, NLR ligands), - Inflammatory cytokines VIII. ANTIGEN PRESENTATION PATHWAYS Two types of MHC molecules important for T cell responses: Major Histocompatibility Complex Class I and Class II - Peptides are loaded onto MHC molecules within cells and are then transported to the surface where they are presented to the T cell. - All nucleated mammalian cells can express Class I MHC. - Only professional APCs normally express Class II MHC. MHC CLASS II (MHC-II) - Expression is mostly limited to dendritic cells, B cells, and macrophages - The TCRs found on CD4 T cells react with MHC-II + peptide Block: Foundations | LYBARGER [14 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES Steps: i. APC takes up exogenous material via endocytosis. ii. endocytic vesicle fuses with a lysosome, where protein contents are digested into short peptides iii. newly-synthesized MHC-II molecules are transported to the lysosome where they bind peptides iv. MHC-II + peptide complex is transported to the cell surface v. If a CD4 T cell has a receptor capable of recognizing the peptide, the T cell can become activated and: - help the macrophage become fully activated - help the B cell to secrete antibody MHC-II peptide-loading pathway: MHC CLASS I (MHC-I) - Expressed by virtually all cell types in the body - The TCRs found on CD8 T cells react with MHC-I + peptide - In the absence of infection, the class I molecules are loaded with peptides from normal cellular proteins – the immune system ignores these “self” peptides. Steps: i. proteins synthesized within the cell are degraded by the proteasome into peptides ii. peptides are transported in the endoplasmic reticulum (ER) of the cell through a dedicated transporter complex iii. newly-synthesized MHC-I molecules are loaded with peptides in the ER lumen iv. MHC-I + peptide complex is transported to the cell surface v. If a CD8 T cell has a receptor capable of recognizing the peptide, the T cell can become activated and: - kill the cell expressing the foreign peptide antigen - secrete interferon-gamma (activates macrophages and upregulates MHC expression) Block: Foundations | LYBARGER [15 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES MHC-I antigen presentation pathway: MHC Molecules – Comparison of MHC-I and -II IX. MHC MOLECULES AND GENETICS - More diseases associated genetically with MHC genes than any other family of genes - MHC is a large locus: ≈ 4 megabases in humans on chromosome 6 that contains >200 genes - Many involved in the immune system, but not all - ‘MHC molecules’ typically refers to just class I or class II gene products - MHC-I and -II are primarily responsible for allograft rejection - MHC in humans is called “Human Leukocyte Antigen; HLA” Block: Foundations | LYBARGER [16 of 17] INTRO TO IMMUNOLOGY - LYMPHOCYTES MHC molecules are peptide-binders MHC genes - Polygenic and Polymorphic: Polygenic (multiple genes) MHC Class I: 3 genes (HLA-A, HLA-B, and HLA-C in humans) - gene encodes the heavy-chain (or -chain), which binds antigen - a separate light chain (2-microglobulin; 2m) completes the structure Class II: 3 genes pairs (HLA-DP, HLA-DQ, and HLA-DR) - two linked genes encode two proteins of equal size (e.g., HLA-DR and HLA-DR); together, the two chains bind peptide Polymorphic (multiple alleles/variants of each gene, leading to a high degree of heterozygosity) >13,000 MHC-I variant alleles and >5,000 MHC-II variant alleles in the human population; MHC- I and -II are the most polymorphic proteins in mammals. - These allelic differences are primarily responsible for transplant rejection reactions and have important implications for immune responses. These features of MHC molecules will be covered in greater detail in the Immunity and Infection Block. - Each variant MHC molecule can bind different sets of peptides from any larger antigenic protein. Block: Foundations | LYBARGER [17 of 17]