DISEASES OF THE IMMUNE SYSTEM Disorders caused: a. Allergic reactions b.

Autoimmune reactions THE NORMAL IMMUNE RESPONSE Classical definition: defense against infectious pathogens Mechanism of protection, 2 categories: a. Innate immunity b. Adaptive immunity

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Leukocyte A. Pattern recognition receptors recognize molecular patterns: a. Pathogen associated molecular patterns: components of related microbes essential for infectivity - recognized by leukocytes and epithelial cells b. Danger-associated molecular patterns: released by necrotic and injured cells - recognized by leukocytes *Toll-like receptors (TLR) specific for components of different bacteria and viruses - Located on cell surface and endosomes - Recognize and initiate cellular response to extracellular and ingested microbes - TLR and other sensors signal by a common pathway activation of cytokines and proteins stimulate microbial activities of cells, especially phagocytes B. Receptors that bind microbes for phagocytosis: - Receptors for mannose residues mannose are typical of microbial and not host glycoproteins - Receptors for opsonins antibodies and complement proteins coating the microbes Monocytes enter tissues and mature macrophages Dendritic cells - produce type I interferon & anti-viral cytokines inhibit viral infection and replication Natural Killer Cells provide early protection Plasma Proteins: a. Proteins of Complement System - activated by microbes(innate immunity) via alternative and lectin pathway - activated by antibodies (adaptive immunity) via classical pathway b. Mannose-binding lectin c. CRP

Adaptive Acquired, specific Develops later, after exposure Stimulated by microbial and nonmicrobial substance More powerful *immune response refers to adaptive immunity INNATE IMMUNITY Major components: 1. Epithelial barriers block entry of microbes 2. Phagocytic cells neutrophils and macrophages 3. Dendritic cells 4. Natural killer cells 5. Plasma proteins complement system, etc Two most important cellular reactions of innate IS: a. Inflammation phagocytic leukocytes are recruited and activated b. Anti-viral defense mediated by dendritic cells and NK cells Epithelial cells - Skin, GI, RT provide mechanical barriers to entry of microbes - Also produce antimicrobial molecules such as defensins - Contains some lymphocytes that combat microbes

Innate Natural, native Present before infection Always ready -> first line of defense

Surfactant component of innate immunity, providing protection against inhaled microbes ADAPTIVE IMMUNITY - Consists of lymphocytes and their products, including antibodies - Receptors are much more diverse than innate IS - Lymphocytes are not inherently specific for microbes can recognize a vast array of foreign substance 2 types of adaptive immunity: Humoral Extracellular microbes B-cells (bone marrow derived) Antibodies

TCR diversity During T cell development, TCR genes rearrange to form many different combinations - Mediated by enzyme produced by RAG 1 and RAG 2 genes (Recombination Activating Genes) defect results in failure to generate mature lymphocytes - only T cell contains rearranged TCR genes but all cells in the body contain TCR genes in germ-line configuration, unarranged - Each T cell expresses TCR molecule of one specificity - Presence of rearranged TCR genes can be demonstrated by molecular analysis, marker of T-lineage cells - Each T cell and progeny have unique DNA arrangement possible to distinguish polyclonal (non-neoplastic) t cell proliferations from monoclonal/neoplastic - Analysis of antigen receptor gene rearrangements is a valuable assay for detecting lymphoid tumors TCR complex: *CD3 protein complex + chains - Involved in transduction of signals into the T cell after the TCR has bound the antigen - identical in all T cells b. TCR recognizes peptides, lipids and small molecules, without requirement for display by MHC - aggregate at epithelial surface of skin, mucosa of GI and UG tract suggest protective role c. NK T-cells small subset of t cells found on NK cells - Recognize glycolipids displayed by CD 1 (MHC-like molecule) Other proteins that assist T-cells: a. CD4 b. CD8 c. CD 2 d. CD 28 e. Integrins

Cell-mediated Intracellular microbes T cells (thymus-derived)

CELLS OF THE IMMUNE SYSTEM - Capable of migrating among lymphoid and other tissues Lymphocytes specialized in molecular properties and function a. Naive lymphocytes have not yet encountered antigen b. Effector cells activated naive lymphocytes by antigens c. Memory cells live in state of heightened awareness T LYMPHOCYTES - Precursor from thymus - Mature T cells are found in a. blood constitute 60-70% of lymphocyte content b. T-cell zones at peripheral lymphoid organs T-Cell Receptor (TCR) antigen specific a. TCR (95% of t cells) - Have disulfide-linked alpha and beta polypeptide chains 2 regions per chain: > Variable region antigen-binding region > Constant region - recognize antigens displayed by MHC molecules on surface of APCs MHC restriction limited specificity of t cells to cell-associated antigens

CD4 and CD8 are expressed exclusively in different TCRs *coreceptors in T-cell activation work with antigen receptor in response to antigen initiate signals that are necessary for activation of T cells CD4 expressed on 60% of CD3+ cells - Function as cytokine-secreting cells that help macrophages and B lymphocytes to combat infections - Binds to type II MHC CD8 expressed in 30% of cytotoxic t cells - Binds to type I MHC B LYMPHOCYTES - Precursor from bone marrow - Mature B cells constitute 10-20% of circulating lymphocytes - Also present in peripheral lymphoid tissues: lymph nodes, spleen - Develop into plasma cells after antigen stimulation secrete antibodies - Recognize antigen via B-cell Antigen Receptor Complex - Antigen binding component: membrane bound IgM and IgD on surface of all mature naive b cells B-cell Antigen Receptor Complex - Dimer: Ig and Ig - unique antigen specificity derived from RAG-mediated rearrangements of Ig genes - Analysis of Ig gene rearrangements is useful for identifying monoclonal B-cell tumors Other expressed molecules: a. Complement receptors - Type 2 complement receptors is also receptor for EBV hence EBV readily infects B cells b. Fc receptors c. CD 40 DENDRITIC CELLS - have numerous fine cytoplasmic processes that resemble dendrites - 2 types

A. Interdigitating dendritic cells - Most important APC for initiating primary t-cell responses against protein antigens Key features: 1. Located under epithelia (common site of entry) and in the interstitia of all tissues (where antigens may be produced) *Langerhans cells immature dendritic cells within epidermis 2. Express many receptors for capturing and responding to microbes: TLR and mannose receptors 3. Can be recruited to T-cell zones of lymphoid organs ideal location to present antigens to T cell 4. Express high levels of molecules needed for presenting antigen and activating CD4+ T cells B. Follicular dendritic cells - Present in germinal centers of lymphoid follicles in the spleen and lymph nodes - Bear Fc receptors for IgG and receptors for C3b - Can trap antigen bound to antibodies or complement proteins role in humoral immunity is presenting antigens to B cells MACROPHAGES - Function as APC in T-cell activation: macrophages process antigens and peptide fragments from phagocytosed microbes to T cells - Fxn in cell-mediated immunity as key effector cells in eliminating intracellular microbes (activated by T cells) - Fxn in effector phase of humoral immunity phagocytose and destroy microbes that are opsonised (coated) by IgG or C3b NATURAL KILLER CELLS - 10-15% of peripheral blood lymphocytes - Do not express TCR or Ig - Larger than lymphocytes and contain abundant azurophilic granules Large Granular Lymphocytes - Can kill variety of infected and tumor cells without prior exposure to or activation by microbes provides early line of defense against viral infections and some tumors - Two cell surface molecules: CD 16 and CD 56 used to identify NK cells

CD 16, an Fc receptor of IgG, confers to NK cells ability to lyse IgGcoated target cells Antibody-dependent Cell-mediated Cytotoxicity (ADCC) Functional activity is regulated by balance between signals from activating and inhibitory receptors:

(+) APC in the nodes sample antigens of microbes carried in the lymph, which entered through epithelia Dendritic cells pick up and transport antigens of microbes from epithelia via lymphatic vessels to lymph nodes - Antigens of microbes that enter through epithelia or colonize tissues become concentrated in draining lymph nodes Spleen abdominal organ that serves same role as that of lymph nodes in response to lymph-borne antigens - Blood entering the spleen flow through a network of sinusoids dendritic cells and macrophages trap blood-borne antigens Mucosal and cutaneous lymphoid tissues - Located under epithelia of the skin, and the GI and respiratory tract respectively - Responds to antigens that breach epithelium - At any time, half of bodys lymphocytes are in mucosal tissues, mostly memory cells - Example of mucosal lymphoid tissues: Pharyngeal tonsils and Peyers patches of intestine

a. NK cell inhibitory receptors recognizes self-class I MHC molecules, which are expressed in all healthy cells - Prevent NK cells from killing normal cells - 2 major families: > killer cell Ig-like receptors > CD94 family of lectins (CHO-recognizing proteins) b. Activating receptors: NKG2D receptors recognize surface molecules induced by various kinds of stress, such as infection and DNA damage - Virus infection or neoplastic transformation (a) induce expression of ligands for activating receptors (b) reduce expression of class I MHC balance is tilted toward activation infected or tumor cell is killed NK cells also secrete cytokines: IFN-y activate macrophage provide early defense against intracellular microbial infections Activity is regulated by: IL-2 and IL-15 stimulate proliferation of NK cells IL-12 activates killing and secretion of IFN-y

Segregation of T lymphocytes and B lymphocytes in lymph nodes: B cells concentrated in discrete structures, follicles, around periphery/cortex of each node - has central region if recently responded to antigen germinal center - Follicles contain follicular dendritic cells that present antigen to T cells T cells concentrated in paracortex, adjacent to follicles; in periarteriolar lymphoid sheaths surrounding small arterioles *when lymphocytes are activated altered expression of chemokine receptors that regulate distribution of b cells and t cells cells migrate toward each other meet at edge of follicles helper t cells interact with b cells to differentiate into antibody-producing cells

TISSUES OF THE IMMUNE SYSTEM A. Generative (primary or central) lymphoid organs in which T and B lymphocytes mature and become competent to respond to antigens 1. Thymus: principal generative lymphoid organ, where t cells develop 2. Bone marrow: site of production of all blood cells, where b lymphocytes mature B. Peripheral (secondary) lymphoid organs in which adaptive responses are initiated Lymph nodes nodular aggregates of lymphoid tissue located along lymphatic channels throughout body

Lymphocyte Recirculation - Lymphocytes constantly circulate - Naive lymphocytes traverse the peripheral lymphoid organs where immune responses are initiated - Effector lymphocytes migrate to sites of infection and inflammation - Plasma cells remain in lymphoid organs and do not need to migrate to sites of infection they secrete antibodies that are carried to distant tissues Naive lymphocyte exit thymus migrate to lymph nodes enter T-cell zones through high endothelial venules (HEV) encounters antigen in the lymph nodes that it specifically recognize on the surface of APC activated and differentiate into effector T cells leave lymph nodes and enter circulation migrate into the tissue that harbour the microbes.

3 groups: Class I MHC expressed on all nucleated cells and platelets - Encoded by HLA-A, HLA-B, HLA-C - Heterodimer linked noncovalently: a. alpha or heavy chain - polymorphic - extracellular region has 3 domains: 1, 2, and 3 - 1 and 2 forma cleft or groove where peptides bind, with polymorphic residues lining the sides and base - 3 has binding site for CD8 b. peptide beta2 microglobulin not encoded within the MHC; nonpolymorhphic Display: peptides that are derived from proteins (ex: viral antigens) that are: a. located in the cytoplasm b. usually produced in the cell transported to ER peptides bind to class I MHC stable dimer is transported to cell surface Recognized by: CD8+ T lymphocytes CD8 binds to heavy chain function as CTL eliminate virus *CD8+ T cells are class I-MHC restricted Class II MHC molecules - encoded in HLA-D 3 subgroups: HLA-DQ, HLA-DP, HLA-DR - heterodimer: and chains (both polymorphic) extracellular portion has 2 domains each: 1 and 2, 1 and 2 2 domain has binding site for CD4 recognized by CD4+ T cells Class II-MHC restricted 1 and 1 domains form peptide-binding clefts facing outward (portion where class II alleles differ) Display: antigen that are internalized into vesicles, derived from extracellular microbes and soluble proteins internalized proteins are proteolytically digested in endosomes or lysosomes peptide

MAJOR HISTOCOMPATIBILITY MOLECULES - Peptide display system of adaptive immunity - Fundamental to recognition by t cells - Discovered as products of genes that evoke rejection of transplanted organs - Responsible for tissue compatibility between individuals - Physiologic function: display peptide fragments of proteins for recognition by antigen-specific t cells - Genes encoding for MHC are clustered on a small segment of Chromosome 6: Major Histocompatibility Complex or Human Leukocyte Antigen (HLA) (MHC-encoded proteins were initially detected on leukocytes) *HLA system is highly polymorphic there are many alleles of each MHC gene in the population each individual inherit different sets barrier in organ transplantation

fragments associate with class II MHC in the vesicles stable class II-peptide complex transported to cell surface *In contrast to class I, class II are mainly expressed on cells that present ingested antigens and respond to T-cell help: macrophages, B lymphocytes, and dendritic cells *MHC locus also contains genes that encode some complement components and cytokines TNF and lymphotoxin *Class II locus contains genes that encode many proteins involved in antigen processing and presentation HLA haplotype combination of HLA alleles in an individual - an individual inherits one set of HLA genes from each parent expresses 2 different molecule per locus - polymorphism of HLA loci innumerable combinations no 2 individuals (other than identical twins) are likely to express same MHC molecules grafts from other individuals are recognized as foreign and attacked MHC role in regulating T-cell mediated immune response: 1. an individual mounts an immune response against a protein only if he/she inherits the gene(s) for those MHC molecule(s) that can bind peptides from the antigen and present it to T cells examples: *individual has class II molecules capable of binding ragweed pollen antigen genetically prone to allergic reaction against pollen *inherited capacity to bind bacterial peptide evoke protective antibody response resistance to infection 2. MHC ensure that correct immune response is mounted against different microbes by segregating cytoplasmic and internalized antigens: *CTL cytoplasmic microbes *T-cell activated antibodies and macrophages extracellular microbes

HLA and Disease Association Most striking: association between ankylosing spondylitis and HLA-B27 - Individuals have 90x greater chance of developing the disease Categories of HLA-associated Diseases: 1. Inflammatory diseases ankylosing spondylitis, some postinfectious arthropathies (HLA-B27) 2. Autoimmune diseases autoimmune endocrinopathies (alleles at DR locus) 3. Inherited errors of metabolism 21-hydropxylase deficiency (HLABW47) and hemochromatosis (HLA-A) Disease Ankylosing spondylitis Postgonococcal arthritis Acute anterior uveitis RA Chronic active hepatitis Primary Sjogren syndrome Type 1 DM HLA Allele B27 DR4 DR3 DR3 DR4 Both Risk % 90-100 14 4 13 9 5 6 20

CYTOKINES - Messenger molecules of immune system - Short-acting secreted mediators - Molecularly defined cytokines are called interleukins they mediate communication between leukocytes - Wide spectrum of effects - Produced by several different cell types Categories: 1. Cytokines of innate immunity - Produced rapidly in response to microbes and other stimuli - Made principally by macrophages, dendritic cells and NK cells - Mediate inflammation and anti-viral defense - TNF, IL-1, IL-2, type I IFN, IFN-y, and chemokines

2. Cytokines of adaptive immunity - Made principally by CD4+ lymphocytes in response to antigens and other signals - Promote lymphocyte proliferation and differentiation - Activate effector cells - IL-2, IL-4, IL-5, IL-17, and IFN-y 3. Colony stimulating factors - cytokines that stimulate hematopoiesis from bone marrow progenitors - increases leukocyte numbers during immune and inflammatory responses, and to replace leukocytes that are consumed Therapeutic application: *TNF antagonist molecularly-targeted therapy for RA *Recombinant cytokines enhance immunity against cancer or microbial infections (immunotherapy)

*CD4+ effector T cells of TH1 subset recognize antigens of microbes ingested by phagocytes activate phagocyte to kill microbe - also induce inflammation *CD8+ CTL kill infected cells harbouring microbes in the cytoplasm *TH2 cells defense against helminthic infections B lymphocytes use antigen receptors (membrane-bound antibody molecules) to recognize antigens of many different chemical types Microbe-elicited innate immune response: - same time as antigens are recognized by T and B lymphocytes - if (+) immunization innate response is induced by the adjuvant given with the antigen microbe activates APCs to express molecules called costimulators and secrete cytokines to stimulate proliferation and differentiation of T lymphocytes *principal costimulators of T-cells: B7 proteins (CD80 and CD86) expressed on APC recognized by CD28 receptors of Naive T cells Signal 1 antigen Signal 2 costimulator - function cooperatively to activate lymphocytes - requirement for signal 2 ensures that adaptive immune response is induced by microbes and not by harmless substances - in tumors or transplants, signal 2 may be provided by substances released from necrotic cells (danger-associated molecular patterns) CELL MEDIATED IMMUNITY: ACTIVATION OF T LYMPHOCYTES - elimination of intracellular microbes Antigen and costimulators in peripheral lymphoid organs Activate naive T cells to proliferate and differentiate Migrate into site where antigen/microbe is present ---CD4+ T helper cells express CD40 ligand Engages CD40 in macrophages or B cells to activate these cells

OVERVIEW OF LYMPHOCYTE ACTIVATION AND IMMUNE RESPONSES ANTIGEN RECOGNITION *Clonal selection hypothesis specific lymphocytes for antigens exist before exposure when antigen enters, it selects the specific cells and activates them microbes and protein antigens captured by dendritic cells (DC) that are resident in epithelia and tissues carry antigenic cargo to draining lymph nodes, DC mature and express high levels of MHC molecules and costimulators MHC-associated peptide antigens are displayed on DC are recognized by naive T-cells activated to proliferate and differentiate into effector and memory T cells Differentiated T-cells enter circulation and migrate to sites of infection

*Helper T Cells functions are mediated by combined actions of CD40ligand (CD40L) and cytokines One of earliest response of CD4+ helper T cells: secretion of IL-2 and expression of high-affinity receptors for IL-2 - IL-2 is a growth factor that acts on T lymphocytes stimulate proliferation increase number *Effector cells can secrete different sets of cytokines, and thus perform different functions Ex: differentiated CD4+ T helper cells TH1 and TH2 subsets TH1 subset: - TFN-y: potent macrophage activator combine with CD40mediated activation induction of microbicidal substances in macrophages destruction of ingested microbes TH2 subset - IL-4: stimulates B cells to differentiate to IgE-secreting plasma cells - IL-5: activates eosinophils *Eosinophils and mast cells bind to IgE-coated microbes such as helminthic parasites function to eliminate helminths TH17 subset discovered recently - IL-17: powerful recruiters of neutrophils and monocytes neutrophilic inflammation in some bacterial and fungal infections TH1 IFN-y IFN-y, IL-12 Macrophage activation IgG production Intracellular microbes Inflammatorymediated, often autoimmune, dses TH2 IL-4, IL-5, IL-13 IL-4 Mast cells and eosinophil activation IgE production Helminthic parasites Allergies TH17 IL-17, IL-22, chemokines TGF-B, IL-6, IL-1, IL-23 Recruitment of neutrophils and monocytes Extracellular bacteria, fungi Inflammatorymediated, often autoimmune, dses

Activated CD8+ lymphocytes differentiate into CTL kill cells harbouring microbes (eliminate reservoirs of infection) HUMORAL IMMUNITY: Activation of B Lymphocytes - Elimination of extracellular microbes - Upon activation, B lymphocytes proliferate and differentiate into plasma cells secrete antibodies with distinct functions *Polysaccharide and lipid antigens have multiple identical antigenic determinants epitopes that are able to engage many antigen receptor molecules on B cells and activate it *Typical globular protein antigens not able to bind to many antigen receptors requires help from CD4+ T cells for full response of B cells B cells ingest protein antigens into vesicles, degrade them and display peptides bound to MHC molecules for recognition by helper T cells helper T cells express CD40L and secrete cytokines work together to activate B cells

Plasma Cells secrete antibodies that have same antigen binding site as B cell receptors/antibodies that first recognized the antigen *polysaccharides and lipids mainly of IgM antibody *protein antigens antibodies of different classes or isotypes: IgG, IgA, IGE - By virtue of CD40L and cytokine-mediated T cell action - Isotype switching can be induced by IFN-y and IL-4 occur mainly in germinal centers formed by proliferating B cells *Affinity maturation helper T cells stimulate production of antibodies with high affinities for the antigen improve quality of humoral immune response Humoral immune responses: Antibodies binds to microbes and prevents them from infecting cells (neutralizing microbes) IgG antibodies coat/opsonize microbes and target them for phagocytosis (phagocytes have receptors for Fc tails of IgG)

IgG and IgM activate complement system by classic pathway complement products promote phagocytosis and destruction of microbes *Opsonizing and complement-fixing IgG is stimulated by TH1 helper cells respond to most bacteria and viruses IgA secreted from mucosal epithelia neutralized microbes in the lumen of RT and GIT (and other mucosal tissues) IgG is actively transported across placenta protects newborn until immune system becomes mature IgE and eosinophils cooperate to kill parasites by release of eosinophilic granules that are toxic to the worms (orchestrated by TH2 cells) Most circulating IgG antibodies have half life of 3 wks Some antibody-secreting plasma cells migrate to bone marrow and live for years and continue to produce low levels of antibodies

Decline of Immune Response and Immunological Memory - Majority of effector lymphocytes die by apoptosis after microbe is eliminated - Generated memory cells are long-lived and survive for years after the infection Memory cells are expanded pool of antigen-specific lymphocytes More numerous than naive cells Respond faster and more effectively when re-exposed to antigen than naive cells Generation of memory cells is an important goal of vaccination