Acquired and Innate Immunity OVERVIEW OF IMMUNOLOGY Lee S.F. Soderberg, Ph.D.

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Objectives: This lecture is an introduction to the major cell types and organs of the immune system. While later lectures will cover these in more detail, you should be familiar with the maturation, identification, and function of the major cell types and the organ sites of their activity. I. HEMATOPOIESIS Hematopoiesis, the generation of blood cells and cells of the immune system, occurs throughout life to provide a constantly renewing supply of these vital cells. Hematopoietic stem cells differentiate early into erythroid, myeloid, and lymphoid progenitors. Myeloid cells give rise to two cell lineages important to the immune system: neutrophils and monocytes. Monocytes differentiate into macrophages and dendritic cells. Neutrophils, monocytes, and macrophages are phagocytes, which engulf and kill microorganisms. Dendritic cells take up foreign antigens, which they use to stimulate lymphocytes. Lymphoid progenitor cells differentiate into B cells, which make antibodies, and two main classes of T cells: CD8+ T cells and CD4+ T cells. CD8+ T cells are cytotoxic for tumor cells and virusinfected cells. CD4+ T cells are also called helper cells, because they modulate the activities of other cells through a variety of factors, called cytokines. Natural killer (NK) cells, CD16 + CD56+, are cytotoxic cells involved in innate immunity. II. INNATE VS. ACQUIRED IMMUNITY Infections depend upon the ability of microorganisms to gain access to internal tissues and to survive and proliferate there. Innate resistance mechanisms non-specifically block and/or kill organisms before they become established. Barriers, such as skin and mucous membranes, are assisted by cilia and secretions to keep organisms out. Enzymes (lysozyme in tears), low pH (stomach acid), and normal flora discourage potential invaders. Organisms that gain access to tissues are phagocytized and killed by scavenger cells, neutrophils and macrophages. Scavenger cells produce antimicrobial peptides, called defensins that can kill certain classes of organisms. They also produce hydrolytic enzymes and toxic oxygen radicals that can kill organisms. Natural killer (NK) cells can kill virus-infected cells and tumor cells. Innate immune mechanisms are normally present and generally do not require induction or activation. However, triggered by microbial components through Toll-like receptors (TLR), scavenger cells are stimulated to increase production of enzymes, defensins, and oxygen radicals. Phagocyte and NK cell activities are enhanced by soluble factors (cytokines) from lymphocytes, macrophages, or NK cells. IMPORTANT CONCEPT 1: Acquired immune responses depend upon the specific activation of lymphocytes by antigen. If innate mechanisms fail to keep microorganisms out, acquired immune mechanisms may become activated. These include B cell production of antibodies, CD8 + cytotoxic T lymphocytes, and CD4+ T cell-mediated activation of other lymphocytes and of macrophages. By definition, the acquired immune mechanisms involve the antigen-specific activation of lymphocytes. This complex process and its regulation will be the principal subjects of this lecture series in immunology.
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III. INNATE IMMUNITY A. Normal flora Microbiological barriers: Normal flora compete with pathogens for nutrients and attachment sites. B. Complement (See complement SDL). Ceratin microbial cell walls bind C3 or C3b, activating the alternative complement pathway. Phagocytes secrete a C1q analogue, mannose-binding lectin (MBL), that binds to microbial cell walls, activating the lectin complement pathway. Complement activation can lyse certain microbes or opsonize microbes, providing a target (C3b) for phagocytosis. Complement by-products, C2a, C5a, and C4a, increase vascular permeability, allowing and attracting (C5a) to the site of infection. C. Phagocytes Macrophages and neutrophils engulf and try to kill microbes. Macrophages are present in tissues, particularly those lining the GI and respiratory tracts, in alveoli, and in the liver. Neutrophils circulate in the blood and are the first-responders to infections, attracted by chemokines. Neutrophils are shorter-lived than macrophages that arrive late, but last longer. Phagocytes recognize microbes by receptors that lack the fine specificity of antibodies anf T cells. Phagocytes express receptors, pathogen recognition receptors (PRR). These receptors include scavenger receptors, mannose receptors (mannose is common in pathogens, but rare in human cells), and Toll-like receptors (TLR). TLR recognize microbial constituents or pathogen associated molecular patterns (PAMP). There are 9 or 10 TLR that bind certain microbial carbohydrates, proteins, or nucleic acids. For instance, TLR-4 binds the Gram-negative bacterial constituent, LPS. IMPORTANT CONCEPT 2: microbes. Phagocytes have limited specificity for recognizing

On engulfment, phagocyte cell membranes surround of microbes forming a vacule, phagosome. Phagocyte vesicles, lysosomes, containing lysosomal enzymes and reactive oxygen intermediates (superoxide, nitric oxide) fuse with the phagosomes. In the resulting phagolysosome, the enzymes and oxygen radicals can kill certain microbes. Chronic granulomatous disease (CGD) is an inherited immunodeficiency caused by a mutation of the NADPH oxidase system, precluding oxygen radical formation, primarily in neutrophils. Patients with CGD suffer from chronic bacterial infections. D. Inflammation Upon recognition/binding of microbes, phagocytes produce mediators, cytokines and chemokines, that stimulate inflammation. Chemokines are a family of 36 or so chemoattractant molecules. Chemokines are classified into four groups, the most prevalent of which are CC (two adjacent cysteines) and CXC (two cysteines separated by
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another amino acid). In general, CC-chemokines attract monocytes, while CXCchemokines attract neutrophils. Inflammatory cytokines include interleukin-1 (IL-1), IL-6, and tumor necrosis factor- (TNF-). The hallmarks of inflammation are heat, pain, erythema and swelling, related to cytokine-induced local dilation of blood capillaries. Vasodilation allows phagocytes to migrate to the site of infection in response to chemokines. E. Anti-viral activity Among the cytokines released in inflammation, are type I interferons (IFN- and IFN-). Type I interferons interfere with viral replication and have anti-tumor activity. Natural Killer cells (NK cells) are large granular lymphocytes that also provide defense against viral infections and tumor cells. NK cell activity will be presented with T cell cytotoxicity. Patients with immunodeficies in NK cell function have persistent viral infections and are more susceptible to neoplasias. IV. ACQUIRED IMMUNITY A. Antigens and antibodies Antigens are compounds that are recognized as foreign by the acquired immune system through binding to antibodies, often on B cells, and/or analogous receptors on T cells (TCR). Very small antigens, called haptens, are recognized as foreign, but are too small to induce an immune response unless bound to a larger antigen. Larger antigens, also called immunogens, can stimulate specific immune responses by inducing T and/or B cell activation through interactions with their receptors. Foreign proteins and carbohydrates generally stimulate the strongest immune responses. Nucleic acids are less immunogenic and lipids are least immunogenic. Poor immunogens become more efficient when covalently bound to proteins. Antibodies are 150 kD (monomer) glycoproteins produced by B cells that have the capability of selectively binding antigens. The small region on an antigen recognized by the antibody is called an antigenic determinant or an epitope. Antibodies can inactivate some toxins and induce the lysis or phagocytosis of certain microorganisms. B. Clonal selection The activation of both T cells and B cells follows the tenets of clonal selection. 1. During T cell differentiation in the thymus and B cell differentiation in the bone marrow, a broad repertory of specificities (~10 7 for T cells and ~10 8-109 or more for B cells) develops in the absence of any foreign antigens. 2. Each cell has only a single specificity to which it can respond. 3. Stimulatory exposure to an antigen causes the proliferation of only those lymphocytes with receptors that bind that antigen. 4. As each stimulated cell proliferates, it forms a clone of cells. Each cell in a clone has antigen receptors with identical specificity and, in the case of B cells, produces antibodies with the same specificity. C. Immunity in neonates

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The newborn baby is generally considered not to be immunocompetent (capable of appropriate and adequate immune responses). It is estimated that the newborn is capable of responding to 104 antigenic specificities. The acquisition of responsiveness is both gradual and predictable (responsiveness to a particular antigen is acquired at about the same age in all normal babies). Thus, administration of childhood vaccines is recommended at specific ages. Administration of a vaccine earlier than recommended will be ineffective and may be deleterious (see Tolerance, Lecture 27). 1. Maternal antibody. While newborn babies have a limited capacity to respond to antigens, they do have passive immunity derived from the circulating antibodies of their mother. Whatever specific antibodies the mother has, will be transferred across the placenta to the baby.

IMPORTANT CONCEPT 3: Passive immunity does not provide lasting protection IgG is the only class of antibodies which crosses the placenta. Because these antibodies have a half-life of 25-35 days, maternal antibodies are gone from the babys circulation by about 6 months after birth. By this time the baby has developed a somewhat better ability to produce its own antibodies. IgG and IgA antibodies are present in colostrum and mother's milk. These antibodies generally stay in the baby's intestinal tract and are important in resistance to enteric infections. D. Immunity in adults 1. Central Lymphoid Organs (immune cell production) Bone marrow a. Formation - hematopoietic function starts at about 4 mo of gestation b. Organization - loose, containing mostly hematopoietic progenitor cells and stromal cells. Stromal cells include macrophages, adipocytes, and fibroblasts and are a major source of hematopoietic growth factors or cytokines. c. Function 1) Growth and differentiation of blood cells and cells of the immune system. 2) B lymphocyte maturation. Membrane antibodies on B cells serve as specific antigen

IMPORTANT CONCEPT 4: receptors

B cells are lymphocytes characterized by the presence of membrane-bound antibodies, which act as specific antigen receptors (binding of antigen is integral to their activation). B cells also express CD19 and CD20 markers. Activated B cells secrete antibodies and differentiate into antibody producing cells, plasma cells. Thymus a. Formation The first organ of the immune system to form, the thymus reaches its peak size and activity at puberty. The thymus progressively atrophies throughout adult life with only 5-10% of cell mass remaining by age 45-50.

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b.

Thymic atrophy probably contributes to immune senescence, the decline in immune responsiveness with aging. Organization 1) Cortex - predominantly lymphocytes from the bone marrow 2) Medulla - largely epithelial cells and some lymphocytes from the cortex (cells move from the outer cortex into the medulla and then into peripheral circulation as they mature.

IMPORTANT CONCEPT 5: The thymus is critical to normal development of the immune system c. Function - T lymphocyte maturation 1) T cells - lymphocytes characterized by T cell receptors (TCR) and other markers (CD, cluster of differentiation). The most important CD's are designated below. T cells (%) 100 100 ~65 ~35 Comments T cell antigen receptor A complex of peptides closely associated with TCR Defines T cell subpopulation designated as helper cells (Th) Defines T cell subpopulation mediating cytotoxicity (CTL)

Marker TCR CD3 CD4 CD8

2) The thymus modulates T cell maturation through the production of thymic hormones (e.g. thymosin) by thymic epithelial cells. 3) The thymus eliminates self-reactive T cells (immune tolerance). 2. Peripheral Lymphoid Organs (sites of immune stimulation) Lymph nodes a. Structure - 1) Cortex - lymphoid tissue segregated into B and T cell-rich areas. B cell rich lymphoid follicles may develop into germinal centers upon stimulation. T cells are concentrated in paracortical areas. 2) Medulla - a network of sinusoids rich in macrophages. c. Function 1) Filter foreign organisms or antigens from lymph fluid bathing local tissues 2) Major site of immune stimulation. Spleen a. Structure 1) Red pulp - sinusoids populated by macrophages and filled with blood 2) White pulp - lymphoid cells a) B cell-dependent - lymphoid follicles b) T cell-dependent - periarteriolar lymphatic sheath (PALS) b. Function 1) Filter foreign organisms from the blood 2) Major site of immune stimulation Mucosal (Gut) Associated Lymphoid Tissue (MALT or GALT)
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b. V.

General structure - lymphoid follicles or germinal centers (B-dependent) with Tdependent areas in between; covered by specialized mucosal epithelium; no medulla 1) Tonsils - poor lymphatic drainage 2) Peyer's patches - good lymphatic drainage to mesenteric lymph nodes. The mucosal epithelium, particularly over Peyers patches, contains specialized M (microfold) cells. M cells have microfolds, instead of microvilli and actively transport antigenic materials to the lymphoid areas. 3) Appendix - good lymphatic drainage to mesenteric lymph nodes 4) Lamina propria and all other mucous membranes - numerous small foci of lymphoid material. Function - immunity along the gastrointestinal, respiratory, and urogenital tracts.

LYMPHOCYTE RECIRCULATION (HOMING) a. Lymphocytes and phagocytes are formed in the bone marrow and migrate through the blood and lymph to other organs of the immune system. While concentrated in lymphoid tissues, cells continually recirculate. If an infection or inflammation is present, T cells home to the draining lymph nodes and phagocytes migrate to affected tissues. Cell migration or homing is regulated by a family of over 30 adhesion molecules (selectins and integrins) expressed on the membranes of immune cells. The corresponding ligands are expressed on endothelial cells or on extracellular matrix (fibronectin).

IMPORTANT CONCEPT 6: Adhesion molecules mediate the migration of immune cells For example, T cells express the adhesion molecule, L-selectin, which binds to carbohydrates on addressins, such as Gly-CAM-1 (Glycan-bearing Cell Adhesion Molecule-1), on high endothelial venule cells in lymph nodes. This loose binding slows the T cells, causing them to roll along the endothelial cells. Chemotactic factors (IL-8, CXCL8) released at the local site, increases the affinity of the integrin, LFA-1 (Lymphocyte Function Antigen-1), on the T cells for protein on ICAM-1 (InterCellular Adhesion Molecule-1) on the endothelial cells. The combination of adhesion molecules slows the cells rolling enough to allow adhesion. Once firmly attached, the cells can migrate through the endothelial barrier (diapedesis), into the lymph node. Other adhesion molecules direct T cell migration to mucosal tissues or to sites of inflammation. Interaction with dendritic cells induces the expression of tissue-specific (skin, mucosal lymphoid tissues, spleen, etc.) adhesion molecules. Inflammation increases the local expression of addressins and ICAM-1 on endothelial cells. READING: The Immune System , 3rd Ed., Ed: P. Parham. Chapter 1 and 2-10 2-16 and 2-20 222.