Chapter 2: Immune Response to Infection

The “maladies” Metchnikoff and the other pioneers of immunology were fighting infections and, for decades, their field was defined in terms of the immune response to infection. We now understand that the immune system is as much a part of everyday human biologic function as the cardiovascular or renal systems. In its adaptive and disordered states, infectious diseases are only one of the major player along with cancer and autoimmune diseases. Students of medicine take up immunology as a separate unit with its own text covering the field broadly. This chapter is not intended to fulfill that function, or to be a shortened but comprehensive version of those sources. It is included as an overview of aspects related to infection for other students and as an internal reference for topics that reappear in later pages of this book. These include some of the greatest successes of medical science. The early and continuing development of vaccines that prevent and potentially eliminate diseases is but one example. In addition, knowledge of the immune response to infection is integral to understanding the pathogenesis of infectious diseases. It turns out that one of the main attributes of a successful pathogen is evading or confounding the immune system.

The immune response to infection is presented as two major components—innate immunity and adaptive immunity. The primary effectors of both are cells that are part of the white blood cell series derived from hematopoietic stem cells in the bone marrow ­(Figure 2–1). Innate immunity includes the role of physical, cellular, and chemical systems that are in place and that respond to all aspects of foreignness. These include mucosal barriers, phagocytic cells, and the action of circulating glycoproteins such as complement. The adaptive side is sometimes called specific immunity because it has the ability to develop new responses that are highly specific to molecular components of infectious agents, called antigens. These encounters trigger the development of new cellular responses and production of circulating antibody, which have a component of memory if the invader returns. Artificially creating this memory is, of course, the goal of vaccines.

Innate immunity acts through a series of specific and nonspecific mechanisms, all working to create a series of hurdles for the pathogen to navigate (Table 2–1). The first are mechanical barriers such as the tough multilayered skin or the softer but fused mucosal layers of internal surfaces. As discussed in Chapter 1, the microbiota on these surfaces presents formidable competition for space and nutrients. Turbulent movement of the mucosal surfaces and enzymes or acid secreted on their surface make it difficult for an organism to persist. Organisms that are able to pass the mucosa encounter a population of cells with the ability to engulf and destroy them. In addition, body fluids contain chemical agents such as complement, which can directly injure the microbe. The entire process has cross-links to the adaptive immune system. The endpoint of phagocytosis and digestion in a macrophage is the presentation of the antigen on its surface; the first step in specific immune recognition.

PHYSICAL BARRIERS

The thick layers of the skin containing insoluble keratins present the most formidable ­barrier to infection. The mucosal membranes of the alimentary and urogenital tract are not as tough but, often, are bathed in secretions inhospitable to invaders. Lysozyme is an enzyme that digests peptidoglycan—a unique structural component of the bacterial cell wall. Lysozyme is secreted onto many surfaces and is particularly concentrated in conjunctival tears. The acid pH of the vagina and particularly the stomach makes colonization difficult for most organisms. Only small particles (5-10 μm) can be inhaled deep into the lung alveoli because the lining of the respiratory tract includes cilia that trap and move them back toward the pharynx.

The skin and mucosal surfaces of the intestinal and respiratory tract also contain concentrations of lymphoid tissue within or just below their surfaces, which provide a next-level defense for invaders surviving the above described barriers. These lymphoid collections are designed to entrap and deliver invaders to some of the phagocytes described in the following text. For example, in the intestine, M cells (Figure 2–2) that lack the villous brush border of their neighbors endocytose bacteria and then release them into a pocket containing macrophages and lymphocytic components (T and B cells) of the adaptive immune system. The enteric pathogen Shigella exploits this receptiveness of the M cell to attack the adjacent enterocytes from the side.

IMMUNORESPONSIVE CELLS AND ORGANS

Not all the cells shown in Figure 2–1 are involved in the immune system; of those that are, not all respond to infection. What the immunoresponsive cells have in common is derivation from hematopoietic stem cells in the bone marrow, which create the myeloid and lymphoid series followed by further differentiation into their mature cell types. Of the types shown, the erythroblast and megakaryocyte do not participate in immune reactions. In the myeloid series, basophils and mast cells are primarily involved in allergic reactions rather than infection. The immunoresponsive cells are found throughout the body in the circulation or at fixed locations in tissues. They are concentrated in the lymph nodes and spleen, and form a unified filtration network designed as a sentinel warning system. In the lymphoid series, cells destined to become T cells mature in the thymus (the source of their name). Thus, the thymus, spleen, and lymph nodes might be thought of as the organs of the immune system. These are collectively referred to as the lymphoid tissues.

Cellular Receptors for Microbes

Fixed and circulating phagocytes express surface receptors which recognize a limited array of uniquely microbial structures based on the pattern of their molecular structure. These pathogen-associated molecular patterns (PAMPs) include bacterial cell wall peptidoglycan, the lipopolysaccharide endotoxin of gram-negative bacteria, mannose and other glycoproteins, lipids and polysaccharides. Nucleic acids are also recognized such as the double-stranded RNA found in many viruses. These PAMP-recognizing receptors may be found on the surface of phagocytes, dendritic cells, and specialized compartments called toll-like receptors (TLRs) of which over 10 types have been described. The engagement of TLR receptors generate transcription signals that generate a range of antimicrobial cytokines specific to the TLR type.

Antimicrobial Peptides

Antimicrobial peptides (AMPs) are small peptide molecules with natural antimicrobial effects. In mammals there are two major families of AMPs called cathelicidins and defensins.­ They are produced by multiple cell types including leukocytes, mast cells, dendritic cells, and platelets in response to tissue damage. They exhibit broad-spectrum activity against bacteria, fungi, parasites, and some enveloped viruses. Their antimicrobial action is by electrostatic interaction with the surfaces of microbes such as bacterial outer membranes, cytoplasmic membranes, and cell walls causing rupture and death. Some AMPs may also disrupt metabolic processes like nucleic acid and protein synthesis.

Cells Responding to Infection

Monocytes

Monocyte is a general morphologic term for cells that include or quickly (within hours) differentiate into macrophages or dendritic cells. These are the cells of the immune system that both phagocytose invaders and process them for presentation to the adaptive immune system. Macrophages are found in the circulation and tissues, where they are sometimes given regional names such as alveolar macrophage. They possess surface receptors such as mannose and fructose, which nonspecifically recognize components commonly found on pathogens and more specialized receptors able to recognize unique components of microbes such as the lipopolysaccharide (LPS) of gram-negative bacteria. They also have receptors that recognize antibody and complement.

Dendritic cells have a distinctive star-like morphology, and are present in the skin and in the mucous membranes of the respiratory and intestinal tracts. Similar to macrophages, they phagocytose and present foreign antigens. Surface recognition includes PAMP recognition. After binding and phagocytosis, dendritic cells migrate to lymphoid tissues where specific adaptive immune responses are triggered. This interaction involving lymphocytes and T cells functions as a bridge between the innate and adaptive immune systems.

Granulocytes

Of the cells in the granulocyte series, the most active is the polymorphonuclear ­neutrophil or PMN. These cells have a distinctive multilobed nucleus and cytoplasmic granules that contain lytic enzymes and antimicrobial substances including peroxidase, lysozyme, defensins, collagenase, and cathelicidins. PMNs have surface receptors for antibody and complement and are active phagocytes. In addition to the digestive enzymes, PMNs have other oxygen-dependent and oxygen-independent pathways for killing microorganisms. Unlike macrophages, they only circulate and are not present in tissues except by migration as part of an acute inflammatory response.

Eosinophils are nonphagocytic cells that participate in allergic reactions along with basophils and mast cells. Eosinophils are also involved in the defense against infectious parasites by releasing peptides and oxygen intermediates into the extracellular fluid. It is felt that these products damage membranes of the parasite.

Lymphocytes

Lymphocytes are the primary effector cells of the adaptive immune system. They are produced from a lymphocyte stem cell in the bone marrow and leave in a static state marked to become T, B, or null cells after further differentiation (Figure 2–3). This requires activation mediated by surface binding, which then stimulates further replication and differentiation.

B cells mature in the bone marrow and then circulate in the blood to lymphoid organs. At these sites, they may become activated to a form called a plasma cell, which produces antibodies. T cells mature in the thymus and then circulate awaiting activation. Their activation results in production of cytokines, which are effector molecules for multiple immunocytes and somatic cells. Some of the uncommitted null cells become natural killer (NK) cells, which have the capacity to directly kill cells infected with viruses by secreting IFN-γ.

Phagocytosis

Phagocytosis is one of the most important defenses against microbial invaders (Figure 2–4). The major cells involved are PMNs, macrophages, and dendritic cells. For all, the process begins with surface–pathogen recognition mechanisms, which may be either dependent on opsonization of the organism with complement or antibody or independent of opsonization. At this point, only the opsonin-independent mechanisms are considered. These use the nonspecific mechanisms already described and hydrophobic interactions between bacteria and the phagocyte surface. More powerful mechanisms include lectins, which bind carbohydrate moieties and protein–protein interactions based on a specific peptide sequence (arginine-glycine-aspartic-acid or RGD). These RGD receptors are present on virtually all phagocytes.

Bound organisms are taken inside the phagocyte in a membrane-bound phagosome destined to fuse with lysosomes inside to form a phagolysosome. This is the main killing ground of the phagocyte. The lysosomal enzymes include hydrolases and proteases that have maximum activity at the acidic pH inside the phagolysosome. In addition, inside the phagocyte are oxidative killing mechanisms created by enzymes that produce reactive oxygen intermediates (superoxide, hydrogen peroxide, singlet oxygen) driven by a metabolic respiratory burst in the cell cytoplasm. These mechanisms are particularly used for killing bacteria. Bacterial pathogens whose pathogenesis involves multiplication rather than destruction inside the phagocyte have mechanisms to block one or more of the preceding steps. For example, some pathogens are able to block fusion of the phagosome with the lysosome; others interfere with the acidification of the phagolysosome.

Another mechanism effective with some viruses, fungi, and parasites is the formation of reactive nitrogen intermediates (nitric oxide, nitrate, and nitrite) delivered into a vacuole or in the cytoplasm. PMN granules contain a variety of other antimicrobial substances, including peptides called defensins. Defensins act by permeabilizing membranes and, in addition to bacteria, are active against enveloped viruses.

INFLAMMATION

Inflammation encompasses a series of events in which the above mentioned cells are deployed in response to an injury—such as a new microbial invader. At the first insult, chemical signals mobilize cells, fluids, and other mediators to the site to contain, combat, and heal. In acute inflammation, the first events may be noticed in minutes, and the entire process resolved over a matter of days to a couple of weeks. Chronic inflammation may follow the incomplete resolution of an acute process or arise as a slow insidious process of its own. The natural history of infections such as tuberculosis, which follow this pattern, run for months, years, even decades.

The first event in acute inflammation is the release of chemical signals (chemokines) that act on adhesion molecules (selectins) in local capillaries. This slows the movement of passing PMNs and activates adhesive integrins on their surface. This leads to tight adhesion to the endothelium followed by squeezing past the endothelial wall to the tissues below. There, chemotactic factors released by the bacteria lead them to the primary site. Increasing acidity of local fluids releases enzymes (kallikrein, bradykinin) that open junctions in capillary walls and allow increased flow of fluids and more leukocytes. Histamine (from mast cells), arachidonic acid, and prostaglandin release complete the picture of swelling and pain.

Chronic inflammation bridges the innate and adaptive immune responses. An acute phase, if present, is usually not noticed, and the cellular infiltrate is composed of lymphocytes and macrophages with relatively few PMNs. It is generally associated with slower-growing pathogens such as mycobacteria, fungi, and parasites in which cell-mediated immunity is the primary adaptive defense. Many of these pathogens have mechanisms that allow them to multiply in nonactivated macrophages. If the macrophages are effectively activated by T cells, the multiplication ceases and the inflammation and injury are minimal. If not, multiplication and chronic inflammation continue sometimes in the form of a granuloma, which is an indication of a destructive hypersensitivity component to the inflammation.

CHEMICAL MEDIATORS

Chemical mediators of innate immunity that have direct antimicrobial activity include cationic proteins and complement. The cationic proteins (cathelicidin, defensins) act on bacterial plasma membranes by the formation of ionic pores, which alter membrane ­permeability. The complement system is a series of glycoproteins, which can directly insert in bacterial membranes or act as receptors for antibody. Cytokines are proteins or glycoproteins released by one cell population that act as signaling molecules for another. They are generally thought of in the context of the adaptive immune system, but they can be stimulated directly by microorganisms.

The Complement System

The complement system consists of more than 30 distinct components and several other precursors. All are in the plasma of healthy individuals in inactive forms that must be enzymatically cleaved to become active. When this happens, a cascade of reactions is triggered, which activates the various components in a fixed sequence (Figure 2–5). The difference between the pathways is in the mechanisms for their initiation. Once started, any pathway can produce the same effects on pathogens, which include enhancing phagocytosis, activation of leukocytes, and lysis of bacterial cell walls. An important step in the process is coating of the organism with serum components, a process called opsonization. The coatings may be mannose-binding proteins, complement components, or antibody. There is no immunologic specificity in complement activation or in its effects.

Alternative Pathway

The alternative pathway is activated by bacterial cell wall components with repetitive surface structures such as LPS. The multiple components come together in the formation of the membrane-attack complex, which inserts directly into bacterial membranes (Figure 2–6), particularly the outer membrane of gram-negative bacteria. This not only injures the organism, but also enhances phagocytosis because the other end of the molecule has receptors for phagocytes. Gram-positive bacteria are less affected because they have no exposed membrane (see Chapter 21). These actions are particularly important for the effectiveness of innate immunity in the early stages of acute infections before the adaptive immune system has time to act. The key complement component for alternate pathway activity is C3b. C3b activation and degradation are regulated by a number of serum factors (factors B, D, and H) that can modulate its activity. A major mechanism for pathogens to block alternate pathway attack is by binding factor H to their surface. This is accomplished by bacterial capsules and surface proteins. This concentration of factor H causes local degradation of C3b (see Chapter 22, Figure 22–4).

Lectin Pathway

Another means of activating the complement system is based on the carbohydrate building of lectins. In this case, the lectins bind to mannose—a common surface component of bacteria, fungi, and some virus envelopes. This binding opsonizes the pathogen and enhances phagocytosis. Thus, as in the alternative pathway, the activation comes from pathogen surfaces and proceeds through the same C3 convertase (Figure 2–5).

Classic Pathway

The classic complement pathway is initiated by the binding of antibodies formed during the adaptive immune response (as described further) with their specific antigens on the surface of a pathogen. This binding is highly specific but amounts to another case of opsonization activating the complement cascade. In this case, specific sites on the Fc portion of immunoglobulin molecules bind and activate the C1 component of complement to start the process. The pathway and sequence of individual complements are characteristic of the classic pathway, but it still reaches C3b, the common point for microbial directed action. As with the alternative pathway, this creates the membrane-attack complex, the mediators of inflammation, and receptors for phagocytes on C3b.

Cytokines

Cytokine is a broad term referring to molecules released from one cell population destined to have an effect on another cell population (Table 2–2). As these proteins and glycoproteins have been discovered, they have been named and classified in relation to biologic effects observed initially only to discover that they have multiple other actions. For infectious diseases, the operative subcategories are chemokines, which are cytokines chemotactic for inflammatory cell migration, and interleukins (IL-1, -2, -3, etc), which regulate growth and differentiation between monocytes and lymphocytes. Tumor necrosis factor (TNF), so named for its cytotoxic effect on tumor cells, can also induce apoptosis (programmed cell death) in phagocytes—a useful feature for pathogens they have taken in. Interferons (INF-α, -β, and -γ) were originally named for their interference with viral replication ­(Figure 2–7), but are now known to be central to activation of T cells and macrophages. Unless commanded to understand specific situations, cytokine is used to represent all these mediators in these pages.

The adaptive immune system differs from the innate immune response in its discrimination between self and nonself and in the magnitude and diversity of highly specific immune responses possible (Table 2–3). In addition, it has a memory function, which is able to mount an accelerated response if an invader returns. The adaptive system operates in two broad arms—humoral immunity and cell-mediated immunity. Humoral immunity comes from bone marrow-derived B cells and acts through the ability of the antibodies it produces to bind foreign molecules called antigens. Cell-mediated (cellular) immunity is mediated through T cells that mature in the thymus and respond to antigens by directly attacking infected cells or by secreting cytokines to activate other cells. As shown in ­Figure 2–8, the B-cell and T-cell systems are interactive.

Antigens and Epitopes

An antigen is any substance (usually foreign) with the ability to stimulate an immune response when presented in an effective fashion. They are usually large structurally complex proteins, polysaccharides, or glycolipids. Each antigen can contain many subregions that are the actual antigenic determinants, or epitopes. These epitopes can consist of separate peptides, carbohydrates, or lipids of the correct size and three-dimensional configuration to fit the combining site of an antibody molecule or a T-cell receptor (TCR) (Figure 2–9). Approximately six amino acids or monosaccharide units provide a correctly sized epitope. Antigens presented by infectious agents typically contain multiple epitopes, including copies of the same epitope. Other small organic molecules that would not ordinarily stimulate an immune response may do so if bound to a larger carrier, such as a protein. These are called haptens, and the specificity of the immune response may be generated for both the hapten and its larger carrier.

A foreign antigen entering a human host may, by chance, encounter a B cell whose surface antibody is able to bind it. This interaction stimulates the B cell to multiply, differentiate, and produce more surface and soluble antibodies of the same specificity. Eventually, the process leads to production of enough antibody to bind more of the antigen. This mechanism is most likely to operate with antigens such as polysaccharides that have repeating subunits, thus improving the possibility that exposed epitopes are recognized.

Large, complex antigens such as proteins and viruses must be processed before their epitopes can be effectively recognized by the immune system. This processing takes place in macrophages or specialized epithelial cells found in the skin and lymphoid organs, where they are adjacent to other immunoresponsive cells. The ingested antigen is degraded to peptides of 10 to 20 amino acids that are presented by major histocompatibility molecules on the host cell surface to be recognized by T cells (Figures 2–10, 2–11).

Recognition of Foreignness

Distinguishing between self and nonself is obviously essential to maintaining integrity and homeostasis. The collection of genes that control these functions is called the major histocompatibility complex (MHC), and it codes for molecules present on the surface of almost all human cells. Of interest in infection are MHC class I and II molecules (Figure 2–10). MHC class I molecules are in the membrane of almost all cells, but MHC class II molecules are present only on certain leukocytes such as macrophages, dendritic cells, and some T and B cells.

Both MHC class I and class II participate in antigen processing, but by distinctly different pathways (Figure 2–11). MHC class I molecules bind to products generated in the cytoplasm by a natural process or a viral infection. Viral proteins are digested to peptides in a cytoplasmic structure called the proteasome, and delivered to the endoplasmic reticulum. Here they find the binding site of the class I molecule and are transported to the surface for presentation of the peptide. MHC class II molecules bind to fragments that originally come from outside the cell, but have been taken into the endocytotic vacuole of a phagocyte. After digestion in the phagolysosome, peptide fragments are combined with class II molecules and move to the surface for presentation. The presented MHC class I peptides are recognized by CD8+ T cells and the MHC class II by CD4+ T cells.

THE T-CELL RESPONSE

T cells originate in the bone marrow and migrate to the thymus for differentiation. Those that recognize self are destroyed. Those that survive are mature but still to be activated. T cells have specific TCRs on their surface, with binding sites extending to the outside ­(Figure 2–12). The two major types of T cells are helper T (CD4+) and cytotoxic (CD8+) T cells. The major roles of T cells in the immune response are as follows:

1. Recognition of peptide epitopes presented by MHC molecules on cell surfaces. This is followed by activation and clonal expansion of T cells in the case of epitopes associated with class II MHC molecules.

2. Production of cytokines that act as intercellular signals and mediate the activation and modulation of various aspects of the immune response and of nonspecific host defenses.

3. Direct killing of foreign cells, of host cells bearing foreign surface antigens along with class I MHC molecules (eg, some virally infected cells), and of some immunologically recognized tumor cells.

CD4+ Helper T Lymphocytes

Helper T cells are stimulated by antigen in the context of MHC class II presentation and are further marked by the presence of the CD4 cell surface antigen. If T cells are of the proper MHC background to recognize the antigen specifically, T-cell activation occurs. The antigen–MHC complex presented to a specific T cell by the macrophage is the specific signal that induces the T cell to become activated and divide. At this point, the T helper (Th) cells differentiate into three major subsets of effector cells each with characteristic cytokines, target cells, and typical microbial pathogen profiles. Th1 cells produce IFN-γ, target macrophages, and are effective against intracellular pathogens like Mycobacterium tuberculosis. Th2 cells produce multiple interleukins, and promote IgE, mast cell, and eosinophil-mediated destruction of parasites. Th17 cells also produce interleukins including IL-17, stimulate neutrophils, and are active against extracellular bacterial and fungal pathogens.

CD8+ Cytotoxic T Lymphocytes

CD8+ cytotoxic T lymphocytes (CTLs) are a second class of effector T cells. They are lethal to cells expressing the epitope against which they are directed when the epitope is presented by class I MHC molecules. They too have specific epitope recognition sites, but they are characterized by the CD8 cell surface marker; thus, they are referred to as CD8+ cytotoxic T cells. These cells recognize the association of antigenic epitopes with class I MHC molecules on a wide variety of cells of the body. In the case of virally infected cells, cytotoxic CD8+ cells prevent viral production and release by eliminating the host cell before viral synthesis or assembly is complete (Figure 2–13). The destruction of the virally infected cell is accomplished through a complement-like action mediated by perforins, which also facilitates entry into the cell of enzymes (granzymes) that activate apoptosis.

Superantigens

A group of antigens have been termed superantigens because they stimulate a much larger number of T cells than would be predicted based on the specificity of combining site ­diversity. This causes a massive cytokine release. The action of superantigens is based on their ability to bind directly to MHC proteins and to particular Vβ regions of the TCR without involving the antigen-combining site. Individual superantigens recognize exposed portions defined by framework residues that are common to the structure of one or more Vβ regions. Any T cells bearing those Vβ sites may be directly stimulated. A variety of microbial products have been identified as superantigens. Superantigens are discussed further in Chapter 22 (see Figure 22–7) and in Chapters 24 and 25, describing their role in toxic shock syndromes caused by Staphylococcus aureus and group A streptococci.

Cell-Mediated Immunity

In the control of infection, cell-mediated immunity is most important in the response to obligate or facultative intracellular pathogens. These include some slow-growing bacteria, such as the mycobacteria and fungi against which antibody responses appear to be ineffective. The mechanisms are complex and involve a number of cytokines with amplifying feedback mechanisms for their production. After the initial processing of antigen to stimulate activation of the antigen-recognizing CD4+ T cell, cytokine feedback from the CD4+ T cells to macrophages further increases their clonal expansion (including memory cells) and activates CD8+ (cytotoxic) T lymphocytes. Other cytokines from CD4+ T cells attract macrophages to the site of infection, hold them there, and activate them to greatly enhance microbiocidal activity. The sum of the individual and collaborative activities of T cells, macrophages, and their products is a progressive mobilization of a range of host defenses to the site of infection and greatly enhanced macrophage activity. In the case of tuberculosis, IFN-γ inhibits the replication of the mycobacteria inside macrophages. In viral infections, CD8+ cytotoxic lymphocytes destroy their cellular habitat leaving already assembled virions accessible to circulating antibody.

B CELLS AND ANTIBODY RESPONSES

B lymphocytes are the cells responsible for antibody responses. They develop from precursor cells in the bone marrow before migrating to other lymphoid tissues. Each mature cell of this series carries a specific epitope recognition site on its surface. This B-cell receptor is actually a monomer of one form of antibody (IgM) oriented with its binding sites facing outward. Upon binding antigen, the receptor-antigen complex is internalized for initiation of antibody production by the stimulated B cell. In this process, the B lymphocytes multiply, differentiating into either memory or plasma cells. Plasma cells are end cells adapted for secretion of large amounts of antibodies. In addition to their essential role in antibody production, B cells can present antigen to T cells.

There are two broad types of antigen triggering: T-dependent and T-independent. T-dependent reactions are those that use collaboration between helper T cells and B cells to initiate the process of antibody production. This is the mechanism evoked by proteins and haptens bound to proteins. The response is strong and includes memory cells, therefore, it can be boosted in the case of immunization.

T-independent responses are those that do not require help by T cells to stimulate B-cell antibody production. It is evoked by large molecules with many repeating units such as polysaccharides which cannot bind to MHC molecules. At first glance, this independence may seem to be an advantage, but T-independent responses are not the same as T-dependent responses. The antibody generally has a lower affinity for its antigen and a shorter duration in circulation. Memory cells are not produced, and T-independent responses mature more slowly than T-dependent responses. This delay in maturation may contribute to the increased susceptibility to some bacterial infections in early life. It certainly contributed to the failure of purified polysaccharide vaccines to effectively immunize children younger than 2 years. For use in children, these vaccines have been replaced with a hapten approach in which the polysaccharide is conjugated to protein. In this form, antibody generated by the T-dependent mechanism (protein carrier) still has specificity for the polysaccharide epitopes.

After challenge with foreign antigen, there is a lag period of 4 to 6 days before antibody can be detected in serum. This period reflects the events involved in the recognition of the antigen, its processing, and the specific activation of the cells of the immune system. The first event is the clearance of antigen from the circulation by what is essentially a metabolic process in which the antigen is recognized in a nonspecific sense and ingested. The vast preponderance of antigen ends up in circulating phagocytes or in stationary macrophages. The macrophages process the antigen; therefore, those immunogenic moieties can be presented to T cells, which then cause the B cells to produce immunoglobulins. The antibody-forming system is a learning system that responds to challenge by foreign molecules by producing large amounts of specific antibody. In addition, the affinity of its binding to the specifically recognized antigen often increases with time or secondary challenge.

Antibody Structure

Antibodies belong to the immunoglobulin family of proteins, which appear in quantity in serum and on the surfaces of B cells. Of the five known structural types, three (IgG, IgM, and IgA) are involved in the defense against infection. The basic structure of an immunoglobulin is illustrated in Figure 2–14, which depicts an IgG molecule. Immunoglobulins have a basic tetrameric structure consisting of two light polypeptide chains and two heavy chains usually associated as light/heavy pairs by disulfide bonds. The two light/heavy pairs are covalently associated by disulfide bonds to form the tetramer. There are two types of light chains, κ and λ, which are the products of distinct genetic loci. The class or isotype of the immunoglobulin is defined by the type of heavy chain expressed.

The Y-shaped structure includes two antigen binding sites (Fab) formed by interaction of the variable domains of the heavy chain and the light chain. The stalk is called the Fc fragment. Antibodies carry out two broad sets of functions: the recognition function is the property of the Fab sites for antigen, and the effector functions are mediated by the constant regions of the heavy chains. Variations in the hypervariable region of the Fab-combining site due to mutations are called idiotypes. Antibodies combine with foreign antigens, but the actual destruction or removal of antigen requires the interaction of portions of the Fc fragment with other molecules such as complement components and phagocytes which have Fc receptors.

Figure 2–15 shows a schematic representation of a serum IgM immunoglobulin. This molecule consists of five subunits of the typical IgG molecule. The molecule occurs as a cyclic pentamer, and a J (joining) chain links the intact structure. When IgM is present on the surface of B cells where it serves as a primary receptor for antigen, it is present as a monomer. Other immunoglobulins showing a difference in arrangement from the typical IgG model are the IgA immunoglobulins. In serum, these immunoglobulins can occur as a monomer, but they can also occur in dimers in which the joining chain is required to stabilize the dimer. IgA molecules in the gut occur as dimers in which both the J chain and an additional polypeptide, termed the secretory component, are present in the complex.

Functional Properties of Immunoglobulins

Immunoglobulin G

Immunoglobulin G (IgG) is the most abundant immunoglobulin in health and provides the most extensive and long-lived antibody response to the various microbial and other antigens that are encountered throughout life. Although at least four subclasses of IgG have been characterized, they are grouped together for the purpose of this chapter. The IgG molecule is bivalent with two identical and specific combining sites. The Fc region does not vary with differences in specificity of combining sites of different antibody molecules. The Fc fragment binding sites for phagocytic cells are made available when the variable region of the antibody molecule has reacted with specific antigen, leaving the Fc facing outward.

IgG antibody is characteristically formed in large amounts during the secondary response to an antigenic stimulus, and usually follows production of IgM (see Immunoglobulin M) in the course of a viral or bacterial infection. Memory cells are programmed for rapid IgG response when another antigenic stimulus of the same type occurs later. Immunoglobulin G antibodies are the most significant antibody class for neutralizing bacterial exotoxins and viruses often by blocking their attachment to cell receptors. Accelerated IgG responses from memory cell expansion frequently confer lifelong immunity when directed against microbial antigens that are determinants of virulence. IgG is the only immunoglobulin class able to cross the placental barrier and, thus, provides passive immune protection to the newborn in the form of maternal antibody.

Immunoglobulin M

Monomers of immunoglobulin M (IgM) constitute the specific epitope recognition sites on B cells that ultimately give rise to plasma cells producing one or another of the different immunoglobulin classes of antibody. Because of its many specific combining sites, IgM is particularly effective in agglutinating particles carrying epitopes against which it is directed. It also contains many sites for binding the first component of complement. These sites become available once the IgM molecule has reacted with antigen. IgM is particularly active in bringing about complement-mediated cytolytic damage to foreign antigen-bearing cells. It is less effective as an opsonizing antibody because its Fc portion is not available to phagocytes.

Immunoglobulin A

Immunoglobulin A (IgA) has a special role as a major determinant of so-called local immunity in protecting epithelial surfaces from colonization and infection. Certain B cells in lymphoid tissues adjacent to, or draining surface epithelia of the intestines, respiratory tract, and genitourinary tract, are encoded for specific IgA production. After antigenic stimulus, the clone expands locally, and some of the IgA-producing cells also migrate to other viscera and secretory glands. At the epithelia, two IgA molecules ­combine with another protein, termed the secretory piece, which is present on the surface of local epithelial cells. The complex, then termed secretory IgA (sIgA), passes through the cells into the mucous layer on the epithelial surface or into glandular secretions, where it exerts its protective effect. The secretory piece not only mediates secretion, but also protects the molecule against proteolysis by enzymes such as those present in the intestinal tract.

The major role of sIgA is to prevent attachment of antigen-carrying particles to receptors on mucous membrane epithelia. Thus, in the case of bacteria and viruses, it reacts with surface antigens that mediate adhesion and colonization and prevents the establishment of local infection or invasion of the subepithelial tissues. sIgA can agglutinate particles, but has no Fc domain for activating the classic complement pathway; however, it can activate the alternative pathway. Reaction of IgA with antigen within the mucous membrane initiates an inflammatory reaction that helps mobilize other immunoglobulin and cellular defenses to the site of invasion. IgA response to an antigen is shorter lived than the IgG response.

Antibody Production

The major events characterizing the time course of antibody production are illustrated in Figure 2–16 and summarized as follows: Initial contact with a new antigen evokes the primary response, which is characterized by a lag phase of approximately 1 week between the challenge and the detection of circulating antibodies. In general, the length of the lag phase depends on the immunogenicity of the stimulating antigen and the sensitivity of the detection system for the antibodies produced. Once antibody is detected in serum, the levels rise exponentially to attain a maximal steady state in approximately 3 weeks. These levels then decline gradually with time if no further antigenic stimulation is given. The first antibodies synthesized in the primary immune response are IgM and, then in the latter phase, IgG antibodies arise and eventually predominate. This transition is termed the IgM/IgG switch.

After a subsequent exposure or booster injection of the same antigen, a different sequence called the secondary response or anamnestic response ensues. This response involves memory. In the secondary response, the lag time between the immunization and the appearance of antibody is shortened, the rate of exponential increase to the maximum steady-state level is more rapid, and the steady-state level itself is higher, representing a larger amount of antibody. Another key factor of the secondary response is that the antibodies formed are predominantly of the IgG class. In addition to higher levels, the secondary IgG antibodies have a higher affinity for their antigen. Figure 2–16 shows the participation of memory T cells created during the primary response in these reactions.

The immune system is no different from any other human system. In balance, we do not even know it is there, but in an exaggerated state we call hypersensitivity, it can cause injury and even chronic disease. Hypersensitivity reactions have been placed into four classes on the basis of their mechanism of immunologic injury. Type I or allergic reactions relate to the action of IgE and the release of powerful mediators, such as histamine from mast cells. Type II or cytotoxic reactions are created when IgG or IgM antibodies are misdirected to host cells. Type III or immune complex reactions are created when an excess of antigen–antibody complexes are deposited and followed by complement-mediated inflammation. Type IV reactions are cell-mediated and often called delayed-type hypersensitivity because of the time delay in invoking the T_H1 response. The hypersensitivity diseases include allergy, anaphylaxis, asthma, transfusion reactions, rheumatoid arthritis, and type 1 diabetes. Infectious diseases are a relatively small part of this spectrum, but involve three of the four mechanisms (II, III, and IV).

ANTIBODY-MEDIATED (TYPE II) HYPERSENSITIVITY

Type II hypersensitivity is antibody-dependent cytotoxicity that occurs when antibody binds to antigens on host cells, leading to phagocytosis, cytotoxic T-cell activity, or complement-mediated lysis. The cells to which the antibody is specifically bound, as well as the surrounding tissues, are damaged because of the inflammatory amplification. In the best understood situations related to infection, the mechanism of antibody stimulation is molecular mimicry. That is, the antibody stimulated by an epitope on the pathogen, unfortunately, also binds to a similar epitope on host cells. In rheumatic fever, the infectious epitope is in a surface protein of the group A streptococcus and the host epitope in the myocardium of the heart (see Chapter 25). The streptococcal protein and cardiac myosin share similar amino acid sequences; therefore, it is a cross-reaction. The result is acute myocarditis.

IMMUNE COMPLEX (TYPE III) HYPERSENSITIVITY

When IgG is mixed in appropriate proportions with multivalent antigen molecules (ie, bearing multiple epitopes), aggregates of many antigen and antibody molecules may form. These antigen–antibody complexes can occur in infection when sufficient amounts of specific antibody and free antigen from an infecting microorganism combine to form an immune complex. These complexes are usually removed by cells of the monocyte-macrophage system, but, in excess, can circulate and become deposited in blood vessels, kidney, or joints. When deposited, they bind complement and stimulate an inflammatory reaction that may injure the local tissue. This is felt to be the mechanism of poststreptococcal acute glomerulonephritis (see Chapter 25), and is suspected to be responsible for some of the manifestations when microorganisms circulate in the bloodstream.

In the past, an immune complex disease called serum sickness used to follow the infusion of antibodies (antisera) produced in horses to combat infection. Human antibody to the foreign horse immunoglobulin was formed. These diseases (diphtheria, tetanus) are now prevented by vaccines that stimulate antibody against the same epitopes in humans. When passive immunization is used, human sources of antibody are now available.

DELAYED-TYPE (TYPE IV) HYPERSENSITIVITY

Type IV delayed-type hypersensitivity (DTH) is a cell-mediated immune reaction. The delay is the time required after initiation of a T_H1 response for antigen to be processed, cytokines produced, and T cells to migrate and accumulate at the antigen site. At the site, cytotoxic T cells, macrophages, and other inflammatory mediators directed at cells containing the antigen also produce injury in the surrounding tissue. The purest form of DTH is the intradermal skin test for tuberculosis. In persons already sensitized to the antigens of M tuberculosis, it takes 1 to 2 days for induration to be produced at the site of inoculation of a standardized antigen called tuberculin. This is a useful diagnostic test, but, in infectious disease, DTH is also the hypersensitivity mechanism that causes the most injury. This occurs in diseases in which immunity is cell-mediated with little or no effective antibody component. If these responses are successful in containing the infection at an early stage, there is little destruction. If they are not successful enough to contain growth of the pathogen, increasing amounts of antigen stimulate continuing DTH-mediated destructive inflammation. This is the primary mechanism of injury in tuberculosis, fungal infections, and many parasitic diseases.

NATURAL IMMUNITY TO INFECTION

The majority of encounters with microorganisms including pathogens end favorably for the host. The heightened immunologic responses following infection usually provide immunity, often for life. This is called natural immunity. In some instances, the gauntlet is long because a pathogen of the same name may exist in multiple antigenic types. Because of the specificity of the adaptive immune response, immunity must be developed individually to each antigenic type. Development of natural immunity need not require a clinical infection. There is ample evidence from population studies that individuals with no history or recollection of infection have evidence of immunity in the form of specific antibody. From the time of birth forward, we have many encounters with infectious agents, most of which lead to immunity without disease.

PASSIVE IMMUNITY

Passive immunity is the transfer of antibodies from one person to another. Because the antibody was not made by the recipient, this antibody is transient and lasts only a few weeks or months. This is a natural process in the case of IgG transferred transplacentally from mother to fetus. The protection provided by this antibody is limited to the immunologic experience of the mother, but covers a particularly vulnerable time in life, lasting as long as 6 months after birth. Passive immunity can also be provided as a therapeutic product in which specific antibodies are infused. Such antisera are available for only a limited number of diseases such as rabies, botulism, and tetanus.

VACCINES

Vaccines artificially stimulate immunity through exposure to an antigenic substance. The early vaccines such as Jenner’s for smallpox and Pasteur’s for anthrax (in animals) were live attenuated strains with the ability to produce a true, if mild, infection. We later learned how to kill the agent in a way that retained its antigenicity. These killed vaccines are ­practical if the number of antigens present is limited as with a virus (polio) or bacterial toxin (diphtheria), but usually too crude if whole bacteria are used. Progress with killed bacterial vaccines required knowledge of just which antigenic component provides protective immunity. This allowed inactivation followed by purification of the selected component. This approach with bacterial polysaccharide capsules has produced a dramatic reduction (> 95%) in childhood meningitis. Genomic approaches are now aimed at producing a protective antigen without growth of the organism itself. For each of the 57 chapters in this book devoted to specific infectious agents, vaccines and the immunologic mechanisms involved are carefully examined.