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Antigen Processing and Presentation

T lymphocytes recognize antigens only in the context of self-MHC molecules on the surface of accessory cells. During processing, intact protein antigens are degraded into peptide fragments. Most epitopes that T cells recognize are peptide chain fragments. B cells and T cells often recognize different epitopes of an antigen leading to both antibody and cell-mediated immune responses. Before antigen can bind to MHC molecules, it must be processed into peptides in the intracellular organelles. CD4+ helper T lymphocytes recognize antigens in the context of class II MHC molecules, a process known as class II MHC restriction. By contrast, CD8+ cytotoxic T lymphocytes recognize antigens in the context of class I molecules, which is known as class I MHC restriction. Following the generation of peptides by proteolytic degradation in antigenpresenting cells, peptide–MHC complexes are presented on the surface of antigen-presenting cells where they may be recognized by T lymphocytes. Antigens derived from either intracellular or extracellular proteins may be processed to produce peptides from either self or foreign proteins that are presented by surface MHC molecules to T cells. In the class II MHC processing pathway, professional antigen-presenting cells such as macrophages, dendritic cells, or B lymphocytes incorporate extracellular proteins into endosomes where they are processed (Figure 5.1 and Figure 5.2). Enzymes within the vesicles of the endosomal pathway cleave proteins in the acidic environment. Class II MHC heterodimeric molecules, united with invariant chain, are shifted to endosomal vesicles from the endoplasmic reticulum. Following cleavage of the invariant chain, DM molecules remove a tiny piece of invariant chain from the MHC molecules’ peptide-binding groove. Following complexing of extracellular-derived peptide with the class II MHC molecule, the MHC–peptide complex is transported to the cell surface where presentation to CD4+ T cells occurs. Proteins in the cytosol, such as those derived from viruses, may be processed through the class I MHC route of antigen presentation. The multiprotein complex in the cytoplasm, known as the proteasome effects, involve proteolytic degradation of proteins in the cytoplasm to yield many of the peptides that are presented by class I MHC molecules. TAP molecules transport peptides from the cytoplasm to the endoplasmic reticulum where they interact and bind to class I MHC dimeric molecules. Once the class I MHC molecules have become

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stabilized through peptide binding, the complex leaves the endoplasmic reticulum entering the Golgi apparatus en route to the surface of the cell. Thus, mechanisms are provided through MHC-restricted antigen presentation to guarantee that peptides derived from extracellular microbial proteins can be presented by class II MHC molecules to CD4+ helper T cells and that peptides derived from intracellular microbes can be presented by class I MHC molecules to CD8+ cytotoxic T lymphocytes. The generation of microbial peptides produced through antigen processing to combine with self MHC molecules is critical to the development of an appropriate immune response. Antigen presentation (Figure 5.3 and Figure 5.4) is the expression of antigen molecules on the surface of a dendritic cell, macrophage, or other antigen-presenting cell in association with MHC class II molecules when the antigen is being presented to a CD4+ T helper lymphocyte or in association with cell surface MHC class I molecules when presentation is to CD8+ cytotoxic T lymphocytes (Figure 4.3). Antigen-presenting cells, known also as accessory cells, include dendritic cells, macrophages, and Langerhans cells of the skin as well as B lymphocytes. Target cells such as fibroblasts present antigen to CD8+ cytotoxic T lymphocytes. Mononuclear phagocytes ingest proteins and split them into peptides in endosomes. These eight to ten amino acid residue peptides link to cell surface MHC class II molecules. For appropriate presentation, it is essential that peptides bind securely to the MHC class II molecules since those that do not bind or are bound only weakly are not presented and fail to elicit an immune response. Following interaction of the presented antigen and MHC class II molecules with the CD4+ helper T cell receptor, the CD4+ lymphocyte is activated, interleukin-2 is released, and IL-2 receptors are expressed on the CT4+ lymphocyte surface. The IL-2 produced by the activated cell stimulates its own receptors as well as those of mononuclear phagocytes, increasing their microbicidal activity. IL-2 also stimulates B cells to synthesize antibodies. Whereas B cells may recognize a protein antigen in its native state, T lymphocytes recognize the peptides that result from antigen processing. Antigen processing is the degradation of proteins into peptides capable of binding to MHC molecules for presentation to T lymphocytes. For presentation by MHC molecules, antigens must be processed into peptides.

An antigenic peptide is a peptide that is able to induce an immune response and one that complexes with major histocompatibility complex (MHC), thereby permitting its recognition by a T cell receptor. The peptide-binding cleft is that part of a major histocompatibility molecule that binds peptides for display to T lymphocytes. Paired α-helices on a floor of an eightstranded β-pleated sheet comprise the cleft. Situated in and around this cleft are polymorphid residues that are the amino acids which differ among various MHC alleles. Anchor residues are amino acid side chains of the peptide whose side chains fit into pockets in the peptide-binding cleft of the MHC molecule. The side chains anchor the peptide in the cleft of the MHC molecule by binding two complementary amino acids in the MHC molecule.

FIGURE 5.1 Capture, processing, and presentation of antigen by an antigen-presenting cell.

CD1 is an antigen that is a cortical thymocyte marker, which disappears at later stages of T cell maturation. The antigen is also found on interdigitating cells, fetal B cells, and Langerhans cells. These chains are associated with β2-microglobulin and the antigen is thus analogous to classical histocompatibility antigens, but coded for by a different chromosome. More recent studies have shown that the molecule is coded for by at least five genes on chromosome 1, of which three produce recognized polypeptide products. CD1 may participate in antigen presentation. Direct antigen presentation refers to cell surface allogeneic MHC molecule presentation by graft cells to T lymphocytes of the graft recipient, leading to T lymphocyte activation. This process does not require processing. Direct recognition of foreign MHC molecules is a crossreaction between a normal TCR that recognizes self-MHC molecules plus foreign antigen and allogeneic MHC molecule–peptide complex. The powerful T cell response to allografts is due in part to direct presentation. A proteasome (Figure 5.6) is a 650-kDa organelle in the cytoplasm termed the low molecular mass polypeptide complex. The proteasome is believed to generate peptides by degradation of proteins in the cytosol. It is a cylindrical structure comprised of as many as 24 protein subunits. The proteasome participates in degradation of proteins in the cytosol that are covalently linked, ubiquinated, prior to presentation to MHC class I-restricted T lymphocytes.

FIGURE 5.2 Processing pathways for class II-restricted antigen presentation.

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FIGURE 5.3 Presentation of MHC histocompatibility antigen HLA-B 270S complexed with nonapeptide ARG-ARG-ILELYS0ALA-ILE-THR-LEU-LYS. The C-terminal amino acid of the antigen-binding domain is protected by a N-methyl group. Three water molecules bridge the binding of the peptide to the histocompatibility protein.

Proteasomes that include MHC gene encoded subunits are especially adept at forming peptides that bind MHC class I molecules. Indirect antigen presentation: In organ or tissue transplantation, the mechanism whereby donor allogeneic MHC molecules are present in microbial proteins. The recipient professional antigen-presenting cells process allogeneic

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MHC proteins. The resulting allogeneic MHC peptides are presented in association with recipient self-MHC molecules to host T lymphocytes. By contrast, recipient T cells recognize unprocessed allogeneic MHC molecules on the surface of the graft cells in direct antigen presentation. The immunological synapse is the nanometer scale gap between a T cell and an antigen-presenting cell, which is

FIGURE 5.4 Antigen presentation.

the site of interaction between T cell antigen receptors and major histocompatibility complex molecule–peptide complexes that initiate adaptive immune responses. Erp57 is a chaperone molecule that participates in the loading of peptide onto MHC class I molecules in the endoplasmic reticulum. Transporter associated with antigen processing (TAP) refers to a TAP-binding heterodimeric protein in the rough endoplasmic reticulum membrane that transports peptides from the cytosol to the endoplasmic reticulum lumen. It is comprised of TAP 1 and TAP 2 subunits that bind peptides to class I MHC molecules. Agrin is an aggregating protein crucial for formation of the neuromuscular junction. It is also expressed in lymphocytes and is important in reorganization of membrane lipid microdomains in setting the threshold for T cell signaling. T cell activation depends on a primary signal delivered through the T cell receptor and a secondary costimulatory signal mediated by coreceptors. Costimulation is believed to act through the specific redistribution and clustering of membrane and intracellular kinase-ridge lipid raft microdomains at the contact site between T cells and antigen-presenting cells. This site is known as the immunological synapse. Endogenous mediators of raft clustering in lymphocytes are essential for T cell activation. Agrin induces the aggregation of signaling proteins and the creation of signaling domains in both immune and nervous systems through a common lipid raft pathway. A lipid raft is a membrane subdomain rich in cholesterol and glycosphingolipid-rich where cellular activation molecules are concentrated. Proteasome genes are two genes in the MHC class II region that encode two proteasome subunits. The proteasome is a protease complex in the cytosol that may participate in the generation of peptides from proteins in the cytosol.

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FIGURE 5.5 A schematic backbone structure of human class I histocompatibility antigen (HLA-A 0201) complexed with a decameric peptide from hepatitis B nucleocapsid protein (residues 18 to 27). Determined by x-ray crystallography.

LMP genes are two genes located in the MHC class II region in humans and mice that code for proteasome subunits. They are closely associated with the two TAP genes. LMP-2 and LMP-7 are catalytic subunits of the organelle (proteasome) that degrades cytosolic proteins into peptides in the class I MHC pathway of antigen presentation. MHC genes encode these two subunits which are upregulated by IFN-Îł and are especially significant in the generation of class I MHC-binding peptides. Tapasin: TAP-associated protein is a chaperone molecule that participates in the assembly of peptide-MHC class I

FIGURE 5.7 Topology of Tap 1 and Tap 2 proteins.

FIGURE 5.6 Longitudinal and transverse section through the 20S proteasome. The 20S proteasome is composed of two outer and two inner rings. The two outer rings each comprise seven copies of the 25.9-kDa Îą subunit. FIGURE 5.8 Antigen-presenting cell.

molecule complexes in the endoplasmic reticulum. Cells deficient in this protein have unstable MHC class I cell surface molecules. Tapasin (TAP-associated protein) is a molecule that is critical in MHC class I molecule assembly. Without this protein, MHC class I molecules are unstable on the cell surface. The transporter in antigen processing (TAP) 1 and 2 genes (Figure 5.7) are in the MHC class II region that must be expressed for MHC class I molecules to be assembled efficiently. TAP 1 and TAP 2 are postulated to encode components of a heterodimeric protein pump that conveys cytosolic peptides to the endoplasmic reticulum. Here they associate with MHC class I heavy chains. TAP 1 and TAP 2 genes: See transporter in antigen processing 1 and 2 genes. An antigen-presenting cell (APC) is a cell that can process a protein antigen, break it into peptides, and present it in conjunction with class II MHC molecules on the cell surface where it may interact with appropriate T cell receptors (Figure 5.8). Dendritic cells, macrophages, Langerhans cells, and B cells process and present antigen to immunoreactive lymphocytes such as CD4 + , helper/inducer T cells (Figure 5.8). A MHC transporter gene-encoded peptide supply factor may mediate peptide

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FIGURE 5.9 Properties and functions of antigen-presenting cells.

antigen presentation. Other antigen-presenting cells that serve mainly as passive antigen transporters include B cells, endothelial cells, keratinocytes, and Kupffer cells. APCs include cells that present exogenous antigen processed in their endosomal compartment and presented together with MHC class II molecules. Other APCs present antigen that has been endogenously produced by the body’s own cells with processing in an intracellular compartment and presentation together with class I MHC molecules. A third group of APCs present exogenous

antigen that is taken into the cell and processed, followed by presentation together with MHC class I molecules. In addition to processing and presenting antigenic peptides in association with MHC class II molecules, an antigenpresenting cell must also deliver a costimulatory signal that is necessary for T cell activation. Professional APCs include dendritic cells, macrophages, and B cells, whereas nonprofessional APCs that function in antigen presentation for only brief periods include thymic epithelial cells and vascular endothelial cells. Dendritic cells, macrophages, and B cells are the principal antigen-presenting cells for T cells, whereas follicular dendritic cells are the main antigen-presenting cells for B cells. Professional antigen-presenting cells are dendritic cells, macrophages, and B cells that are capable of initiating T lymphocyte responsiveness to antigen. These cells display antigenic peptide fragments in association with the proper class of MHC molecules and also bear costimulatory surface molecules. Dendritic cells are the most important professional APCs for initiating primary T lymphocyte responses. Among the three major antigen-presenting cells, dendritic cells are the only ones that continuously express high levels of costimulatory B7 and can present antigen via both class I MHC molecules and class II MHC molecules. Thus, they can activate both CD8 and CD4 T cells directly. APC is the abbreviation for antigen-presenting cell. Cathepsins are thiol and aspartyl proteases that have broad substrate specificities. Cathepsins represent the most abundant proteases of endosomes in antigen-presenting cells. They are believed to serve an important function in the generation of peptide fragments from exogenous protein antigens that bind to class II MHC molecules. CD2 is a T cell adhesion molecule that binds to the LFA3 adhesion molecule of antigen-presenting cells. Also called LFA-2. A circulating dendritic cell is one that has taken up antigen and is migrating to a secondary lymphoid tissue such as a lymph node. CD8 is a cell surface glycoprotein on T cells that recognizes antigens presented by MHC class I molecules. It binds to MHC class I molecules on antigen-presenting cells and serves as a coreceptor to facilitate the T cell’s response to antigen. The CD8 molecule is a heterodimer of an α and β chain that are covalently associated by disulfide bond. The two chains of a dimer have similar structures, each having a single domain resembling an immunoglobulin variable domain and a stretch of peptide believed to be in a relatively extended conformation.

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CD8 T cells comprise the T cell subset that expresses CD8 coreceptor and recognizes peptide antigens presented by MHC class I molecules. CD20 is a B cell marker with a molecular weight of 33, 35, and 37 kDa that appears relatively late in the B cell maturation (after the pro-B cell stage) and then persists for some time before the plasma cell stage. Its molecular structure resembles that of a transmembrane ion channel. The gene is on chromosome 11 at band q12-q13. It may be involved in regulating B cell activation. CD21 is a 145-kDa glycoprotein. Component of the B cell receptor. CD21 is a membrane molecule that participates in transmitting growth-promoting signals to the interior of the B cell. It is the receptor for the C3d fragment of the third component of complement, CR2. The CD21 antigen is a restricted B cell antigen expressed on mature B cells. It is present at high density on follicular dendritic cells (FDC), the accessory cells of the B zones. Also called complement receptor 2 (CR2). CD22 is a molecule with an α130- and β140-kDa mol wt that is expressed in the cytoplasm of B cells of the pro-B and pre-B cell stage and on the cell surface on mature B cells with surface Ig. The antigen is lost shortly before the terminal plasma cell phase. The molecule has five extracellular immunoglobulin domains and shows homology with myelin adhesion glycoprotein and with N-CAM (CD56). It participates in B cell adhesion to monocytes and T cells. Also called BL-CAM. CD28 is a T cell low-affinity receptor that interacts with B7 costimulatory molecules to facilitate T lymphocyte activation. More specifically, B7-1 and B7-2 ligands are expressed on the surface of activated antigen-presenting cells (APCs). Signals from CD28 to the T cell elevate expression of high affinity IL-2 receptor and increase the synthesis of numerous cytokines, including IL-2. CD28 regulates the responsiveness of T cells to antigen when they are in contact with APCs. It serves as a costimulatory receptor because its signals are synergistic with those provided by the T cell antigen receptor (TCR-CD3) in promoting T cell activation and proliferation. Without the signal from TCR-CD3, CD28’s signal is only able to stimulate minimal T cell proliferation and may even lead to T cell unresponsiveness. Tp44 (CD28) is a T lymphocyte receptor that regulates cytokine synthesis, thereby controlling responsiveness to antigen. Its significance in regulating activation of T lymphocytes is demonstrated by the ability of monoclonal antibody against CD28 receptor to block T cell stimulation by specific antigen. During antigen-specific activation of T lymphocytes, stimulation of the CD28 receptor occurs when it combines with the B7/BB1 coreceptor during the interaction between T and B lymphocytes. CD28 is a T

lymphocyte differentiation antigen that four fifths of CD3/Ti positive lymphocytes express. It is a member of the immunoglobulin superfamily. CD28 is found only on T lymphocytes and on plasma cells. There are 134 extracellular amino acids with a transmembrane domain and a brief cytoplasmic tail in each CD28 monomer. A costimulator is an antigen-presenting cell surface molecule that supplies a stimulus, serving as a second signal required for activation of naïve T lymphocytes, in addition to antigen (the “first signal”). An example of a costimulator is the B7 molecule on professional antigen-presenting cells that binds to the CD28 molecule on T lymphocytes. Costimulatory molecules are membrane bound or secreted products of accessory cells that activate signal transduction events in addition to those induced by MHC/TCR interactions. They are required for full activation of T cells, and it is thought that adjuvants may work by enhancing the expression of costimulator molecules by accessory cells. The interaction of CD28/CTLA-4 with B7 to induce full transcription of IL-2 mRNA is an example of costimulator mechanisms. A costimulatory signal is an extra signal requisite to induce proliferation of antigen-primed T lymphocytes. It is generated by the interaction of CD28 on T cells with B7 on antigen-presenting cells or altered self cells. In B cell activation an analogous second signal is illustrated by the interaction of CD40 on B cells with CD40L on activated TH cells. Cross-priming is the activation or priming of a naïve CD4+ cytotoxic T lymphocyte specific for antigens of a third cell such as a virus-infected cell or tumor cell by a professional antigen-presenting cell. Cross-priming takes place when a professional antigen-presenting cell ingests an infected cell and the microbial antigens are processed and presented in association with class I MHC molecules. The professional antigen-presenting cell also costimulates the T cells. Also referred to as cross-presentation. A coreceptor is a cell surface protein that increases the sensitivity of an antigen receptor to antigen by binding to associated ligands and facilitating in signaling for activation. CD4 and CD8 are T cell coreceptors that bind nonpolymorphic parts of a MHC molecule concurrently with the TCR binding to polymorphic residues and the bound peptide. It is a structure on the surface of a lymphocyte that binds to a part of an antigen simultaneously with membrane immunoglobulin (Ig) or T cell receptor (TCR) binding of antigen and which transmits signals required for optimal lymphocyte activation. CD4 and CD8 represent T cell coreceptors that bind nonpolymorphic regions of a major histocompatibility complex (MHC) molecule simultaneously with the binding of the T cell receptor to polymorphic residues and the exhibited peptide.

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MHC restriction is the recognition of an antigen in the context of either class I or class II molecules by the T cell receptor for antigen. In the afferent limb of the immune response, when antigen is being presented at the surface of a macrophage or other antigen-presenting cell to CD4+ T lymphocytes, this presentation must be in the context of MHC class II molecules for the CD4+ lymphocyte to recognize the antigen and proliferate in response to it. By contrast, cytotoxic (CD8+) T lymphocytes recognize foreign antigen, such as viral antigens on infected target cells, only in the context of class I MHC molecules. Once this recognition system is in place, the cytotoxic T cell can fatally injure the target cell through release of perforin molecules that penetrate the target cell surface. T cells recognize a firm peptide antigen only in the context of a specific allelic form of an MHC molecule to which it is bound. CTLA-4 is a molecule that is homologous to CD28 and expressed on activated T cells (Figure 5.10). The genes for CD28 and CTLA-4 are closely linked on chromosome 2. The binding of CTLA-4 to its ligand B7 is an important costimulatory mechanism (see CD28 and costimulatory molecules). CTLA-4 is a high affinity receptor for B7 costimulatory molecules on T lymphocytes. CTLA4-Ig is a soluble protein composed of the CD28 homolog CTLA and the constant region of an IgG1 molecule. It is used experimentally to inhibit the immune response by blocking CD28-B7 interaction. CTLA4-Ig is a soluble protein composed of the CD28 homolog CTLA and the constant region of an IgG1 molecule. It is used experimentally to inhibit the immune response by blocking CD28–B7 interaction. Immune costimulatory molecules B7-1 and B7-2, together with their receptors CD28 and CTLA-4, constitute one of the dominant costimulatory pathways that regulate T- and B-cell responses. Although both CTLA-4 and CD28 can bind to the same ligands, CTLA-4 binds to B71 and B7-2 with a 20- to 100-fold higher affinity than CD28 and is involved in the downregulation of the immune response. B7-1 is expressed on activated B and activated T cells and macrophages. B7-2 is constitutively expressed on interdigitating dendritic cells, Langerhans cells, peripheral blood dendritic cells, memory B cells, and germical center B cells. It has been observed that both human and mouse B7-1 and B7-2 can bind to either human or mouse CD28 and CTLA-4, suggesting that there are conserved amino acids which form the B7-1/B72/CD28/CTLA-4 critical binding sites. B7, B7-2: B7 is the ligand for CD28. B7 is expressed by accessory cells and is important in costimulataory mechanisms (Figure 5.11). Some APCs may upregulate expression of B7 following activation by various stimuli including

FIGURE 5.10 Participation of CTLA-4 molecules during antigen presentation.

FIGURE 5.12 Invariant chain promotes assembly of class II heterodimers from free chains.

FIGURE 5.11 B7.

An invariant (Ii) chain is a nonpolymorphic, 31-kDa glycoprotein that associates with class II histocompatibility molecules in the endoplasmic reticulum (Figure 5.12). It inhibits the linking of endogenous peptides with the class II molecule, conveying it to appropriate intracellular compartments. Truncation of the invariant chain stimulates a second signal that may function in the trans-Golgi network, prior to the conveyance of MHC class II molecules to the cell surface.

IFN-a, endotoxin, and MHC class II binding. B7 is also termed CD80. B7-2 is a costimulatory molecule whose sequence resembles that of B7. Dendritic cells, monocytes, activated T cells, and activated B lymphocytes may express B7-2.

A class II vesicle (CIIV) is a murine B cell membranebound organelle that is critical in the class II MHC pathway of antigen presentation. It contains all constituents requisite for the formation of peptide antigen and class II MHC molecular complexes, including the enzymes that

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FIGURE 5.13 The schematic representation of the interaction of the class II MHC, processed peptide antigen, and T cell receptor molecules during antigen presentation.

degrade protein antigens, class II molecules, invariant chain, and HLA-DM. I invariant (Ii): See invariant chain. Desetope is a term derived from “determinant selection.� It describes that region of class II histocompatibility molecules that reacts with the antigen during antigen presentation (Figure 5.13 and Figure 5.14). Allelic variation permits these contact residues to vary, which is one of the factors in histocompatibility molecule selection of a particular epitope that is being presented. An agretope refers to the region of a protein antigen that combines with an MHC class II molecule during antigen presentation. This is then recognized by the T cell receptor MHC class II complex. Amino acid sequences differ in their reactivity with MHC class II molecules. A histotope is the portion of an MHC class II histocompatibility molecule that reacts with a T lymphocyte receptor. A restitope is that segment of a T cell receptor that makes contact and interacts with a class II histocompatibility antigen molecule during antigen presentation.

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FIGURE 5.14 Costimulator for the activation of T cells.

CLIP is the processed fragment of invariant chain. In the MHC class II transport pathway, the peptide binding groove must be kept free of endogenous peptides. The cell uses one protein, called invariant chain (and its processed

fragment CLIP), to block the binding site until needed. HLA-DM facilitates release of CLIP peptides and their exchange for antigenic peptides as they become available. As long as CLIP remains in the binding groove, antigenic peptides cannot bind. A superantigen is an antigen such as a bacterial toxin that is capable of stimulating multiple T lymphocytes, especially CD4+ T cells, leading to the release of relatively large quantities of cytokines. Selected bacterial toxins may stimulate all T lymphocytes in the body that contain a certain family of V β T cell receptor genes. Superantigens may induce proliferation of 10% of CD4+ T cells by combining with the T cell receptor V β and to the MHC HLA-DR α-1 domain. Superantigens are thymus-dependent (TD) antigens that do not require phagocytic processing. Instead of fitting into the TCR internal groove where a typical processed peptide antigen fits, superantigens bind to the external region of the αβ TCR and simultaneously link to DP, DQ, or DR molecules on antigen-presenting cells (Figure 5.15). Superantigens react with multiple TCR molecules whose peripheral structure is similar. Thus, they stimulate multiple T cells that augment a protective T and B cell antibody response. This enhanced responsiveness to antigens such as toxins produced by staphylococci and

streptococci is an important protective mechanism in the infected individual (Figure 5.16). Several staphylococcal enterotoxins are superantigens and may activate many T cells resulting in the release of large quantities of cytokines and producing a clinical syndrome resembling septic shock.

FIGURE 5.15 Superantigen.

FIGURE 5.16 Cellular and molecular interactions in antigen presentation.

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FIGURE 5.17 Granuloma.

MHC-I antigen presentation: Proteins in the cytosol, such as those derived from viruses, may be processed through the class I MHC route of antigen presentation. The multiprotein complex in the cytoplasm, known as the proteasome effects, involve proteolytic degradation of proteins in the cytoplasm to yield many of the peptides that are presented by class I MHC molecules. TAP molecules transport peptides from the cytoplasm to the endoplasmic reticulum, where they interact and bind to class I MHC dimeric molecules. Once the class I MHC molecules have become stabilized through peptide binding, the complex leaves the endoplasmic reticulum, entering the Golgi apparatus en route to the surface of the cell. Thus, mechanisms are provided through MHC-restricted antigen presentation to guarantee that peptides derived from extracellular microbial proteins can be presented by class II MHC molecules to CD4+ helper T cells and that peptides derived from intracellular microbes can be presented by class I MHC molecules to CD8+ cytotoxic T lymphocytes. The generation of microbial peptides produced through antigen processing to combine with self MHC molecules is critical to the development of an appropriate immune response. A granuloma is a tissue reaction characterized by altered macrophages (epithelioid cells), lymphocytes, and fibroblasts

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(Figure 5.17). These cells form microscopic masses of mononuclear cells. Giant cells form from some of these fused cells. Granulomas may be of the foreign body type, such as those surrounding silica or carbon particles, or of the immune type that encircle particulate antigens derived from microorganisms. Activated macrophages trap antigen which may cause T cells to release lymphokines causing more macrophages to accumulate. This process isolates the microorganism. Granulomas appear in cases of tuberculosis and develop under the influence of helper T cells that react against Mycobacterium tuberculosis. Some macrophages and epithelioid cells fuse to form multinucleated giant cells in immune granulomas. There may also be occasional neutrophils and eosinophils. Necrosis may develop. It is a delayed type of hypersensitivity reaction that persists as a consequence of the continuous presence of foreign body or infection. HAM-1 and HAM-2 (histocompatibility antigen modifier): These are two murine genes that determine formation of permeases that are antigen transporters (oligopeptides) from the cytoplasm to a membrane-bound compartment where antigen complexes with MHC class I and class II molecules. In man, the equivalents of HAM-1 and HAM2 are termed ATP-binding cassette transporters.

complexes have been successfully used in delineating the different responses of B and T cells to each part of this complex. Immunization of a rabbit or other animal with a particular hapten–carrier complex will induce a primary immune response, and a second injection of the same hapten–carrier conjugate will induce a secondary immune response. However, linkage of the same hapten to a different carrier elicits a much weaker secondary response in an animal primed with the original hapten–carrier complex. This is termed the carrier effect. B lymphocytes recognize the hapten and T lymphocytes the carrier.

FIGURE 5.18 T–B cell interactions.

T lymphocyte–B lymphocyte cooperation refers to the association of B cell and helper T cell through a number of receptor–ligand interactions at the surfaces of both cell types which leads to B cell proliferation and differentiation into plasma cells that synthesize and secrete specific antibody specific for thymus-dependent antigen (Figure 5.15). B cell immunoglobulin receptors react with protein antigens. This is followed by endocytosis, antigen processing, and presentation to helper T lymphocytes. Their antigen-specific T cell receptors recognize processed antigens only in the context of MHC class II molecules on the B cell surface during antigen presentation. CD4+ helper T cells secrete lymphokines, including IL-2, which promote B cell growth and differentiation into plasma cells that secrete specific antibodies. T cells are required for B cells to be able to switch from forming IgM to synthesizing IgG or IgA. B and T lymphocytes recognize different antigens. B cells may recognize peptides, native proteins, or denatured proteins. T cells are more complex in their recognition system in that a peptide antigen can be presented to them only in the context of MHC class II or class I histocompatibility molecules. Hapten–carrier

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By light microscopy, resting lymphocytes appear as a distinct and homogeneous population of round cells, each with a large, spherical or slightly kidney-shaped nucleus which occupies most of the cell and is surrounded by a narrow rim of basophilic cytoplasm with occasional vacuoles. The nucleus usually has a poorly visible single indentation and contains densely packed chromatin. Occasionally, nucleoli can be distinguished. The small lymphocyte variant, which is the predominant morphologic form, is slightly larger than an erythrocyte. Larger lymphocytes, ranging between 10 and 20 µm in diameter, are difficult to differentiate from monocytes. They have more cytoplasm and may show azurophilic granules. Intermediate-size forms between the two are described. By phase contrast microscopy, living lymphocytes show a feeble motility with ameboid movements that give the cells a hand-mirror shape. The mirror handle is called a uropod. In large lymphocytes, mitochondria and lysosomes are better visualized, and some cells show a spherical, birefringent, 0.5-µm diameter inclusion, called a gall body. Lymphocytes do not spread on surfaces. The different classes of lymphocytes cannot be distinguished by light microscopy. By scanning electron microscopy, B lymphocytes sometimes show a hairy (rough) surface, but this is apparently an artifact. Electron microscopy does not provide additional information except for visualization of the cellular organelles which are not abundant. This suggests that the small, resting lymphocytes are end-stage cells. However, under appropriate stimulation, they are capable of considerable morphologic changes. Cooperation refers to T lymphocyte–B lymphocyte cooperation. Cooperativity refers to the effect observed when two binding sites are linked to their ligand to yield an effect of binding to both that is greater than the sum of each binding site acting independently. Cognate interaction refers to the interaction of processed antigen on a B cell surface interacting with a T cell receptor for antigen resulting in B cell differentiation into an antibody-producing cell. Cognate recognition refers to cognate interaction.

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