免疫学（英文版）：Chapter 12

a73 Properties of Cytokines a73 Cytokine Receptors a73 Cytokine Antagonists a73 Cytokine Secretion by T H 1 and T H 2 Subsets a73 Cytokine-Related Diseases a73 Therapeutic Uses of Cytokines and Their Receptors a73 Cytokines in Hematopoiesis Class I Cytokine Receptors Cytokines T ?? ??????????? ?? ?? ????????? ?????? response involves lymphoid cells, inflammatory cells, and hematopoietic cells. The complex inter- actions among these cells are mediated by a group of pro- teins collectively designated cytokines to denote their role in cell-to-cell communication. Cytokines are low-molecular- weight regulatory proteins or glycoproteins secreted by white blood cells and various other cells in the body in response to a number of stimuli. These proteins assist in reg- ulating the development of immune effector cells, and some cytokines possess direct effector functions of their own. This chapter focuses on the biological activity of cyto- kines, the structure of cytokines and their receptors, signal transduction by cytokine receptors, the role of cytokine abnormalities in the pathogenesis of certain diseases, and therapeutic uses of cytokines or their receptors. The impor- tant role of cytokines in the inflammatory response is described in Chapter 15. Properties of Cytokines Cytokines bind to specific receptors on the membrane of target cells, triggering signal-transduction pathways that ultimately alter gene expression in the target cells (Figure 12-1a). The susceptibility of the target cell to a particular cytokine is determined by the presence of specific mem- brane receptors. In general, the cytokines and their receptors exhibit very high affinity for each other, with dissociation constants ranging from 10 –10 to 10 –12 M. Because their af- finities are so high, cytokines can mediate biological effects at picomolar concentrations. A particular cytokine may bind to receptors on the mem- brane of the same cell that secreted it, exerting autocrine action; it may bind to receptors on a target cell in close prox- imity to the producer cell, exerting paracrine action; in a few cases, it may bind to target cells in distant parts of the body, exerting endocrine action (Figure 12-1b). Cytokines regu- late the intensity and duration of the immune response by stimulating or inhibiting the activation, proliferation, and/ or differentiation of various cells and by regulating the secre- tion of antibodies or other cytokines. As described later, binding of a given cytokine to responsive target cells gener- ally stimulates increased expression of cytokine receptors and secretion of other cytokines, which affect other target cells in turn. Thus, the cytokines secreted by even a small number of lymphocytes activated by antigen can influence the activity of numerous cells involved in the immune response. For example, cytokines produced by activated T H cells can influence the activity of B cells, T C cells, natural killer cells, macrophages, granulocytes, and hematopoietic stem cells, thereby activating an entire network of interact- ing cells. Cytokines exhibit the attributes of pleiotropy, redun- dancy, synergy, antagonism, and cascade induction, which permit them to regulate cellular activity in a coordinated, interactive way (Figure 12-2). A given cytokine that has different biological effects on different target cells has a pleiotropic action. Two or more cytokines that mediate sim- ilar functions are said to be redundant; redundancy makes it difficult to ascribe a particular activity to a single cytokine. Cytokine synergism occurs when the combined effect of two cytokines on cellular activity is greater than the additive chapter 12 effects of the individual cytokines. In some cases, cytokines exhibit antagonism; that is, the effects of one cytokine inhibit or offset the effects of another cytokine. Cascade induction occurs when the action of one cytokine on a target cell induces that cell to produce one or more other cytokines, which in turn may induce other target cells to produce other cytokines. The term cytokine encompasses those cytokines secreted by lymphocytes, substances formerly known as lympho- kines, and those secreted by monocytes and macrophages, substances formerly known as monokines. Although these other two terms continue to be used, they are misleading because secretion of many lymphokines and monokines is not limited to lymphocytes and monocytes as these terms imply, but extends to a broad spectrum of cells and types. For this reason, the more inclusive term cytokine is preferred. Many cytokines are referred to as interleukins, a name indicating that they are secreted by some leukocytes and act upon other leukocytes. Interleukins 1–25 have been identi- fied. There is reason to suppose that still other cytokines will be discovered and that the interleukin group will expand further. Some cytokines are known by common names, including the interferons and tumor necrosis factors. Re- cently gaining prominence is yet another another subgroup of cytokines, the chemokines, a group of low-molecular- weight cytokines that affect chemotaxis and other aspects of leukocyte behavior. These molecules play an important role in the inflammatory response and are described in Chapter 15. Because cytokines share many properties with hormones and growth factors, the distinction between these three classes of mediators is often blurred. All three are secreted soluble factors that elicit their biological effects at picomolar concentrations by binding to receptors on target cells. Growth factors tend to be produced constitutively, whereas cytokines and hormones are secreted in response to discrete stimuli, and secretion is short-lived, generally ranging from a few hours to a few days. Unlike hormones, which generally act long range in an endocrine fashion, most cytokines act over a short distance in an autocrine or paracrine fashion. In addition, most hormones are produced by specialized glands and tend to have a unique action on one or a few types of tar- get cell. In contrast, cytokines are often produced by, and bind to, a variety of cells. The activity of cytokines was first recognized in the mid- 1960s, when supernatants derived from in vitro cultures of lymphocytes were found to contain factors that could regulate proliferation, differentiation, and maturation of allogeneic Cytokines CHAPTER 12 277 Gene activation Biological effects Signal Cytokine gene Inducing stimulus (a) Cytokine-producing cell Target cell Cytokine Receptor (b) Endocrine action Circulation Distant cell Paracrine action Nearby cell Autocrine action FIGURE 12-1 (a) Overview of the induction and function of cytokines. (b) Most cytokines exhibit autocrine and/or paracrine action; fewer exhibit endocrine action. immune-system cells. Soon after, it was discovered that pro- duction of these factors by cultured lymphocytes was induced by activation with antigen or with nonspecific mitogens. Bio- chemical isolation and purification of cytokines was ham- pered because of their low concentration in culture super- natants and the absence of well-defined assay systems for individual cytokines. A great advance was made with the development of gene-cloning techniques during the 1970s and 1980s, which made it possible to produce pure cytokines by expressing the protein from cloned genes. The discovery of 278 PART III Immune Effector Mechanisms Activated T H cells PLEIOTROPY Mast cell IL-4 Thymocyte B cell Activation Proliferation Differentiation Proliferation Target Cell Effect IL-4 Proliferation IL-2 IL-5 Activated T H cells REDUNDANCY IL-4 + IL-5 Activated T H cells SYNERGY Induces class switch to IgE Blocks class switch to IgE induced by IL-4 IL-4 IFN-γ Activated T H cells ANTAGONISM Proliferation B cell B cell B cell Activated T H cells Activated T H cells IFN-γ, TNF, IL-2, and other cytokines Macrophage IL-12 CASCADE INDUCTION IFN-γ (a) (b) FIGURE 12-2 Cytokine attributes of (a) pleiotropy, redundancy, synergy (synergism), antagonism, and (b) cascade induction. cell lines whose growth depended on the presence of a partic- ular cytokine provided researchers with the first simple assay systems. The derivation of monoclonal antibodies specific for each of the more important cytokines has made it possible to develop rapid quantitative immunoassays for each of them (Figure 12-3). Cytokines Belong to Four Structural Families Once the genes encoding various cytokines had been cloned, sufficient quantities of purified preparations became avail- able for detailed studies on their structure and function. Cytokines generally have a molecular mass of less than 30 kDa. Structural studies have shown that the cytokines characterized so far belong to one of four groups: the hematopoietin family, the interferon family, the chemokine family, or the tumor necrosis factor family. The structures of two members of the hematopoietin family, IL-2 and IL-4, are depicted in Figure 12-4. Although the amino acid sequences of these family members differ considerably, all of them have a high degree of H9251-helical structure and little or no H9252-sheet structure. The molecules Cytokines CHAPTER 12 279 Add enzyme- conjugated secondary antibody Cytokine bound to antibody coated onto well Add substrate and measure color S S EEE E (a) (b) Concentration of IL-12 (pg/ml) Optical density 10 1 0.1 0.01 0.1 1 10 100 α SS 105 C N 6 133 114 30 36 42 73 80 96 52 58 AD C B C 105 B–C loop β strand C–D loop A–B loop COOH NH 2 A C B ' B D α P6 5 C 58 (a) Interleukin 2 (b) Interleukin 4 A B C 2 1 D FIGURE 12-3 ELISA assay of a cytokine. (a) The sample contain- ing the cytokine of interest is captured by specific antibody (blue) coated onto wells of a microtiter plate. A second specific antibody (blue), conjugated to an enzyme (E) such as horseradish peroxidase, forms a sandwich with the captured cytokine, immobilizing the en- zyme in the microtiter well. A chromogenic substrate (S) is added, and the enzyme generates a color whose intensity is proportional to the amount of cytokine bound to the capture antibody. The optical density of this color produced by the unknown is compared with val- ues on an appropriately determined standard curve. (b) The standard curve shown here is for human interleukin 12 (IL-12). It is clear that this assay is sufficiently sensitive to detect as little as 1 picogram of IL-12. [Part (b) courtesy of R&D Systems.] FIGURE 12-4 Several representations of structures in the hemato- poietin family. (a) Left: Topographical representation of the primary structure of IL-2 showing H9251-helical regions (H9251 and A–D) and connect- ing chains of the molecule. Right: Proposed three-dimensional model of IL-2. (b) Ribbon model of IL-4 deduced from x-ray crystallographic analysis of the molecule. In (a) and (b) the H9251 helices are shown in red and the H9252 sheets in blue. The structures of other cytokines belonging to the hematopoietin family are thought to be generally similar. [Part (b) from J. L. Boulay and W. E. Paul, 1993, Curr. Biol. 3:573.] share a similar polypeptide fold, with four H9251-helical regions (A–D) in which the first and second helices and the third and fourth helices run roughly parallel to one another and are connected by loops. Cytokines Have Numerous Biological Functions Although a variety of cells can secrete cytokines, the two principal producers are the T H cell and the macrophage. Cytokines released from these two cell types activate an entire network of interacting cells (Figure 12-5). Among the numerous physiologic responses that require cytokine in- volvement are development of cellular and humoral im- mune responses, induction of the inflammatory response, regulation of hematopoiesis, control of cellular proliferation and differentiation, and the healing of wounds. Although the immune response to a specific antigen may include the pro- duction of cytokines, it is important to remember that cytokines act in an antigen-nonspecific manner. That is, they affect whatever cells they encounter that bear appropriate receptors and are in a physiological state that allows them to respond. Cytokines are involved in a staggeringly broad array of biological activities including innate immunity, adaptive immunity, inflammation, and hematopoiesis. Altogether, the total number of proteins with cytokine activity easily exceeds 100 and research continues to uncover new ones. Table 12-1 summarizes the activities of some cytokines and places them into functional groups. An expanded list of cytokines can be found in the Appendix. It should be kept in mind that most of the listed functions have been identified from analysis of 280 PART III Immune Effector Mechanisms VISUALIZING CONCEPTS Inflammation Macrophage Resting T H cell Adaptive immunity Cytokine promotion of macrophage activation Cytokine promotion of T H cell differentiation Hemato- poiesis Cytokine promotion of activation, differentiation, proliferation, or cell death of T cells, B cells, macrophages, dendritic cells, NK cells, and other leukocytes. FIGURE 12-5 Interaction of antigen with macrophages and the subsequent activation of resting T H cells leads to release of nu- merous cytokines (blue arrows), generating a complex network of interacting cells in the immune response. the effects of recombinant cytokines, often at nonphysiologic concentrations, added individually to in vitro systems. In vivo, however, cytokines rarely, if ever, act alone. Instead, a target cell is exposed to a milieu containing a mixture of cytokines, whose combined synergistic or antagonistic ef- fects can have very different consequences. In addition, cytokines often induce the synthesis of other cytokines, re- sulting in cascades of activity. The nonspecificity of cytokines seemingly conflicts with the established specificity of the immune system. What keeps the nonspecific cytokines from activating cells in a nonspe- cific fashion during the immune response? One way in which specificity is maintained is by careful regulation of the ex- pression of cytokine receptors on cells. Often cytokine recep- tors are expressed on a cell only after that cell has interacted with antigen. In this way cytokine activation is limited to antigen-activated lymphocytes. Another means of maintain- ing specificity may be a requirement for direct interaction between the cytokine-producing cell and the target cell to trigger cytokine secretion, thus ensuring that effective concen- trations of the cytokine are released only in the vicinity of the intended target. In the case of the T H cell, a major producer of cytokines, close cellular interaction occurs when the T-cell receptor recognizes an antigen-MHC complex on an appro- priate antigen-presenting cell, such as a macrophage, den- dritic cell, or B lymphocyte. Cytokines secreted at the junction of these interacting cells reach high enough local concentra- tions to affect the target APC but not more distant cells. In addition, the half-life of cytokines in the bloodstream or other extracellular fluids into which they are secreted is usually very short, ensuring that they act for only a limited period of time and thus over a short distance. Cytokines CHAPTER 12 281 TABLE 12-1 Functional groups of selected cytokines 1 Cytokine* Secreted by** Targets and effects SOME CYTOKINES OF INNATE IMMUNITY Interleukin 1 (IL-1) Monocytes, macrophages, Vasculature (inflammation); hypothalamus (fever); endothelial cells, epithelial cells l iver (induction of acute phase proteins) Tumor Necrosis Macrophages Vasculature (inflammation); liver (induction of acute phase Factor-H9251 (TNF-H9251) proteins); loss of muscle, body fat (cachexia); induction of death in many cell types; neutrophil activation Interleukin 12 (IL-12) Macrophages, dendritic cells NK cells; influences adaptive immunity (promotes T H 1 subset) Interleukin 6 (IL-6) Macrophages, endothelial cells Liver (induces acute phase proteins); influences adaptive immunity (proliferation and antibody secretion of B cell lineage) Interferon H9251 (IFN-H9251) Macrophages Induces an antiviral state in most nucleated cells; increases MHC (This is a family of molecules) class I expression; activates NK cells Interferon H9252 (IFN-H9252) Fibroblasts Induces an antiviral state in most nucleated cells; increases MHC class I expression; activates NK cells SOME CYTOKINES OF ADAPTIVE IMMUNITY Interleukin 2 (IL-2) T cells T-cell proliferation; can promote AICD. NK cell activation and proliferation; B-cell proliferation Interleukin 4 (IL-4)T H 2 cells; mast cells Promotes T H 2 differentiation; isotype switch to IgE Interleukin 5 (IL-5 H 2 cells Eosinophil activation and generation Interleukin 25 (IL-25) Unknown Induces secretion of T H 2 cytokine profile Transforming growth T cells, macrophages, Inhibits T-cell proliferation and effector functions; inhibits factor H9252 (TGF-H9252) other cell types B-cell proliferation; promotes isotype switch to IgE; inhibits macrophages Interferon H9253 (IFN-H9253)T H 1 cells; CD8 + cells; NK cells Activates macrophages; increases expression MHC class I and class II molecules; increases antigen presentation 1 Many cytokines play roles in more than one functional category. *Only the major cell types providing cytokines for the indicated activity are listed; other cell types may also have the capacity to synthesize the given cytokine. **Also note that activated cells generally secrete greater amounts of cytokine than unactivated cells. Cytokine Receptors As noted already, to exert their biological effects, cytokines must first bind to specific receptors expressed on the mem- brane of responsive target cells. Because these receptors are expressed by many types of cells, the cytokines can affect a diverse array of cells. Biochemical characterization of cyto- kine receptors initially progressed at a very slow pace because their levels on the membrane of responsive cells is quite low. As with the cytokines themselves, cloning of the genes encod- ing cytokine receptors has led to rapid advances in the iden- tification and characterization of these receptors. Cytokine Receptors Fall Within Five Families Receptors for the various cytokines are quite diverse struc- turally, but almost all belong to one of five families of recep- tor proteins (Figure 12-6): a73 Immunoglobulin superfamily receptors a73 Class I cytokine receptor family (also known as the hematopoietin receptor family) a73 Class II cytokine receptor family (also known as the interferon receptor family) a73 TNF receptor family a73 Chemokine receptor family Many of the cytokine-binding receptors that function in the immune and hematopoietic systems belong to the class I cytokine receptor family. The members of this receptor family have conserved amino acid sequence motifs in the extracellu- lar domain consisting of four positionally conserved cysteine residues (CCCC) and a conserved sequence of tryptophan- serine-(any amino acid)-tryptophan-serine (WSXWS, where X is the nonconserved amino acid). The receptors for all the cytokines classified as hematopoietins belong to the class I cytokine receptor family, which also is called the hematopoi- etin receptor family. The class II cytokine receptors possess the conserved CCCC motifs, but lack the WSXWS motif pre- sent in class I cytokine receptors. Initially only the three interferons, H9251, H9252, and H9253, were thought to be ligands for these receptors. However, recent work has shown that the IL-10 receptor is also a member of this group. Another feature common to most of the hematopoietin (class I cytokine) and the class II cytokine receptor families is 282 PART III Immune Effector Mechanisms RECEPTOR FAMILY LIGANDS (a) Immunoglobulin superfamily receptors IL-1 M-CSF C-Kit S S S S S S (b) Class I cytokine receptors (hematopoietin) IL-2 IL-3 IL-4 IL-5 IL-6 IL-7 IL-9 IL-11 IL-12 IL-13 IL-15 GM-CSF G-CSF OSM LIF CNTF Growth hormone Prolactin Conserved cysteines WSXWS (c) Class II cytokine receptors (interferon) IFN-α IFN-β IFN-γ IL-10 C C C C (d) TNF receptors C1 C3 C2 C1 C3 C2 C1 C3 C2 C1 C3 C2 TNF-α TNF-β CD40 Nerve growth factor (NGF) FAS (e) Chemokine receptors IL-8 RANTES MIP-1 PF4 MCAF NAP-2 G-protein FIGURE 12-6 Schematic diagrams showing the structural features that define the five types of receptor proteins to which most cytokines bind. The receptors for most of the interleukins belong to the class I cytokine receptor family. C refers to conserved cysteine. multiple subunits, often including one subunit that binds specific cytokine molecules and another that mediates signal transduction. Note, however, that these two functions are not always confined to one subunit or the other. Engagement of all of the class I and class II cytokine receptors studied to date has been shown to induce tyrosine phosphorylation of the receptor through the activity of protein tyrosine kinases closely associated with the cytosolic domain of the receptors. Subfamilies of Class I Cytokine Receptors Have Signaling Subunits in Common Several subfamilies of class I cytokine receptors have been identified, with all the receptors in a subfamily having an identical signal-transducing subunit. Figure 12-7 schemati- cally illustrates the members of three receptor subfamilies, named after GM-CSF, IL-2, and IL-6. The sharing of signal-transducing subunits among recep- tors explains the redundancy and antagonism exhibited by some cytokines. Consider the GM-CSF receptor subfamily, which includes the receptors for IL-3, IL-5, and GM-CSF (see Figure 12-7a). Each of these cytokines binds to a unique low- affinity, cytokine-specific receptor consisting of an H9251 subunit only. All three low-affinity subunits can associate noncova- lently with a common signal-transducing H9252 subunit. The re- sulting dimeric receptor not only exhibits increased affinity for the cytokine but also can transduce a signal across the membrane after binding the cytokine (Figure 12-8a). Inter- estingly, IL-3, IL-5, and GM-CSF exhibit considerable redun- dancy. IL-3 and GM-CSF both act upon hematopoietic stem cells and progenitor cells, activate monocytes, and induce megakaryocyte differentiation. All three of these cytokines induce eosinophil proliferation and basophil degranulation with release of histamine. Since the receptors for IL-3, IL-5, and GM-CSF share a common signal-transducing H9252 subunit, each of these cyto- kines would be expected to transduce a similar activation sig- nal, accounting for the redundancy among their biological effects (Figure 12-8b). In fact, all three cytokines induce the same patterns of protein phosphorylation. Furthermore, IL-3 and GM-CSF exhibit antagonism; IL-3 binding has been shown to be inhibited by GM-CSF, and conversely, binding Cytokines CHAPTER 12 283 GM-CSF IL-3 IL-5 ββ β (a) GM-CSF receptor subfamily (common β subunit) (c) IL-2 receptor subfamily (common γ subunit) IL-6 CNTF IL-11 (b) IL-6 Receptor subfamily (common gp130 subunit) CNTFR IL-2Rβ IL-2Rβ IL-2Rα IL-15Rα IL-7R IL-9R γγγγ IL-15 IL-9IL-7 IL-4R γ IL-4IL-2 GM-CSFRα IL-3R IL-5R LIF/OSM gp130gp130gp130gp130 FIGURE 12-7 Schematic diagrams of the three subfamilies of class I cytokine receptors. All members of a subfamily have a common sig- nal-transducing subunit (blue), but a unique cytokine-specific subunit. In addition to the conserved cysteines (double black lines) and WSXWS motifs (red lines) that characterize class I cytokine receptors, immu- noglobulin-like domains are present in some of these receptors. CNTF = ciliary neurotrophic factor; LIF/OSM = leukemia-inhibitory factor/ oncostatin. [Adapted from K. Sugamura et al., 1996, Annu. Rev. Im- munol. 14:179.] of GM-CSF has been shown to be inhibited by IL-3. Since the signal-transducing H9252 subunit is shared between the re- ceptors for these two cytokines, their antagonism is due to competition for a limited number of H9252 subunits by the cytokine-specific H9251 subunits of the receptors (Figure 12-8c). A similar situation is found among the IL-6 receptor sub- family, which includes the receptors for IL-6, IL-11, leukemia- inhibitory factor (LIF), oncostatin M (OSM), and ciliary neurotrophic factor (CNTF) (see Figure 12-7b). In this case, a common signal-transducing subunit called gp130 associ- ates with one or two different cytokine-specific subunits. LIF and OSM, which must share certain structural features, both bind to the same H9251 subunit. As expected, the cytokines that bind to receptors in this subfamily display overlapping bio- logical activities: IL-6, OSM, and LIF induce synthesis of acute-phase proteins by liver hepatocytes and differentiation of myeloid leukemia cells into macrophages; IL-6, LIF, and CNTF affect neuronal development, and IL-6, IL-11, and OSM stimulate megakaryocyte maturation and platelet pro- duction. The presence of gp130 in all receptors of the IL-6 subfamily explains their common signaling pathways as well as the binding competition for limited gp130 molecules that is observed among these cytokines. A third signal-transducing subunit defines the IL-2 recep- tor subfamily, which includes receptors for IL-2, IL-4, IL-7, IL-9, and IL-15 (see Figure 12-7c). The IL-2 and the IL-15 receptors are heterotrimers, consisting of a cytokine-specific H9251 chain and two chains—H9252 and H9253—responsible for signal transduction. The IL-2 receptor H9253 chain functions as the sig- nal-transducing subunit in the other receptors in this sub- family, which are all dimers. Recently, it has been shown that congenital X-linked severe combined immunodeficiency (XSCID) results from a defect in the H9253-chain gene, which maps to the X chromosome. The immunodeficiencies ob- served in this disorder are due to the loss of all the cytokine functions mediated by the IL-2 subfamily receptors. The IL-2R Is One of the Most Thoroughly Studied Cytokine Receptors Because of the central role of IL-2 and its receptor in the clonal proliferation of T cells, the IL-2 receptor has received intensive study. As noted in the previous section, the com- plete trimeric receptor comprises three distinct subunits— the H9251, H9252, and H9253 chains. The H9252 and H9253 chains belong to the class I cytokine receptor family, containing the characteristic CCCC and WSXWS motifs, whereas the H9251 chain has a quite different structure and is not a member of this receptor family (see Figure 12-7c). The IL-2 receptor occurs in three forms that exhibit dif- ferent affinities for IL-2: the low-affinity monomeric IL-2RH9251, the intermediate-affinity dimeric IL-2RH9252H9253, and the high- affinity trimeric IL-2RH9251H9252H9253 (Figure 12-9). Because the H9251 chain is expressed only by activated T cells, it is often referred to as the TAC (T-cell activation) antigen. A monoclonal anti- 284 PART III Immune Effector Mechanisms β α High-affinity receptors β α β α α Low-affinity receptors β subunit αα Exterior Membrane Interior IL-3 IL-5 GM-CSF (a) α Same signal from all three αα (b) GM-CSFIL-3 αααααα βββ βββ ααα ααα (c) FIGURE 12-8 Interactions between cytokine-specific subunits and a common signal-transducing subunit of cytokine receptors. (a) Sche- matic diagram of the low-affinity and high-affinity receptors for IL-3, IL-5, and GM-CSF. The cytokine-specific subunits exhibit low-affinity binding and cannot transduce an activation signal. Noncovalent association of each subunit with a common H9252 subunit yields a high-affinity dimeric re- ceptor that can transduce a signal across the membrane. (b) Associa- tion of cytokine-specific subunits with a common signaling unit, the H9252 subunit, allows the generation of cytokine-specific signals despite the generation of the same signal by the different cytokine receptors shown. (c) Competition of ligand-binding chains of different receptors for a common subunit can produce antagonistic effects between cytokines. Here binding of IL-3 by H9251 subunits of the IL-3 receptor allows them to out-compete H9251 chains of the GM-CSF receptor for H9252 subunits. [Part (a) adapted from T. Kishimoto et al., 1992, Science 258:593.] body, designated anti-TAC, which binds to the 55-kDa H9251 chain, is often used to identify IL-2RH9251 on cells. Signal trans- duction by the IL-2 receptor requires both the H9252 and H9253 chains, but only the trimeric receptor containing the H9251 chain as well binds IL-2 with high affinity. Although the H9253 chain appears to be constitutively expressed on most lymphoid cells, expression of the H9251 and H9252 chains is more restricted and is markedly enhanced after antigen has activated resting lymphocytes. This phenomenon ensures that only antigen- activated CD4 + and CD8 + T cells will express the high- affinity IL-2 receptor and proliferate in response to physio- logic levels of IL-2. Activated T cells express approximately 5 H11003 10 3 high-affinity receptors and ten times as many low- affinity receptors. NK cells express the H9252 and H9253 subunits con- stitutively, accounting for their ability to bind IL-2 with an intermediate affinity and to be activated by IL-2. Engaged Cytokine Receptors Activate Signaling Pathways While some important cytokine receptors lie outside the class I and class II families, the majority are included within these two families. As mentioned previously, class I and class II cytokine receptors lack signaling motifs (e.g., intrinsic tyrosine kinase domains). Yet, early observations demon- strated that one of the first events after the interaction of a cytokine with one of these receptors is a series of pro- tein tyrosine phosphorylations. While these results were ini- tially puzzling, they were explained when a unifying model emerged from studies of the molecular events triggered by binding of interferon gamma (IFN-H9253) to its receptor, a mem- ber of the class II family. IFN-H9253 was originally discovered because of its ability to induce cells to block or inhibit the replication of a wide vari- ety of viruses. Antiviral activity is a property it shares with IFN-H9251 and IFN-H9252. However, unlike these other interferons, IFN-H9253 plays a central role in many immunoregulatory processes, including the regulation of mononuclear phago- cytes, B-cell switching to certain IgG classes, and the support or inhibition of the development of T H -cell subsets. The dis- covery of the major signaling pathway invoked by interaction of IFN-H9253 with its receptor led to the realization that signal transduction through most, if not all, class I and class II cytokine receptors involves the following steps, which are the basis of a unifying signaling model (Figure 12-10). a73 The cytokine receptor is composed of separate subunits, an H9251 chain required for cytokine binding and for signal transduction and a H9252 chain necessary for signaling but with only a minor role in binding. a73 Different inactive protein tyrosine kinases are associated with different subunits of the receptor. The H9251 chain of the receptor is associated with a novel family of protein tyrosine kinases, the Janus kinase (JAK)* family. The association of the JAK and the receptor subunit occurs spontaneously and does not require the binding of cytokine. However, in the absence of cytokine, JAKs lack protein tyrosine kinase activity. a73 Cytokine binding induces the association of the two separate cytokine receptor subunits and activation of the receptor-associated JAKs. The ability of IFN-H9253,which binds to a class II cytokine receptor, to bring about the association of the ligand-binding chains of its receptor has been directly demonstrated by x-ray crystallographic studies, as shown in Figure 12-11. a73 Activated JAKs create docking sites for the STAT transcription factors by phosphorylation of specific tyrosine residues on cytokine receptor subunits. Once receptor- associated JAKs are activated, they phosphorylate specific tyrosines in the receptor subunits of the Cytokines CHAPTER 12 285 Subunit composition: Dissociation constant (K d ): Cells expressed by: IL-2Rβ IL-2Rγ IL-2Rα IL-2Rβ IL-2Rγ IL-2Rα 10 ?9 M 10 ?11 M 10 ?8 M Affinity constant (K a ): 10 7 M 10 11 M 10 8 M NK cells Resting T cells (low numbers) Activated CD4 + and CD8 + T cells Activated B cells (low numbers) β γ α α β γ Intermediate affinity IL-2R High affinity IL-2R Low affinity IL-2R FIGURE 12-9 Comparison of the three forms of the IL-2 receptor. Signal transduction is mediated by the H9252 and H9253 chains, but all three chains are required for high-affinity binding of IL-2. *The Roman god Janus had two faces. Kinases of the Janus family have two sites, a binding site at which they link with the cytokine receptor subunit and a catalytic site that, when activated, has protein tyrosine kinase activity. Some bio- chemists, wearied by the multitude of different protein kinases that have been discovered, claim JAK means Just Another Kinase. complex. Members of a family of transcription factors known as STATs (signal transducers and activators of transcription) bind to these phosphorylated tyrosine residues. Specific STATs (see Table 12-2) play essential roles in the signaling pathways of a wide variety of cytokines. The binding of STATs to receptor subunits is mediated by the joining of the SH2 domain on the STAT with the docking site created by the JAK-mediated phosphorylation of a particular tyrosine on receptor subunits. a73 After undergoing JAK-mediated phosphorylation, STAT transcription factors translocate from receptor docking sites at the membrane to the nucleus, where they initiate the transcription of specific genes. While docked to receptor subunits, STATs undergo JAK-catalyzed phosphorylation of a key tyrosine. This is followed by the dissociation of the STATs from the receptor subunits and their dimerization. The STAT dimers then translocate into the nucleus and induce the expression of genes containing appropriate regulatory sequences in their promoter regions. In addition to IFN-H9253, a number of other class I and class II ligands have been shown to cause dimerization of their re- ceptors. An important element of cytokine specificity derives from the exquisite specificity of the match between cytokines and their receptors. Another aspect of cytokine specificity is that each particular cytokine (or group of redundant cyto- kines) induces transcription of a specific subset of genes in a given cell type; the resulting gene products then mediate the various effects typical of that cytokine. The specificity of cytokine effects is then traceable to three factors. First, par- ticular cytokine receptors start particular JAK-STAT path- ways. Second, the transcriptional activity of activated STATs 286 PART III Immune Effector Mechanisms α αβ β Dimerization of receptor Activation of JAK family tyrosine kinases, phosphorylation of receptor Tyrosine phosphorylation of STAT by JAK kinase Dimerization of STAT DNA Specific gene transcription P P P P P P P P P P STAT SH 2 JAK Cytokine FIGURE 12-10 General model of signal transduction mediated by most class I and class II cytokine receptors. Binding of a cytokine in- duces dimerization of the receptor subunits, which leads to the activa- tion of receptor-subunit-associated JAK tyrosine kinases by reciprocal phosphorylation. Subsequently, the activated JAKs phosphorylate vari- ous tyrosine residues, resulting in the creation of docking sites for STATs on the receptor and the activation of the one or more STAT tran- scription factors. The phosphorylated STATs dimerize and translocate to the nucleus, where they activate transcription of specific genes. FIGURE 12-11 The complex between IFN-H9253 and the ligand-binding chains of its receptor. This model is based on the x-ray crystallo- graphic analysis of a crystalline complex of interferon-H9253 (violet and blue) bound to ligand-binding H9251 chains of the receptor (green and yellow). Note that IFN-H9253 is shown in its native dimeric form; each member of the dimer engages the H9251 chain of an IFN-H9253 receptor, thereby bringing about receptor dimerization and signal transduc- tion. [From M. R. Walter et al., 1995, Nature 376:230, courtesy M. Walter, University of Alabama.] is specific because a particular STAT homodimer or het- erodimer will only recognize certain sequence motifs and thus can interact only with the promoters of certain genes. Third, only those target genes whose expression is permitted by a particular cell type can be activated within that variety of cell. That is, in any given cell type only a subset of the potential target genes of a particular STAT may be permitted expression. For example, IL-4 induces one set of genes in T cells, another in B cells, and yet a third in eosinophils. Cytokine Antagonists A number of proteins that inhibit the biological activity of cytokines have been reported. These proteins act in one of two ways: either they bind directly to a cytokine receptor but fail to activate the cell, or they bind directly to a cytokine, inhibiting its activity. The best-characterized inhibitor is the IL-1 receptor antagonist (IL-1Ra), which binds to the IL-1 receptor but has no activity. Binding of IL-1Ra to the IL-1 receptor blocks binding of both IL-1H9251 and IL-1H9252,thus accounting for its antagonistic properties. Production of IL-1Ra has been thought by some to play a role in regulating the intensity of the inflammatory response. It has been cloned and is currently being investigated as a potential treat- ment for chronic inflammatory diseases. Cytokine inhibitors are found in the bloodstream and extracellular fluid. These soluble antagonists arise from enzy- matic cleavage of the extracellular domain of cytokine recep- tors. Among the soluble cytokine receptors that have been de- tected are those for IL-2, -4, -6, and -7, IFN-H9253 and -H9251,TNF-H9252, and LIF. Of these, the soluble IL-2 receptor (sIL-2R), which is released in chronic T-cell activation, is the best characterized. A segment containing the amino-terminal 192 amino acids of the H9251 subunit is released by proteolytic cleavage, forming a 45-kDa soluble IL-2 receptor. The shed receptor can bind IL-2 and prevent its interaction with the membrane-bound IL-2 receptor. The presence of sIL-2R has been used as a clin- ical marker of chronic T-cell activation and is observed in a number of diseases, including autoimmunity, transplant re- jection, and AIDS. Some viruses also produce cytokine-binding proteins or cytokine mimics. The evolution of such anti-cytokine strate- gies by microbial pathogens is good biological evidence of the importance of cytokines in organizing and promoting ef- fective anti-microbial immune responses. The poxviruses, for example, have been shown to encode a soluble TNF-binding protein and a soluble IL-1–binding protein. Since both TNF and IL-1 exhibit a broad spectrum of activities in the inflam- matory response, these soluble cytokine-binding proteins may prohibit or diminish the inflammatory effects of the cytokines, thereby conferring upon the virus a selective ad- vantage. Epstein-Barr virus produces an IL-10–like molecule (viral IL-10 or vIL-10) that binds to the IL-10 receptor and, like cellular IL-10, suppresses T H 1-type cell-mediated re- sponses (see the next section), which are effective against many intracellular parasites such as viruses. Molecules pro- duced by viruses that mimic cytokines allow the virus to manipulate the immune response in ways that aid the sur- vival of the pathogen. This is an interesting and powerful modification some viruses have undergone in their continu- ing struggle to overcome the formidable barrier of host immunity. Table 12-3 lists a number of viral products that mimic cytokines or their receptors. Cytokines CHAPTER 12 287 TABLE 12-2 STAT and JAK interaction with selected cytokine receptors during signal transduction Cytokine receptor JAK STAT IFN-H9253 JAK1 and JAK2 Stat1 IFN-H9251/H9252 JAK1 and Tyk-2 Stat2 IL-2 JAK1 and JAK3 Stat5 IL-3 JAK2 Stat5 IL-4 JAK1 and JAK3 Stat6 IL-6 JAK1 (and sometimes others) Stat3 IL-10 JAK1 and Tyk-2* Stat3 IL-12 JAK2 and Tyk-2* Stat4 *Despite its name, Tyk-2 is also a Janus kinase. SOURCE: Adapted from E. A. Bach, M. Aguet, and R. D. Schreiber, 1997, Annu. Rev. Immun. 15:563. TABLE 12-3 Viral mimics of cytokine and cytokine receptors Virus Product Leporipoxvirus Soluble IFN-H9253 receptor (a myxoma virus) Several poxviruses Soluble IFN-H9253 receptor Vaccinia, smallpox virus Soluble IL-1 H9252 receptor Epstein-Barr IL-10 homolog Human herpesvirus-8 IL-6 homolog; also homologs of the chemokines MIP-I and MIP-II Cytomegalovirus Three different chemokine receptor homologs, one of which binds three different soluble chemokines (RANTES, MCP-1, and MIP-1H9251) Cytokine Secretion by T H 1 and T H 2 Subsets The immune response to a particular pathogen must induce an appropriate set of effector functions that can eliminate the disease agent or its toxic products from the host. For exam- ple, the neutralization of a soluble bacterial toxin requires antibodies, whereas the response to an intracellular virus or to a bacterial cell requires cell-mediated cytotoxicity or delayed-type hypersensitivity. A large body of evidence im- plicates differences in cytokine-secretion patterns among T H -cell subsets as determinants of the type of immune response made to a particular antigenic challenge. CD4 + T H cells exert most of their helper functions through secreted cytokines, which either act on the cells that produce them in an autocrine fashion or modulate the re- sponses of other cells through paracrine pathways. Although CD8 + CTLs also secrete cytokines, their array of cytokines generally is more restricted than that of CD4 + T H cells. As briefly discussed in Chapter 10, two CD4 + T H -cell subpopu- lations designated T H 1 and T H 2, can be distinguished in vitro by the cytokines they secrete. Both subsets secrete IL-3 and GM-CSF but differ in the other cytokines they produce (Table 12-4). T H 1 and T H 2 cells are characterized by the fol- lowing functional differences: a73 The T H 1 subset is responsible for many cell-mediated functions (e.g., delayed-type hypersensitivity and activation of T C cells) and for the production of opsonization-promoting IgG antibodies (i.e. antibodies that bind to the high-affinity Fc receptors of phagocytes and interact with the complement system). This subset is also associated with the promotion of excessive inflammation and tissue injury. a73 The T H 2 subset stimulates eosinophil activation and differentiation, provides help to B cells, and promotes the production of relatively large amounts of IgM, IgE, and noncomplement-activating IgG isotypes. The T H 2 subset also supports allergic reactions. The differences in the cytokines secreted by T H 1 and T H 2 cells determine the different biological functions of these two subsets. A defining cytokine of the T H 1 subset, IFN-H9253,acti- vates macrophages, stimulating these cells to increase micro- bicidal activity, up-regulate the level of class II MHC, and secrete cytokines such as IL-12, which induces T H cells to dif- ferentiate into the T H 1 subset. IFN-H9253 secretion by T H 1 cells also induces antibody-class switching to IgG classes (such as IgG2a in the mouse) that support phagocytosis and fixation of complement. TNF-H9252 and IFN-H9253 are cytokines that medi- ate inflammation, and it is their secretion that accounts for the association of T H 1 cells with inflammatory phenomena such as delayed hypersensitivity (Chapter 16). T H 1 cells pro- duce IL-2 and IFN-H9253 cytokines that promote the differentia- tion of fully cytotoxic T C cells from CD8 + precursors. This pattern of cytokine production makes the T H 1 subset partic- ularly suited to respond to viral infections and intracellular pathogens. Finally, IFN-H9253 inhibits the expansion of the T H 2 population. The secretion of IL-4 and IL-5 by cells of the T H 2 subset in- duces production of IgE and supports eosinophil-mediated attack on helminth (roundworm) infections. IL-4 promotes a pattern of class switching that produces IgG that does not activate the complement pathway (IgG1 in mice, for exam- ple). IL-4 also increases the extent to which B cells switch from IgM to IgE. This effect on IgE production meshes with eosinophil differentiation and activation by IL-5, because eosinophils are richly endowed with FcH9280 receptors, which bind IgE. Typically, roundworm infections induce T H 2 re- sponses and evoke anti-roundworm IgE antibody. The anti- body bound to the worm binds to the Fc receptors of eosinophils, thus forming an antigen-specific bridge between the worm and the eosinophils. The attack of the eosinophil on the worm is triggered by crosslinking of the FcH9280-bound IgE. Despite these beneficial actions of IgE, it is also the Ig 288 PART III Immune Effector Mechanisms TABLE 12-4 Cytokine secretion and principal functions of mouse TH1 and TH2 subsets Cytokine/function T H 1 T H 2 CYTOKINE SECRETION IL-2 +– IFN-H9253 ++ – TNF-H9252 ++ – GM-CSF ++ + IL-3 ++ ++ IL-4 –++ IL-5 IL-10 –++ IL-13 FUNCTIONS Help for total antibody production + ++ Help for IgE production – ++ Help for IgG2a production ++ + Eosinophil and mast-cell production – ++ Macrophage activation ++ – Delayed-type hypersensitivity ++ – T C -cell activation ++ – SOURCE: Adapted from F. Powrie and R. L. Coffman, 1993, Immunol. Today 14:270. class responsible for allergy. Finally, IL-4 and IL-10 suppress the expansion of T H 1 cell populations. Because the T H 1 and T H 2 subsets were originally identi- fied in long-term in vitro cultures of cloned T-cell lines, some researchers doubted that they represented true in vivo sub- populations. They suggested instead that these subsets might represent different maturational stages of a single lineage. Also, the initial failure to locate either subset in humans led some to believe that T H 1, T H 2, and other subsets of T helper cells did not occur in this species. Further research corrected these views. In many in vivo systems, the full commitment of populations of T cells to either the T H 1 or T H 2 phenotype often signals the endpoint of a chronic infection or allergy. Hence it was difficult to find clear T H 1 or T H 2 subsets in studies employing healthy human subjects, who would not be at this stage of a response. Experiments with transgenic mice demonstrated conclusively that T H 1 and T H 2 cells arise independently. Furthermore, it was possible to demonstrate T H 1 or T H 2 populations in T cells isolated from humans dur- ing chronic infectious disease or chronic episodes of allergy. It is also important to emphasize that many helper T cells do not show either a T H 1 or a T H 2 profile; individual cells have shown striking heterogeneity in the T H -cell population. One of the best described of these is the T H 0 subset, which secretes IL-2, IL-4, IL-5, IFN-H9253, and IL-10, as well as IL-3 and GM-CSF. Numerous reports of studies in both mice and humans now document that the in vivo outcome of the immune response can be critically influenced by the relative levels of T H 1-like or T H 2-like activity. Typically, the T H 1 profile of cytokines is higher in response to intracellular pathogens, and the T H 2 profile is higher in allergic diseases and hel- minthic infections. The Development of T H 1 and T H 2 Subsets Is Determined by the Cytokine Environment The cytokine environment in which antigen-primed T H cells differentiate determines the subset that develops (Figure 12-12). In particular, IL-4 is essential for the development of a T H 2 response, and IFN-H9253, IL-12, and IL-18 all are impor- tant in the physiology of the development of T H 1 cells. The source of IL-12, one of the key mediatiors of T H 1 differentia- tion, is typically macrophages or dendritic cells activated by an encounter with intracellular bacteria, with bacterial products such as LPS, or with a number of other intracellular parasites. T H 1 development is also critically dependent on IFN-H9253, which induces a number of changes, including the up-regulation of IL-12 production by macrophages and den- dritic cells, and the activation of the IL-12 receptor on acti- vated T cells, which it accomplishes by up-regulating expres- sion of the H9252 chain of the IL-12 receptor. At the beginning of an immune response, IFN-H9253 is generated by stimulation of T cells and can also come from activated NK cells.Yet another cytokine, IL-18, promotes proliferation and IFN-H9253 produc- tion by both developing and fully differentiated T H 1 cells and by NK cells. So a regulatory network of cytokines posi- tively controls the generation of T H 1 cells. The critical role played by each of these cytokines and their receptors has been demonstrated in a series of experiments in which either the cytokine or its receptor has been knocked out. Mice in which the genes for any of these critical components have been knocked out fail to generate populations of T H 1 cells. Just as T H 1 cells require IL-12 and IFN-H9253, the generation of T H 2 cells depends critically on IL-4. Exposing naive helper cells to IL-4 at the beginning of an immune response causes them to differentiate into T H 2 cells. In fact, this influence of IL-4 is predominant in directing T H cells to the T H 2 route. Provided a threshold level of IL-4, T H 2 development is Cytokines CHAPTER 12 289 Macrophage or dendritic cell Positive feedback Positive feedback IL-2 Naive CD4 + T cell T H 2 T H 1 ?Delayed-type hypersensitivity ?Macrophage activation ?Certain opsonic or complement-fixing IgGs IL-4 , IL-5, IL-10, IL-13 ?Eosinophil activation ?IgE and some other antibody classes IL-4 IL-12 IFN-δ , IL-2, TNF-β ? ? FIGURE 12-12 Cytokine-mediated generation and cross regula- tion of T H subsets. Antigen-activated naive CD4 + T cell produces IL-2 and proliferates. If it proliferates in an IL-12 dominated environment, it generates a population of T H 1 cells that secretes a characteristic profile of cytokines including interferon H9253. A positive feedback loop is established when IFN-H9253 secreted by the expanding T H 1 population stimulates dendritic cells or macrophages to produce more IL-12. If the environment is dominated by IL-4, a T H 2 population emerges and secretes a profile of cytokines that promotes eosinophil activa- tion and the synthesis of certain antibody classes. Key cytokines pro- duced by each subset positively regulate the subset that produces it and negatively regulate the other subset. [Adapted from J. Rengarajan, S. Szabo, and L. Glimcher, 2000, Immunology Today 21:479.] greatly favored over T H 1 even if IL-12 is present. The critical role of signals from IL-4 in T H 2 development is shown by the observation that knocking out the gene that encodes IL-4 prevents the development of this T-cell subset. Additional evidence supporting the central role of IL-4 comes from an experiment that interrupted the IL-4 signal-transduction pathway. Like so many other cytokines, IL-4 uses a pathway that involves JAK and STAT proteins. The Stat6 transcription factor is the one activated in signaling by IL-4. Consequently, in mice in which the gene for Stat6 has been disrupted (Stat6 knockouts), IL-4 mediated processes are severely inhibited or absent. The observation that Stat6 knockout mice have very few T H 2 cells confirms the importance of IL-4 for the differ- entiation of this subset. Cytokine Profiles Are Cross-Regulated The critical cytokines produced by T H 1 and T H 2 subsets have two characteristic effects on subset development. First, they promote the growth of the subset that produces them; sec- ond, they inhibit the development and activity of the oppo- site subset, an effect known as cross-regulation, (see Figure 12-12). For instance, IFN-H9253 (secreted by the T H 1 subset) preferentially inhibits proliferation of the T H 2 subset, and IL-4 and IL-10 (secreted by the T H 2 subset) down-regulate secretion of IL-12, one of the critical cytokines for T H 1 dif- ferentiation, by both macrophages and dendritic cells. Simi- larly, these cytokines have opposing effects on target cells other than T H subsets. IFN-H9253 secreted by the T H 1 subset pro- motes IgG2a production by B cells but inhibits IgG1 and IgE production. On the other hand, IL-4 secreted by the T H 2 sub- set promotes production of IgG1 and IgE and suppresses production of IgG2a. The phenomenon of cross-regulation explains the observation that there is often an inverse rela- tionship between antibody production and cell-mediated immunity; that is, when antibody production is high, cell- mediated immunity is low, and vice versa. Furthermore, recent research has shown that IL-4 and IFN-H9253 make mem- bers of the T-cell subset that releases them less responsive to the cytokine that directs differentiation of the other T-cell subset. Thus, IL-4 enhances T H 2 cell development by making T H cells less susceptible to the cytokine signals that cause these cells to enter a differentiation pathway that would lead to T H 1 development. On the other hand, as explained below, IFN-H9253 up-regulates the expression of a key regulatory mole- cule that favors the differentiation and activity of T H 1 cells. Recent work has given insight into the molecular basis for the cytokine-mediated cross-regulation by which one subset promotes its own expansion and development while inhibiting the development of the opposite subset. Two tran- scription factors, T-Bet and GATA-3, are key elements in de- termining subset commitment and cross-regulation. The ex- pression of T-Bet drives cells to differentiate into T H 1 cells and suppresses their differentiation along the T H 2 pathway. Expression of GATA-3 does the opposite, promoting the de- velopment of naive T cells into T H 2 cells while suppressing their differentiation into T H 1 cells. As shown in Figure 12-13, the expression of T-Bet versus GATA-3 is determined by the cytokines IFN-H9253and IL-4. In the presence of IFN-H9253, T cells up- regulate the expression of T-Bet and down-regulate GATA-3. This IFN-H9253 receptor/Stat1–dependent process shifts the cyto- kine profile to the production of IFN-H9253, the signature cytokine of T H 1 cells, and other cytokines typical of the T H 1 set. On the other hand, in a process that involves the IL-4 receptor and Stat6, IL-4 induces the cell to produce IL-4 and other T H 2 cytokines. Further study has revealed that the up-regulation of T-Bet represses the expression of GATA-3. Similarly, expression of GATA-3 down-regulates T-Bet. Consequently, cytokine signals that induce one of these transcription fac- tors set in motion a chain of events that repress the other. At the intracellular level, the differentiation of a T cell along the T H 1 pathway, prevents its development of T H 2 characteris- tics and vice versa. 290 PART III Immune Effector Mechanisms APC TCR Promotes T H 1 Promotes T H 2 Stat1 T-Bet IFN-δ IL-4 IL-5 Stat6 GATA-3 IL-4IFN-δ ? ? ? + + + ?? FIGURE 12-13 Cross-regulation at the intracellular level. Signals through the TCR and cytokine receptors determine whether the cell will produce the T H 1-promoting transcription factor, T-Bet, or the T H 2-promoting transcription factor, GATA-3. Experimental evidence supports a model in which exposure of cells bearing receptors for IFN-H9253 to IFN-H9253 induces the formation of T-Bet, which up-regulates the synthesis of IFN-H9253 and represses the expression of GATA-3. Ex- posure of IL-4 R–bearing cells to IL-4 induces the formation of GATA- 3, which up-regulates the synthesis of IL-4 and IL-5 but represses the expression of T-Bet. [Adapted from J. Rengarajan, S. Szabo, and L. Glimcher, 2000, Immunology Today 21:479.] The cross-regulation of T H 1 cells by IL-10 secreted from T H 2 cells is not a direct inhibition of the T H 1 cells; instead, IL-10 acts on monocytes and macrophages, interfering with their ability to activate the T H 1 subset. This interference is thought to result from the demonstrated ability of IL-10 to down-regulate the expression of class II MHC molecules on these antigen-presenting cells. IL-10 has other potent immu- nosuppressant effects on the monocyte-macrophage lineage, such as suppressing the production of nitric oxide and other bactericidal metabolites involved in the destruction of path- ogens, and also suppressing the production of various inflam- matory mediators (e.g., IL-1, IL-6, IL-8, GM-CSF, G-CSF, and TNF-H9253). These suppressive effects on the macrophage serve to further diminish the biologic consequences of T H 1 activation. The T H 1/T H 2 Balance Determines Disease Outcomes The progression of some diseases may depend on the balance between the T H 1 and T H 2 subsets. In humans, a well-studied example of this phenomenon is leprosy, which is caused by Mycobacterium leprae, an intracellular pathogen that can sur- vive within the phagosomes of macrophages. Leprosy is not a single clinical entity; rather, the disease presents as a spec- trum of clinical responses, with two major forms of disease, tuberculoid and lepromatous, at each end of the spectrum. In tuberculoid leprosy, a cell-mediated immune response forms granulomas, resulting in the destruction of most of the mycobacteria, so that only a few organisms remain in the tissues. Although skin and peripheral nerves are damaged, tuberculoid leprosy progresses slowly and patients usually survive. In lepromatous leprosy, the cell-mediated response is depressed and, instead, humoral antibodies are formed, sometimes resulting in hypergammaglobulinemia. The myco- bacteria are widely disseminated in macrophages, often reach- ing numbers as high as 10 10 per gram of tissue. Lepromatous leprosy progresses into disseminated infection of the bone and cartilage with extensive nerve damage. The development of lepromatous or tuberculoid leprosy depends on the balance of T H 1 and T H 2 cells (Figure 12-14). In tuberculoid leprosy, the immune response is characterized by a T H 1-type response with delayed-type hypersensitivity and a cytokine profile consisting of high levels of IL-2, IFN-H9253, and TNF-H9252. In lepromatous leprosy, there is a T H 2-type im- mune response, with high levels of IL-4, IL-5, and IL-10. This cytokine profile explains the diminished cell-mediated im- munity and increased production of serum antibody in lep- romatous leprosy. There is also evidence for changes in T H -subset activity in AIDS. Early in the disease, T H 1 activity is high, but as AIDS progresses, some workers have suggested, there may be a shift from a T H 1-like to a T H 2-like response. In addition, some pathogens may influence the activity of the T H subsets. The Epstein-Barr virus, for instance, produces vIL-10, which has IL-10–like activity and, like cellular IL-10, tends to suppress T H 1 activity by cross-regulation. Some researchers have spec- ulated that vIL-10 may reduce the cell-mediated response to the Epstein-Barr virus, thus conferring a survival advantage on the virus. Cytokine-Related Diseases Defects in the complex regulatory networks governing the expression of cytokines and cytokine receptors have been implicated in a number of diseases. This section describes several diseases resulting from overexpression or underex- pression of cytokines or cytokine receptors. Bacterial Septic Shock Is Common and Potentially Lethal The role of cytokine overproduction in pathogenesis can be illustrated by bacterial septic shock. This condition may develop a few hours after infection by certain gram-negative bacteria, including E. coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Enterobacter aerogenes, and Neisseria meningitidis. The symptoms of bacterial septic shock, which is often fatal, include a drop in blood pressure, fever, diarrhea, and wide- spread blood clotting in various organs. This condition afflicts about 500,000 Americans annually and causes more than 70,000 deaths. The annual cost for treating bacterial septic shock is estimated to be in excess of $5 billion. Bacterial septic shock apparently develops because bacte- rial cell-wall endotoxins stimulate macrophages to over- produce IL-1 and TNF-H9251 to levels that cause septic shock. In Cytokines CHAPTER 12 291 FIGURE 12-14 Correlation between type of leprosy and relative T H 1 or T H 2 activity. Messenger RNA isolated from lesions from tu- berculoid and lepromatous leprosy patients was analyzed by South- ern blotting using the cytokine probes indicated. Cytokines produced by T H 1 cells predominate in the tuberculoid patients, while cytokines produced by T H 2 cells predominate in the lepromatous patients. [From P. A. Sieling and R. L. Modlin, 1994, Immunobiology 191: 378.] TH1 activity TH2 activity Tuberculoid Lepromatous Tuberculoid Lepromatous IL-2 IFN-H9253 IFN-H9252 IL-4 IL-5 IL-10 one study, for example, higher levels of TNF-H9251 were found in patients who died of meningitis than in those who recovered. Furthermore, a condition resembling bacterial septic shock can be produced by injecting mice with recombinant TNF-H9251 in the absence of gram-negative bacterial infection. Several studies offer some hope that neutralization of TNF-H9251 or IL-1 activity with monoclonal antibodies or antagonists may pre- vent this fatal shock from developing in these bacterial infec- tions. In one study, monoclonal antibody to TNF-H9251 protected animals from endotoxin-induced shock. Another study has shown that injection of a recombinant IL-1 receptor antago- nist (IL-1Ra), which prevents binding of IL-1 to the IL-1 receptor, resulted in a three-fold reduction in mortality. It is hoped that these experimental results will lead to clinically useful products for the treatment of bacterial septic shock in humans. Bacterial Toxic Shock Is Caused by Superantigens A variety of microorganisms produce toxins that act as super- antigens. As Chapter 10 described, superantigens bind simul- taneously to a class II MHC molecule and to the V H9252 domain of the T-cell receptor, activating all T cells bearing a particular V H9252 domain (Figure 10-16). Because of their unique binding abil- ity, superantigens can activate large numbers of T cells irre- spective of their antigenic specificity. Although less than 0.01% of T cells respond to a given conventional antigen, between 5% and 25% of T cells can respond to a given superantigen. The large proportion of T cells responsive to a particular superantigen results from the limited number of TCR V H9252 genes carried in the germ line. Mice, for example, have about 20 V H9252 genes. Assuming that each V H9252 gene is expressed with equal frequency, then each superantigen would be expected to interact with 1 in 20 T cells, or 5% of the total T-cell population. A number of bacterial superantigens have been implicated as the causative agent of several diseases such as bacterial toxic shock and food poisoning. Included among these bacterial superantigens are several enterotoxins, exfoliating toxins, and toxic-shock syndrome toxin (TSST1) from Staphylococcus aureus; pyrogenic exotoxins from Streptococcus pyrogenes; and Mycoplasma arthritidis supernatant (MAS). The large number of T cells activated by these superantigens results in excessive production of cytokines. The toxic-shock syndrome toxin, for example, has been shown to induce extremely high levels of TNF and IL-1. As in bacterial septic shock, these elevated concentrations of cytokines can induce systemic reactions that include fever, widespread blood clotting, and shock. Cytokine Activity Is Implicated in Lymphoid and Myeloid Cancers Abnormalities in the production of cytokines or their recep- tors have been associated with some types of cancer. For ex- ample, abnormally high levels of IL-6 are secreted by cardiac myxoma cells (a benign heart tumor), myeloma and plasma- cytoma cells, and cervical and bladder cancer cells. In myeloma cells, IL-6 appears to operate in an autocrine man- ner to stimulate cell proliferation. When monoclonal anti- bodies to IL-6 are added to in vitro cultures of myeloma cells, their growth is inhibited. In addition, transgenic mice that express high levels of IL-6 have been found to exhibit a mas- sive, fatal plasma-cell proliferation, called plasmacytosis. Although these plasma cells are not malignant, the high rate of plasma-cell proliferation possibly contributes to the devel- opment of cancer. Chagas’ Disease Is Caused by a Parasite The protozoan Trypanosoma cruzi is the causative agent of Chagas’ disease, which is characterized by severe immune suppression. The ability of T. cruzi to mediate immune sup- pression can be observed by culturing peripheral-blood T cells in the presence and in the absence of T. cruzi and then evaluating their immune reactivity. Antigen, mitogen, or anti-CD3 monoclonal antibody normally can activate peri- pheral T cells, but in the presence of T. cruzi, T cells are not activated by any of these agents. The defect in these lympho- cytes has been traced to a dramatic reduction in the expres- sion of the 55-kDa H9251 subunit of the IL-2 receptor. As noted earlier, the high-affinity IL-2 receptor contains H9251, H9252, and H9253 subunits. The H9251 subunit is specific for cytokine binding (see Figure 12-9). Co-culturing of T cells with T. cruzi and sub- sequent staining with fluorescein-labeled anti-TAC, which binds to the H9251 subunit, revealed a 90% decrease in the level of the H9251 subunit. Although the mechanism by which T. cruzi suppresses expression of the H9251 subunit is still unknown, the suppression can be induced across a filter that prevents contact between the lymphocytes and protozoa. This finding suggests that a diffusible factor mediates suppression. Such a factor, once isolated, might have numerous clinical applications for regu- lating the level of activated T cells in leukemias and autoim- mune diseases. Therapeutic Uses of Cytokines and Their Receptors The availability of purified cloned cytokines and soluble cytokine receptors offers the prospect of specific clinical therapies to modulate the immune response. A few cyto- kines—notably, interferons (see Clinical Focus)—and colony- stimulating factors, such as GM-CSF, have proven to be ther- apeutically useful. However, despite the promise of cytokines as powerful mediators of immune and other biological 292 PART III Immune Effector Mechanisms Cytokines CHAPTER 12 293 responses, not many have made their way into clinical prac- tice. A number of factors are likely to raise difficulties in adapting cytokines for safe and effective routine medical use. One of these is the need to maintain effective dose levels over a clinically significant period of time. During an immune response, interacting cells produce sufficiently high concen- trations of cytokines in the vicinity of target cells, but achiev- ing such local concentrations when cytokines must be ad- ministered systemically for clinical treatment is difficult. In addition, cytokines often have a very short half-life, so that continuous administration may be required. For example, recombinant human IL-2 has a half-life of only 7–10 min when administered intravenously. Finally, cytokines are ex- tremely potent biological response modifiers and they can cause unpredictable and undesirable side effects. The side ef- fects from administration of recombinant IL-2, for instance, range from mild (e.g., fever, chills, diarrhea, and weight gain) to serious, such as anemia, thrombocytopenia, shock, respi- ratory distress, and coma. Despite these difficulties, the pro- mise of cytokines for clinical medicine is great and efforts to develop safe and effective cytokine-related strategies con- tinue, particularly in areas such as inflammation, cancer therapy, and modification of the immune response during organ transplantation, infectious disease, and allergy. Some specific examples of various approaches being ex- plored include cytokine receptor blockade and the use of cytokine analogs and cytokine-toxin conjugates. For instance, proliferation of activated T H cells and activation of T C cells can be blocked by anti-TAC, a monoclonal antibody that binds to the H9251 subunit of the high-affinity IL-2 receptor (Fig- ure 12-15a, left panel). Administration of anti-TAC has pro- longed the survival of heart transplants in rats. Similar results have been obtained with IL-2 analogs that retain their ability to bind the IL-2 receptor but have lost their biological activity (Figure 12-15a, right panel). Such analogs have been produced by site-directed mutagenesis of cloned IL-2 genes. Finally, cytokines conjugated to various toxins (e.g., the H9252 chain of diphtheria toxin) have been shown to diminish re- jection of kidney and heart transplants in animals. Such con- jugates containing IL-2 selectively bind to and kill activated T H cells (Figure 12-15b). Cytokines in Hematopoiesis Early work in Australia and Israel demonstrated that soluble factors could support the growth and differentiation of red and white blood cells. The first of these soluble factors to be IL-2R T C cell T H cell Antibodies to IL-2R IL-2 IL-2 analogs IL-2R T C cell T H cell IL-2R Toxin conjugated to IL-2 (b) Destruction of activated T H cells T H cell (a) Suppression of T H -cell proliferation and T C -cell activation FIGURE 12-15 Experimental cytokine-related therapeutic agents offer the prospect of selectively modulating the immune response. (a) The anti-IL-2R monoclonal antibody binds to the cytokine recep- tor (IL-2R) on the cell surface, thereby preventing interaction of the cytokine with its receptor. (b) Conjugation of a toxin with a cytokine results in destruction of cells expressing the cytokine receptor. characterized, erythropoietin, was isolated from the urine of anemic patients and shown to support the develop- ment of red blood cells. Subsequently, many cytokines have been shown to play essential roles in hematopoiesis (see Table 12-5). During hematopoiesis, cytokines act as develop- mental signals that direct commitment of progenitor cells into and through particular lineages. As shown in Figure 12-16, a myeloid progenitor in the presence erythropoietin 294 PART III Immune Effector Mechanisms a73 IFN-H9251 (also known by its trade names Roferon and Intron-A) has been used for the treatment of hepatitis C and hepatitis B. It has also found a number of different applications in cancer therapy. A type of B-cell leukemia known as hairy-cell leukemia (because the cells are covered with fine, hairlike cytoplasmic projections) responds well to IFN-H9251. Chronic myelogenous leukemia, a disease characterized by increased numbers of granulocytes, begins with a slowly developing chronic phase that changes to an accelerated phase and terminates in a blast phase, which is usually resistant to treatment. IFN-H9251 is an effective treatment for this leukemia in the chronic phase (70% response rates have been reported) and some patients (as many as 20% in some studies) undergo complete remission. Kaposi’s sarcoma, the cancer most often seen in American AIDS patients, also responds to treatment with IFN-H9251, and there are reports of a trend toward longer survival and fewer opportunistic infections in patients treated with this agent. IFN-H9253 has also been used, with varying degrees of success, to treat a variety of malignancies that include non- Hodgkin’s lymphoma, cutaneous T-cell lymphoma, and multiple myeloma. Most of the effects mentioned above have been obtained in clinical studies that used IFN-H9251 alone. Future clinical trials in which IFN-H9251 is used in combinations with other agents may improve the effectiveness of this interferon in cancer therapy. a73 IFN-H9252 has emerged as the first drug capable of producing clinical improvement in multiple sclerosis (MS). Young adults are the primary target of this autoimmune neurologic disease, in which nerves in the central nervous system (CNS) undergo demyelination. This results in progressive neurologic dysfunction, which leads to significant and, in many cases, severe disability. This disease is often characterized by periods of nonprogression and remission alternating with periods of relapse. Treatment with IFN-H9252 provides longer periods of remission and reduces the severity of relapses. Furthermore, magnetic-resonance- imaging studies of CNS damage in treated and untreated patients revealed that MS-induced damage was less in a group of IFN-H9252-treated patients than in untreated ones. a73 IFN-H9253 has found application in the clinic as an agent for the treatment of chronic granulomatous disease (CGD). This disease is hereditary and quite rare. Its central feature is a serious impairment of the ability of phagocytic cells to kill ingested microbes. Patients with CGD are beset with recurring infections by a number of bacteria (Staphylococcus aureus, Klebsiella, Pseudomonas, and others) and fungi such as Aspergillus and Candida. Before interferon therapy, standard treatment for the disease was attempts to avoid infection, aggressive administration of Interferons are an extraordinary group of proteins whose antiviral activity led to their discovery almost 50 years ago. Subsequent studies showed that interferons have other effects, including the capacity to induce cell differentiation, to inhibit proliferation by some cell types, to inhibit angiogene- sis, and to function in various immu- noregulatory roles. Their effects on the immune system are important and dra- matic. Interferons induce increases in the expression of class I and class II MHC molecules, and augment NK-cell activity. Increased class I expression increases the display of antigen to CD8 + cells, a class that includes most of the T C popu- lation. This enhanced display of antigen not only makes the antigen-presenting cells more effective in inducing cytotoxic T-cell populations, it also makes them better targets for attack by T C cells. In addition to up-regulating class I MHC expression of many cell types, IFN-H9253 in- creases the expression of class II MHC molecules on such antigen-presenting cells as macrophages and dendritic cells. This makes them better presenters of antigen to T H cells. IFN-H9253 is also a potent inducer of macrophage activation and general promoter of inflammatory re- sponses. Cloning of the genes that en- code all three types of interferon, IFN-H9251, IFN-H9252, and IFN-H9253, has made it possible for the biotechnology industry to produce large amounts of all of these interferons at costs that make their clinical use prac- tical. Some clinical uses of each type of interferon are described here: CLINICAL FOCUS Therapy with Interferons would proceed down a pathway that leads to the production of erythrocytes; suitable concentrations of a group of cyto- kines including IL-3, GM-CSF, IL-1, and IL-6 will cause it to enter differentiation pathways that lead to the generation of monocytes, neutrophils, and other leukocytes of the myeloid group. The participation of leukocytes in immune responses often results in their death and removal. However, both adap- tive and innate immune responses generate cytokines that Cytokines CHAPTER 12 295 osteopetrosis, (not osteoporosis) a life-threatening congenital disorder characterized by overgrowth of bone which results in blindness and deafness. Another problem presented by this disease is that the buildup of bone reduces the amount of space available for bone marrow and the decrease in hematopoiesis results in fewer red blood cells and anemia. The decreased generation of white blood cells causes an increased susceptibility to infection. The use of interferons in clinical prac- tice is likely to expand as more is learned about their effects in combination with other therapeutic agents. Although, in common with other cytokines, interfer- ons are powerful modifiers of biological responses, fortunately, the side effects accompanying their use are much milder. Typical side effects include flu- like symptoms, such as headache, fever, chills, and fatigue. These symptoms can largely be managed with acetaminophen (Tylenol) and diminish in intensity dur- ing continued treatment. Although inter- feron toxicity is usually not severe, seri- ous manifestations such as anemia and depressed platelet and white-blood-cell counts have been seen. antibiotics, and surgical drainage of abscesses. A failure to generate microbicidal oxidants (H 2 O 2 , superoxide, and others) is the basis of CGD, and the administration of IFN-H9253 significantly reverses this defect. Therapy of CGD patients with IFN-H9253 significantly reduces the incidence of infections. Also, the infections that are contracted are less severe and the average number of days spent by patients in the hospital goes down. a73 IFN-H9253 has also been shown to be effective in the treatment of Cytokine-Based Therapies In Clinical Use Agent Nature of agent Clinical application Enbrel Chimeric TNF-receptor/IgG Rheumatoid arthritis constant region Remicade Monoclonal antibody Rheumatoid arthritis against TNF-H9251 receptor Interferon H9251-2a Antiviral cytokine Hepatitis B Hairy cell leukemia Kaposi’s sarcoma Interferon H9251-2b Antiviral cytokine Hepatitis C Melanoma Interferon H9252 Antiviral cytokine Multiple sclerosis Actimmune Interferon H9253 Chronic granulomatous disease (CGD) Osteopetrosis Neupogen G-CSF (hematopoietic cytokine) Stimulates production of neutrophils Reduction of infection in cancer patients treated with chemotherapy Leukine GM-CSF (hematopoietic cytokine) Stimulates production of myeloid cells after bone-marrow transplantation Neumega Interleukin 11 (IL-11), a Stimulates production of platelets hematopoietic cytokine Epogen Erythopoietin (hematopoietic Stimulates red-blood-cell production cytokine) stimulate and support the production of leukocytes. The steps at which a number of cytokines participate in hema- topoiesis is shown in Figure 12-16. SUMMARY a73 Cytokines are low-molecular-weight proteins that are pro- duced and secreted by a variety of cell types. They play major roles in the induction and regulation of the cellular interactions involving cells of the immune, inflammatory and hematopoietic systems. a73 The biological activities of cytokines exhibit pleiotropy, re- dundancy, synergy, antagonism, and, in some instances, cascade induction. a73 There are over 200 different cytokines, most of which fall into one of the following families: hematopoietins, inter- ferons, chemokines, and tumor necrosis factors. a73 Cytokines act by binding to cytokine receptors, most of which can be classified as immunoglobulin superfamily receptors, class I cytokine receptors, class II cytokine recep- tors, members of the TNF receptor family, and chemokine receptors. a73 A cytokine can only act on a cell that expresses a receptor for it. The activity of particular cytokines is directed to spe- cific cells by regulation of the cell’s profile of cytokine receptors. a73 Cytokine-induced multimerization of class I and class I cyto- kine receptors activates a JAK/STAT signal-transduction pathway. a73 Antigen stimulation of T H cells in the presence of certain cytokines can lead to the generation of subpopulations of helper T cells known as T H 1 and T H 2. Each subset displays characterisic and different profiles of cytokine secretion. a73 The cytokine profile of T H 1 cells supports immune re- sponses that involve the marshalling of phagocytes, CTLs, and NK cells to eliminate intracellular pathogens. T H 2 cells produce cytokines that support production of particular immunoglobulin isotypes and IgE-mediated responses. a73 Therapies based on cytokines and cytokine receptors have entered clinical practice. References Abbas, A., K. M. Murphy, and A. Sher. 1996. Functional diversity of helper T lymphocytes. Nature 383:787. Alcami, A., and U. H. Koszinowski. 2000. Viral mechanisms of immune evasion. Immunol. Today 9:447–455. Bach, E. A., M. Aguet, and R. D. Schreiber. 1998. The IFN-H9253 receptor: a paradigm for cytokine receptor signaling. Ann. Rev. Immunol. 15:563. Darnell, J. E. Jr. 1997. STATs and gene regulation. Science 5332:1630–1635. 296 PART III Immune Effector Mechanisms TABLE 12-5 Haematopoietic cytokines Haematopoietic growth factor Sites of production Main functions Erythropoietin Kidney, liver Erythrocyte production G-CSF Endothelial cells, fibroblasts, macrophages Neutrophil production Thrombopoietin Liver, kidney Platelet production M-CSF Fibroblasts, endothelial cells, macrophages Macrophage and osteoclast production SCF/c-kit ligand Bone marrow stromal cells, constitutively Stem cell, progenitor cells survival/division; mast cell differentiation Flt-3 ligand Fibroblasts, endothelial cells Early progenitor cell expansion; pre-B cells GM-CSF T cells (T H 1 and T H 2), macrophages, Macrophage, granulocyte production; dendritic mast cells cell maturation and activation IL-3 T cells (T H 1 and T H 2), macrophages Stem cells and myeloid progenitor cell growth; mast cells IL-5 Activated helper T cells –T H 2 response only Eosinophil production murine B-cell growth IL-6 Activated T cells monocytes, fibroblasts, Progenitor cell stimulation; platelet production; endothelial cells immunoglobulin production in B cells IL-11 As above As LIF IL-7 T-cell survival G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; IL, interleukin; M-CSF, macrophage colony-stimulating factor; SCF, stem cell factor. Adapted from D. Thomas and A. Lopez, 2001. Encyclopedia of Life Sciences: Haematopoietic growth factors, Nature Publishing Group. Go to www.whfreeman.com/immunology Self-Test Review and quiz of key terms Fitzgerald, K. A., et al. 2001. The Cytokine Facts Book, second edi- tion. Academic Press. Flynn, J. L., and J. Chan. 2001. Immunology of tuberculosis. Annu. Rev. Immunol. 19:93–129. Gadina, M., et al. 2001. Signaling by type I and II cytokine recep- tors: ten years after. Curr. Opin. Immunol. 3:363–373. Jaeckel, E., et al. 2001. Treatment of acute hepatitis C with inter- feron H9251-2b. N. Engl. J .Med. 345:1452–1457. Mossman, T. R., H. Cherwinski, M. W. Bond, M. A. Gledlin, and R. L. Coffman. 1986. Two types of murine helper T cell clone. I. Definition according to profiles of lymphokine activities and secreted proteins. J. Immunology 136:2348. Rengarajan, J., S. J. Szabo, and L. H. Glimcher. 2000. Transcrip- tional regulation of Th H 1/Th H 2 polarization. Immunol. Today 10:479–483. Szabo, S. J. et al. 2000. A novel transcription factor, T-bet, directs T H 1 lineage commitment. Cell 100:655–669. Walter, M. R., et al. 1995. Crystal structure of a complex between interferon-H9253 and its soluble high-affinity receptor. Nature 376: 230. Cytokines CHAPTER 12 297 Self-renewing hematopoietic stem cell Common myeloid progenitor Common lymphoid progenitor IL-3, GM-CSF, IL-6 Erythroid progenitor Erythro- poietin IL-5 IL-7, others ? IL-7, others SCF, others SCF, others SCF, others G-CSF M-CSF IL-5 IL-3, GM-CSF Thrombpoietin, IL-11 Megakaryocyte Eosinophil progenitor Granulocyte-monocyte progenitor Basophil progenitor T lymphocytes Natural killer cell B lymphocytes Eosinophil MonocyteNeutrophilBasophilPlateletsErythrocyte FIGURE 12-16 Hematopoietic cytokines and hematopoiesis. A vari- ety of cytokines are involved in supporting the growth and directing the differentiation of hematopoietic cells. Note that additional factors may be required for some of the developmental pathways shown in the dia- gram. CFU = colony-forming unit, a cell capable of generating a colony of cells from which the fully differentiated cell type emerges. USEFUL WEB SITES http://www.rndsystems.com/ The cytokine minireviews found at R&D Systems Web site provide extensive, detailed, well-referenced, and often strik- ingly illustrated reviews of many cytokines and their receptors. http://www.ncbi.nlm.nih.gov:80/LocusLink/index.html LocusLink provides access to sequence and descriptive infor- mation about genetic loci of cytokines and other proteins. It also references papers discussing the basic biology (function and structure) of the gene or protein of interest. Study Questions CLINICAL FOCUS QUESTION Cytokines are proving to be powerful drugs, but their use is accompanied by side effects that can be harmful to patients. What are some of the side effects produced by Actimmune, Roferon, and interferon beta? (Hint: Manufac- turer’s Web sites often provide detailed information on the side effects of drugs they produce.) 1. Indicate whether each of the following statements is true or false. If you think a statement is false, explain why. a. The high-affinity IL-2 receptor consists of two trans- membrane proteins. b. The anti-TAC monoclonal antibody recognizes the IL-1 receptor on T cells. c. All cytokine-binding receptors contain two or three sub- units. d. Expression of the H9252 subunit of the IL-2 receptor is indica- tive of T-cell activation. e. Some cytokine receptors possess domains with tyrosine kinase activity that function in signal transduction. f. All members of each subfamily of the class I cyto- kine (hematopoietin) receptors share a common signal- transducing subunit. 2. When IL-2 is secreted by one T cell in a peripheral lymphoid organ, do all the T cells in the vicinity proliferate in response to the IL-2 or only some of them? Explain. 3. Briefly describe the similarities and differences among cyto- kines, growth factors, and hormones. 4. Indicate which subunit(s) of the IL-2 receptor are expressed by the following types of cells: a. Resting T cells b. Activated T cells c. Activated T cells + cyclosporin A d. Resting T C cells e. CTLs f. NK cells 5. Superantigens have been implicated in several diseases and have been useful as research tools. a. What properties of superantigens distinguish them from conventional antigens? b. By what mechanism are bacterial superantigens thought to cause symptoms associated with food poisoning and toxic-shock syndrome? c. Does the activity of superantigens exhibit MHC restric- tion? 6. IL-3, IL-5, and GM-CSF exhibit considerable redundancy in their effects. What structural feature of the receptors for these cytokines might explain this redundancy? 7. Considerable evidence indicates the existence of two T H -cell subsets, differing in the pattern of cytokines they secrete. a. What type of immune response is mediated by the T H 1 subset? What type of antigen challenge is likely to induce a T H 1-mediated response? b. What type of immune response is mediated by the T H 2 subset? What type of antigen challenge is likely to induce aT H 1-mediated response? 298 PART III Immune Effector Mechanisms