《免疫学》（英文版） Chapter 12 Cytokines

okInes chapter 12 HE DEVELOPMENT OF AN EFFECTIVE IMMUNE response involves lymphoid cells, inflammatory cells, and hematopoietic cells. The complex inter- actions among these cells are mediated by a group of pro- ins collectively designated cytokines to denote their role in cell-to-cell communication. Cytokines are low-molecular- weight regulatory proteins or glycoproteins secreted by ite blood cells and various other cells in the body in response to a number of stimuli. These proteins assist in reg cytokines possess direct effector functions of their owl me ulating the development of immune effector cells, and some This chapter focuses on the biological activity of cyto- Class I Cytokine Receptors ines, the structure of cytokines and their receptors, signal transduction by cytokine receptors, the role of cytokine abnormalities in the pathogenesis of certain diseases, and a Properties of Cytokines therapeutic uses of cytokines or their receptors. The impor Cytokine Receptors tant role of cytokines in the inflammatory response is a Cytokine Antagonists described in Chapter 15 a Cytokine Secretion by TH1 and TH2 Subsets a Cytokine-Related Diseases Properties of Cytokines a Therapeutic Uses of Cytokines and Their Receptors Cytokines bind to specific receptors on the membrane of Cytokines in Hematopoiesis target cells, triggering signal-transduction pathways that ultimately alter gene expression in the target cells(figu 12-la). 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 receptor exhibit very high affinity for each other, with dissociatio constants ranging from 10 to 10M. Because their af- finities are so high, cytokines can mediate biological effects at picomolar concentrations the activity of numerous cells involved in the immune a particular cytokine may bind to receptors on the mem- response. For example, cytokines produced by activated TH brane of the same cell that secreted it, exerting autocrine cells can influence the activity of B cells, Tc cells, natural ction; it may bind to receptors on a target cell in close prox- killer cells, macrophages, granulocytes, and hematopoietic imity to the producer cell, exerting paracrine action; in a few stem cells, thereby activating an entire network of interact- cases, it may bind to target cells in distant parts of the body exerting endocrine action( Figure 12-1b). Cytokines regu Cytokines exhibit the attributes of pleiotropy, redun- late the intensity and duration of the immune response by dancy, synergy, antagonism, and cascade induction, which stimulating or inhibiting the activation, proliferation, and/ permit them to regulate cellular activity in a coordinated or differentiation of various cells and by regulating the secre- interactive way(Figure 12-2). a given cytokine that has tion of antibodies or other cytokines. As described later, different biological effects on different target cells has a binding of a given cytokine to responsive target cells gener- pleiotropic action. Two or more cytokines that mediate sim- ally stimulates increased expression of cytokine receptors ilar functions are said to be redundant; redundancy makes it and secretion of other cytokines, which affect other target difficult to ascribe a particular activity to a single cytokine cells in turn. Thus, the cytokines secreted by even a small Cytokine synergism occurs when the combined effect of two number of lymphocytes activated by antigen can influence cytokines on cellular activity is greater than the additive

■ Properties of Cytokines ■ Cytokine Receptors ■ Cytokine Antagonists ■ Cytokine Secretion by TH1 and TH2 Subsets ■ Cytokine-Related Diseases ■ Therapeutic Uses of Cytokines and Their Receptors ■ 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 cytokinesto 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 TH cells can influence the activity of B cells, TC 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

Cytokines cHAPTER 12 277 (b) Cytokine-producing cell Autocrine action Paracrine action Nearby cell Gene Target cell Endocrine action Distant cell Biological effects JRE 12-1(a)Overview of the induction and function of es. (b)Most cytokines exhibit autocrine and/or paracrine action fewer exhibit endocrine action effects of the individual cytokines. In some cases, cytokines of cytokines, the chemokines, a group of low-molecular- exhibit antagonism; that is, the effects of one cytokine inhibit weight cytokines that affect chemotaxis and other aspects or offset the effects of another cytokine Cascade induction of leukocyte behavior. These molecules play an important occurs when the action of one cytokine on a target cell role in the inflammatory response and are described in induces that cell to produce one or more other cytokines, Chapter 15 which in turn may induce other target cells to produce other Because cytokines share many properties with hormones cytokines and growth factors, the distinction between these three The term cytokine encompasses those cytokines secreted classes of mediators is often blurred. All three are secreted by lymphocytes, substances formerly known as lympho- soluble factors that elicit their biological effects at picomolar kines, and those secreted by monocytes and macrophages, concentrations by binding to receptors on target cells substances formerly known as monokines. Although these growth factors tend to be produced constitutively, whereas other two terms continue to be used, they are misleading tokines and hormones are secreted in response to discrete because secretion of many lymphokines and monokines is stimuli, and secretion is short-lived, generally ranging from a not limited to lymphocytes and monocytes as these terms few hours to a few days. Unlike hormones, which generally imply, but extends to a broad spectrum of cells and types. act long range in an endocrine fashion, most cytokines act For this reason, the more inclusive term cytokine is preferred. over a short distance in an autocrine or paracrine fashion. In lany cytokines are referred to as interleukins, a name addition, most hormones are produced by specialized glands indicating that they are secreted by some leukocytes and act and tend to have a unique action on one or a few types of tar upon other leukocytes. Interleukins 1-25 have been identi- get cell. In contrast, cytokines are often produced by, and fied. There is that still other cytokines will bind to, a variety of cells be discovered and that the interleukin group will expand The activity of cytokines was first recognized in the mid further. Some cytokines are known by common names, 1960s, when supernatants derived from in vitro cultures of including the interferons and tumor necrosis factors. Re- lymphocytes were found to contain factors that could regulate cently gaining prominence is yet another another subgroup proliferation, differentiation, and maturation of allogeneic

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

278 paRT I Immune Effector mechanisms get Cell Effect Activatio Activated Tu cells PLEIOTROPY IL-4 Proliferation IFN-Y Thymocyte Activated TH cells Mast cel REDUNDANCY 9→出彐 Proliferation 12 Activated Tu cells B cell SYNERGY IL-4 Induces class switch to lge IL-5 Activated TH cells ANTAGONISM Activated TH cells IL-4 Blocks class switch to IgE induced by IL-4 IFN-Y IFN-Y TNF, IL-2, and Activated Tu cells B cell other cytokines FIGURE 12.2 Cytokine attributes of (a)pleiotropy, redundancy. synergy(synergism), antagonism, and(b) cascade induction immune-system cells. Soon after, it was discovered that pro- natans and the absence of well-defined assay systems for duction of these factors by cultured lymphocytes was induced individual cytokines. a great advance was made with the by activation with antigen or with nonspecific mitogens. Bio- development of gene-cloning techniques during the 1970s chemical isolation and purification of cytokines was ham- and 1980s, which made it possible to produce pure cytokines red because of their low concentration in culture super- by expressing the protein from cloned genes. The discovery of

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 TH cells PLEIOTROPY Mast cell IL-4 Thymocyte B cell Activation Proliferation Differentiation Proliferation Target Cell Effect IL-4 Proliferation IL-2 IL-5 Activated TH cells REDUNDANCY IL-4 + IL-5 Activated TH cells SYNERGY Induces class switch to IgE Blocks class switch to IgE induced by IL-4 IL-4 IFN-γ Activated TH cells ANTAGONISM Proliferation B cell B cell B cell Activated TH cells Activated TH 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

Cytokines cHAPTER 12 characterized so far belong to one of four groups: the Cytokine bound 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 Add substrate considerably, all of them have a high degree of a-helical and measure structure and little or no B-sheet structure The molecules (a)Interleukin 2 36—42 strand 05 C-D Concentration of IL-12(pg/ml) FIGURE 12-3 ELISA assay of a cytokine. (a)The sample contain- ing the cytokine of interest is captured by specific antibody(blue) NHo COOH 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 (b)Interleukin 4 yme 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 shown here is for human interleukin 12(IL-12). It is clear that assay is sufficiently sensitive to detect as little as 1 picogram of IL-12 /Part (b)courtesy of R&D Systems. cell lines whose growth depended on the presence of a partic lar 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) FIGURE 12.4 Several representations of structures in the hemato. Cytokines Belong to Four Structural poietin family. (a) Left: Topographical representation of the primary Families structure of IL-2 showing a-helical regions(a and A-D)and connect ing chains of the molecule. Right: Proposed three-dimensional model Once the genes encoding various cytokines had been cloned, of IL-2. (b) Ribbon model of IL-4 deduced from x-ray crystallographic sufficient quantities of purified preparations became avail- analysis of the molecule In(a)and ( b)the a helices are shown in red able for detailed studies on their structure and function. and the B sheets in blue. The structures of other cytokines belonging to Cytokines generally have a molecular mass of less than the hematopoietin family are thought to be generally similar. /Part(b) 30 kDa. Structural studies have shown that the cytokines from/L Boulay and W.E. Paul, 1993, Curr Biol. 3: 573/

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 -helical structure and little or no -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 E E E E S (a) (b) Concentration of IL-12 (pg/ml) Optical density 10 1 0.1 0.01 0.1 1 10 100 α S S 105 C N 133 6 114 30 36 42 73 80 96 52 58 D A C B C 105 B–C loop β strand C–D loop A–B loop COOH NH2 A C B' B D α P65 C58 (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 -helical regions ( 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 helices are shown in red and the 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.]

280 PART III Immune Effector Mechanisms VISUALIZING CONCEPTS immunity Cytokine promotion of activation, differentiation, proliferation, or cell death of T cells, B cells, macrophages. dendritic cells, NK cells, and other leukocytes. FIGURE12.5 Interaction of antigen with macrophages and the merous cytokines(blue arrows), generating a complex network of ubsequent activation of resting TH cells leads to release of nu- interacting cells in the immune response share a similar polypeptide fold, with four a-helical regions and differentiation, and the healing of wounds. Although the (A-D)in which the first and second helices and the third and immune response to a specific antigen may include the pro- fourth helices run roughly parallel to one another and are duction of cytokines, it is important to remember that connected by loops. cytokines act in an antigen-nonspecific manner. That is, they affect whatever cells they encounter that bear appropriate Cytokines Have Numerous Biological receptors and are in a physiological state that allows them Functions to respo Cytokines are involved in a staggeringly broad array of Although a variety of cells can secrete cytokines, the two biological activities including innate immunity, adaptive principal producers are the TH cell and the macrophage. immunity, inflammation, and hematopoiesis. Altogether, the Cytokines released from these two cell types activate an total number of proteins with cytokine activity easily exceeds entire network of interacting cells(Figure 12-5). Among the 100 and research continues to uncover new ones. Table 12-1 numerous physiologic responses that require cytokine in- summarizes the activities of some cytokines and places them volvement are development of cellular and humoral im- into functional groups. An expanded list of cytokines can be mune responses, induction of the inflammatory response, found in the Appendix. It should be kept in mind that most regulation of hematopoiesis, control of cellular proliferation of the listed functions have been identified from analysis of

share a similar polypeptide fold, with four -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 TH 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 TH cell Adaptive immunity Cytokine promotion of macrophage activation Cytokine promotion of TH 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 TH cells leads to release of nu￾merous cytokines (blue arrows), generating a complex network of interacting cells in the immune response.