The Role of Growth Hormone in Modulation of the Immune Responsea KEITH W. KELLEY Laboratory of Immunophysiology Department of Animal Sciences University of Illinois Urbana, IIlinois 61801
INTRODUCTION T lymphocytes synthesize and secrete a wide variety of molecules called lymphokines, and these proteins affect the differentiation, growth, and function of leukocytes. It was originally believed that each of these cytokines served a single function, but this idea is no longer tenable.' For example, interleukin-2 (IL-2) is a growth factor for both T and B lymphocytes. IL-4 and IL-2 both function as T cell growth factors. IL-6 augments the secretion of antibody from plasma cells. but it is also the major stimulus for induction of acute phase proteins from the liver. IL-6 was originally identified for its antiviral properties. The finding that lymphokines have more than one function (pleiotropy) and that many lymphokines can cause the same effect (redundancy) on leukocytes is now widely recognized by immunologists? The ideas of pleiotropy and redundancy of lymphokines have recently been extended to the neuroendocrine system. IL-6 increases the synthesis of nerve growth factor by astr~cytes.~ Specific receptors for a macrophage-derived protein, IL-I, have been identified in the brain," and there are IL-l-containing neurons in the brain? It is now abundantly clear that another action of IL-1 is to increase the concentrationof adrenal-derived glucocorticoids in the blood, an effect mediated by the central nervous system, by direct action of IL-1 on the pituitary gland, or by both (reviewed in Reference 6). Indeed, since glucocorticoids suppress the synthesis of IL-1, it is likely that IL-1 is part ofa regulatory feedback loop with the pituitarygland that controls its own production. It should come as no surprise that hormones now join the list of molecules that not only affect their classical target organs, but they are also molecules that alter functional activities of leukocytes as well. A number of hormones are even
"This work was supported by grants from the NIH (AG06246-Ol),Office of Naval Research (N00014-89-5-1956),United States Department of Agriculture (89-37265-4536).and Moorman Mfg. Co. 95
96
ANNALS NEW YORK ACADEMY OF SCIENCES
synthesized by leukocytes (reviewed in Reference 7). Growth hormone is a protein consisting of 191 amino acids and is synthesized by acidophils in the pars distalis. Growth hormone induces a variety of metabolic changes, and these effects have been recently summarized? The recombinant form of human growth hormone was first approved for humans with a growth hormone deficiency in 1985. Growth hormone is now used extensively for these subjects, and its use continues to increase annually. It is therefore important to understand all of the biologic properties of this important hormone. It is the purpose of this short chapter to describe the modulatory role of growth hormone in the immune response. The most recent findings will be emphasized.
GENERAL IMMUNOMODULATORY PROPERTIES OF GROWTH HORMONE There has been an interest in the role of pituitary hormones in regulating the immune response since Smithg demonstrated in 1930 that the thymus gland is atrophied in hypophysectomized rats. Research conducted by Baroni and Fabris in Italy showed that antibody synthesis is defective in the Snell-Bagg pituitary dwarf mouse, and that growth hormone can reverse this defect.I0 In 1968, Pierpaoli and Sorkin” also showed that an antiserum to growth hormone induces thymic atrophy in mice, and this effect is reversed by growth hormone. Istvan Berczi and his colleagues subsequently demonstrated that both antibody synthesis and contact sensitivityreactions are augmented when growth hormone is injected into hypophysectomized rats (discussed in Reference 12). Jim Roth and colleague^'^ also showed that growth hormone injections augment growth of the thymus gland in dwarf dogs, and we subsequently demonstrated that growth hormone- and prolactin-secreting pituitary cells can reverse the age-associated involution of the thymus gland.14 Most recently, Douglas Weigent and collaborators’5 at the University of Alabama at Birmingham have demonstrated that growth hormone is actually synthesized by leukocytes. A broad overview of the effects of growth hormone on a number of immune responses has recently been published,16and a summary of these effects is presented in TABLE 1. Selected aspects of the immunomodulatory effects of growth hormone are discussed below.
THE THYMUS GLAND The thymus gland is now known to be responsible for gene rearrangement of specific antigen receptors on T lymphocytes and deletion of autoreactive T cell clones. Pituitary-deficient mice, rats, and dogs have small thymus glands, and the size and structural organization of the thymus is reconstituted in these animals by injections of growth hormone. Thymic involution that normally occurs during aging is reversed by injections of growth hormone” or by implanting growth hormone- and prolactin-secreting GH, pituitary cells.14 Similarly, injections of growth hormone into aged rats augment lymphocyte proliferation that is triggered by T cell lectins.18
KELLEY THE ROLE OF GROWTH HORMONE IN MODULATION
97
T h e increase in size of the thymus gland is mediated by augmenting the number of cortical thymocytes. However, growth hormone also affects thymic epithelial cells because it increases synthesis of a thymic hormone known as t h y m ~ 1 i n . l ~
TABLE 1.
Growth Hormone Modulates Cells Involved in the Immune Response16
Growth hormone deficiencies and immunoregulation Thymic atrophy and wasting in mice and dogs Reduced antibody synthesis in mice Delayed skin graft rejection in mice Normal lymphoid cell subsets and thymic histology with reduction in peripheral T and B cells Pituitary hypoplasia and thymic atrophy in humans X-linked growth hormone deficiency and complete inability to synthesize antibodies Reduction in activity of natural killer cells in humans Defective allogeneic mixed lymphocyte reaction Reduction in plasma thymulin in humans and mice Normal immunoglobulin concentrations and lymphoid cell subsets in humans Decreased insulin-inducedgrowth hormone response in patients with telangiectasis and bowel disease Growth hormone and the thymus gland Increases thymic size and DNA synthesis in young rodents Improves thymic size and morphology in aged animals Increases plasma thymulin in humans and dogs Growth hormone and lymphoid cells Lymphocytes have receptors for growth hormone Augments antibody synthesis and reduces skin graft survival in vivo Increases lectin-induced T cell proliferation and IL-2 synthesis in vivo Stimulates proliferation of human lymphoblastoid cells Augments basal lymphocyte proliferation in vitro Increases activity of cytotoxic T lymphocytes in vitro Augments activity of natural killer cells in vivo Synthesized by lymphoid cells Growth hormone and phagocytic cells Primes macrophages for superoxide anion release in vitro and in vivo Augments respiratory burst in neutrophils from growth hormone-deficient patients in vivo Increases basal respiratory burst of human neutrophils and inhibits activated burst in vitro Growth hormone and hemopoiesis Augments neutrophil differentiation in vitro Augments erythropoiesis
CYTOLYTIC CELLS Lymphocytes express specific, high-affinity receptors for growth hormone?' but the distribution of these receptors on specific subsets of lymphocytes is unknown. Growth hormone augments t h e activity of cytotoxic T lymphocytes in vitro.2' Natural killer cell activity is deficient in hypophysectomized rats and aged humans and rats, and growth hormone consistently augments natural killer cell activity in these subject^.'^^^^^^^ Leukocytes from growth hormone-deficient children have low levels of natural killer cell activity, and this appears to be due to a developmental defect in the generation of natural killer cells."
ANNALS NEW YORK ACADEMY OF SCIENCES
98
GRANULOCYTE DIFFERENTIATION Multipotential progenitor stem cells are found in the bone marrow that ultimately form all cells that circulate in blood. Defined differentiation signals, such as granulocytelmacrophage colony-stimulating factor (GM-CSF), induces myeloid precursor cells to differentiate into granulocytes and monocyteshacrophages. Merchav et aLZSrecently demonstrated that the addition of recombinant human growth hormone to human bone marrow cells incubated with GM-CSF doubled the number of precursor cells that differentiated into granulocytes (TABLE2). The growth hormone-dependent protein, IGF-I, also increased the number of granulocyte colonies when added to this culture system. As expected, the IGF-I-induced enhancement was abrogated by addition of an antibody that is specific for the IGFI receptor. The provocative finding was that this antibody also blocked the enhancing effect of growth hormone on development of granulocyte colonies. These data establish two important points. ( I ) Both growth hormone and IGFI augment the development of granulocytic cells from primitive bone marrow-derived precursors in the presence of GM-CSF. However, neither growth hormone nor IGF-I can replace the requirement for GM-CSF in granulocyte differentiation. (2) Adherent human bone marrow cells may secrete IGF-I after stimulation with growth hormone. This bone marrow-derived IGF-I is the molecule that is responsible for augmentation of granulocyte differentiation. This conclusion is compatible with two recent reports that have shown that adherent mononuclear cells can express transcripts for IGF-126and synthesize and secrete IGF-I."
PRIMING OF MACROPHAGES Priming for Toxic Oxygen Metabolites
Phagocytosis and the subsequent destruction of invading microorganisms by phagocytic cells are considered to be the first line of defense of the host against acute bacterial and fungal infections. Myeloid cells can be triggered to produce reactive oxygen intermediates that nonspecifically kill ingested organisms. Triggering of phagocytic cells can be accomplished with a variety of substances, such as opsonized bacteria, the bacterial product formyl-methionyl-leucyl-phenylalanine, or pharmacological activators of protein kinase C (e.g., phorbol myristate acetate). These TABLE 2. Growth Hormone and IGF-I Augment the Formation of Granulocyte
Colonies from Human Bone Marrow Progenitor Cells Incubated with Recombinant GM-CSF, and the Enhancement Caused by Growth Hormone Is Blocked by an Antibody to the IGF-I Receptor z5a Treatment
Granulocyte Colonies No Antibody Plus Antibody
Medium Human growth hormone
IGF-I
36 * 6 86 * 10 82 * 12
44 * I 37 * 6b 45 * 6* ~
"Mean * SEM. *Significantly different from control.
~-
KELLEY THE ROLE OF GROWTH HORMONE IN MODULATION
99
Native Rat Growth Hormone and Recombinant Porcine Growth Hormone Enhance the Production of Superoxide Anion from Peritoneal Macrophages of Hypophysectomized Rats2*
TABLE 3.
Item Hypophysectomized rats + Vehicle + Rat IFN-y + Native rat GH
+ Recombinant porcine GH
Dose/ Ratmay
200 pl 500 U 24 12" PCg 48pg 96 Pg pg
l2 " 24 PCg
Growth (Gmay) 0.392" 0.282" 0.252' 1.8504 * 2.32V 2.8701 3.440" 0.815b 1.067b.c 1.297'
nMol0,- / mg protein/h 4@ -17 417b
3" 267b 247b
309
-62 280b 344b
Means with different superscriptsare different (p
INTRODUCTION T lymphocytes synthesize and secrete a wide variety of molecules called lymphokines, and these proteins affect the differentiation, growth, and function of leukocytes. It was originally believed that each of these cytokines served a single function, but this idea is no longer tenable.' For example, interleukin-2 (IL-2) is a growth factor for both T and B lymphocytes. IL-4 and IL-2 both function as T cell growth factors. IL-6 augments the secretion of antibody from plasma cells. but it is also the major stimulus for induction of acute phase proteins from the liver. IL-6 was originally identified for its antiviral properties. The finding that lymphokines have more than one function (pleiotropy) and that many lymphokines can cause the same effect (redundancy) on leukocytes is now widely recognized by immunologists? The ideas of pleiotropy and redundancy of lymphokines have recently been extended to the neuroendocrine system. IL-6 increases the synthesis of nerve growth factor by astr~cytes.~ Specific receptors for a macrophage-derived protein, IL-I, have been identified in the brain," and there are IL-l-containing neurons in the brain? It is now abundantly clear that another action of IL-1 is to increase the concentrationof adrenal-derived glucocorticoids in the blood, an effect mediated by the central nervous system, by direct action of IL-1 on the pituitary gland, or by both (reviewed in Reference 6). Indeed, since glucocorticoids suppress the synthesis of IL-1, it is likely that IL-1 is part ofa regulatory feedback loop with the pituitarygland that controls its own production. It should come as no surprise that hormones now join the list of molecules that not only affect their classical target organs, but they are also molecules that alter functional activities of leukocytes as well. A number of hormones are even
"This work was supported by grants from the NIH (AG06246-Ol),Office of Naval Research (N00014-89-5-1956),United States Department of Agriculture (89-37265-4536).and Moorman Mfg. Co. 95
96
ANNALS NEW YORK ACADEMY OF SCIENCES
synthesized by leukocytes (reviewed in Reference 7). Growth hormone is a protein consisting of 191 amino acids and is synthesized by acidophils in the pars distalis. Growth hormone induces a variety of metabolic changes, and these effects have been recently summarized? The recombinant form of human growth hormone was first approved for humans with a growth hormone deficiency in 1985. Growth hormone is now used extensively for these subjects, and its use continues to increase annually. It is therefore important to understand all of the biologic properties of this important hormone. It is the purpose of this short chapter to describe the modulatory role of growth hormone in the immune response. The most recent findings will be emphasized.
GENERAL IMMUNOMODULATORY PROPERTIES OF GROWTH HORMONE There has been an interest in the role of pituitary hormones in regulating the immune response since Smithg demonstrated in 1930 that the thymus gland is atrophied in hypophysectomized rats. Research conducted by Baroni and Fabris in Italy showed that antibody synthesis is defective in the Snell-Bagg pituitary dwarf mouse, and that growth hormone can reverse this defect.I0 In 1968, Pierpaoli and Sorkin” also showed that an antiserum to growth hormone induces thymic atrophy in mice, and this effect is reversed by growth hormone. Istvan Berczi and his colleagues subsequently demonstrated that both antibody synthesis and contact sensitivityreactions are augmented when growth hormone is injected into hypophysectomized rats (discussed in Reference 12). Jim Roth and colleague^'^ also showed that growth hormone injections augment growth of the thymus gland in dwarf dogs, and we subsequently demonstrated that growth hormone- and prolactin-secreting pituitary cells can reverse the age-associated involution of the thymus gland.14 Most recently, Douglas Weigent and collaborators’5 at the University of Alabama at Birmingham have demonstrated that growth hormone is actually synthesized by leukocytes. A broad overview of the effects of growth hormone on a number of immune responses has recently been published,16and a summary of these effects is presented in TABLE 1. Selected aspects of the immunomodulatory effects of growth hormone are discussed below.
THE THYMUS GLAND The thymus gland is now known to be responsible for gene rearrangement of specific antigen receptors on T lymphocytes and deletion of autoreactive T cell clones. Pituitary-deficient mice, rats, and dogs have small thymus glands, and the size and structural organization of the thymus is reconstituted in these animals by injections of growth hormone. Thymic involution that normally occurs during aging is reversed by injections of growth hormone” or by implanting growth hormone- and prolactin-secreting GH, pituitary cells.14 Similarly, injections of growth hormone into aged rats augment lymphocyte proliferation that is triggered by T cell lectins.18
KELLEY THE ROLE OF GROWTH HORMONE IN MODULATION
97
T h e increase in size of the thymus gland is mediated by augmenting the number of cortical thymocytes. However, growth hormone also affects thymic epithelial cells because it increases synthesis of a thymic hormone known as t h y m ~ 1 i n . l ~
TABLE 1.
Growth Hormone Modulates Cells Involved in the Immune Response16
Growth hormone deficiencies and immunoregulation Thymic atrophy and wasting in mice and dogs Reduced antibody synthesis in mice Delayed skin graft rejection in mice Normal lymphoid cell subsets and thymic histology with reduction in peripheral T and B cells Pituitary hypoplasia and thymic atrophy in humans X-linked growth hormone deficiency and complete inability to synthesize antibodies Reduction in activity of natural killer cells in humans Defective allogeneic mixed lymphocyte reaction Reduction in plasma thymulin in humans and mice Normal immunoglobulin concentrations and lymphoid cell subsets in humans Decreased insulin-inducedgrowth hormone response in patients with telangiectasis and bowel disease Growth hormone and the thymus gland Increases thymic size and DNA synthesis in young rodents Improves thymic size and morphology in aged animals Increases plasma thymulin in humans and dogs Growth hormone and lymphoid cells Lymphocytes have receptors for growth hormone Augments antibody synthesis and reduces skin graft survival in vivo Increases lectin-induced T cell proliferation and IL-2 synthesis in vivo Stimulates proliferation of human lymphoblastoid cells Augments basal lymphocyte proliferation in vitro Increases activity of cytotoxic T lymphocytes in vitro Augments activity of natural killer cells in vivo Synthesized by lymphoid cells Growth hormone and phagocytic cells Primes macrophages for superoxide anion release in vitro and in vivo Augments respiratory burst in neutrophils from growth hormone-deficient patients in vivo Increases basal respiratory burst of human neutrophils and inhibits activated burst in vitro Growth hormone and hemopoiesis Augments neutrophil differentiation in vitro Augments erythropoiesis
CYTOLYTIC CELLS Lymphocytes express specific, high-affinity receptors for growth hormone?' but the distribution of these receptors on specific subsets of lymphocytes is unknown. Growth hormone augments t h e activity of cytotoxic T lymphocytes in vitro.2' Natural killer cell activity is deficient in hypophysectomized rats and aged humans and rats, and growth hormone consistently augments natural killer cell activity in these subject^.'^^^^^^^ Leukocytes from growth hormone-deficient children have low levels of natural killer cell activity, and this appears to be due to a developmental defect in the generation of natural killer cells."
ANNALS NEW YORK ACADEMY OF SCIENCES
98
GRANULOCYTE DIFFERENTIATION Multipotential progenitor stem cells are found in the bone marrow that ultimately form all cells that circulate in blood. Defined differentiation signals, such as granulocytelmacrophage colony-stimulating factor (GM-CSF), induces myeloid precursor cells to differentiate into granulocytes and monocyteshacrophages. Merchav et aLZSrecently demonstrated that the addition of recombinant human growth hormone to human bone marrow cells incubated with GM-CSF doubled the number of precursor cells that differentiated into granulocytes (TABLE2). The growth hormone-dependent protein, IGF-I, also increased the number of granulocyte colonies when added to this culture system. As expected, the IGF-I-induced enhancement was abrogated by addition of an antibody that is specific for the IGFI receptor. The provocative finding was that this antibody also blocked the enhancing effect of growth hormone on development of granulocyte colonies. These data establish two important points. ( I ) Both growth hormone and IGFI augment the development of granulocytic cells from primitive bone marrow-derived precursors in the presence of GM-CSF. However, neither growth hormone nor IGF-I can replace the requirement for GM-CSF in granulocyte differentiation. (2) Adherent human bone marrow cells may secrete IGF-I after stimulation with growth hormone. This bone marrow-derived IGF-I is the molecule that is responsible for augmentation of granulocyte differentiation. This conclusion is compatible with two recent reports that have shown that adherent mononuclear cells can express transcripts for IGF-126and synthesize and secrete IGF-I."
PRIMING OF MACROPHAGES Priming for Toxic Oxygen Metabolites
Phagocytosis and the subsequent destruction of invading microorganisms by phagocytic cells are considered to be the first line of defense of the host against acute bacterial and fungal infections. Myeloid cells can be triggered to produce reactive oxygen intermediates that nonspecifically kill ingested organisms. Triggering of phagocytic cells can be accomplished with a variety of substances, such as opsonized bacteria, the bacterial product formyl-methionyl-leucyl-phenylalanine, or pharmacological activators of protein kinase C (e.g., phorbol myristate acetate). These TABLE 2. Growth Hormone and IGF-I Augment the Formation of Granulocyte
Colonies from Human Bone Marrow Progenitor Cells Incubated with Recombinant GM-CSF, and the Enhancement Caused by Growth Hormone Is Blocked by an Antibody to the IGF-I Receptor z5a Treatment
Granulocyte Colonies No Antibody Plus Antibody
Medium Human growth hormone
IGF-I
36 * 6 86 * 10 82 * 12
44 * I 37 * 6b 45 * 6* ~
"Mean * SEM. *Significantly different from control.
~-
KELLEY THE ROLE OF GROWTH HORMONE IN MODULATION
99
Native Rat Growth Hormone and Recombinant Porcine Growth Hormone Enhance the Production of Superoxide Anion from Peritoneal Macrophages of Hypophysectomized Rats2*
TABLE 3.
Item Hypophysectomized rats + Vehicle + Rat IFN-y + Native rat GH
+ Recombinant porcine GH
Dose/ Ratmay
200 pl 500 U 24 12" PCg 48pg 96 Pg pg
l2 " 24 PCg
Growth (Gmay) 0.392" 0.282" 0.252' 1.8504 * 2.32V 2.8701 3.440" 0.815b 1.067b.c 1.297'
nMol0,- / mg protein/h 4@ -17 417b
3" 267b 247b
309
-62 280b 344b
Means with different superscriptsare different (p
