TGF-β and cancer

The relationships between transforming growth factor-β (TGF-β) and cancer are varied and complex. The paradigm that is emerging from the experimental evidence accumulated over the past decade or so is that TGF-β can play two different and opposite roles with respect to the process of malignant progression. During early stages of carcinogenesis, TGF-β acts predominantly as a potent tumor suppressor and may mediate the actions of chemopreventive agents such as retinoids and nonsteroidal anti-estrogens. However, at some point during the development and progression of malignant neoplasms, bioactive TGF-βs make their appearance in the tumor microenvironment and the tumor cells escape from TGF-β-dependent growth arrest. In many cases, this resistance to TGF-β is the consequence of loss or mutational inactivation of the genes that encode signaling intermediates. These include the types I and II TGF-β receptors, as well as receptor-associated and common-mediator Smads. The stage of tumor development or progression at which TGF-β-resistant clones come to dominate the tumor cell population in different types of neoplasm remains to be defined. The phenotypic switch from TGF-β-sensitivity to TGF-β-resistance that occurs during carcinogenesis has several important implications for cancer prevention and treatment.     

TGF-β: 20 years and counting

The history of transforming growth factor-beta (TGF-β) as a bifunctional agent in the immune system is briefly described. The importance of cellular context in understanding the role of TGF-β in regulating immune response is emphasized.     

TGF-β: from latent to active

The transforming growth factor-betas (TGF-βs) are synthesized as precursor proteins that are modified intracellularly prior to secretion. One of the most relevant intracellular modifications is the cleavage of the C-terminal pro-region from the N-terminal portion of the protein. The C-terminal pro-region is referred to as the latency-associated peptide (LAP) while the N-terminal region is called the mature TGF-β or active TGF-β. However, with some exceptions the LAP noncovalently associates with the mature TGF-β prior to secretion. When the mature TGF-β is associated with the LAP it is called L-TGF-β and cannot interact with its receptor and has no biological effect. The TGF-βs and their receptors are very ubiquitously expressed, suggesting that the regulation of TGF-β activity is likely to be complex and multifactorial. However, one of the most important means of controlling the biological effects of TGF-β is the regulation of converting L-TGF-β to active TGF-β. The current literature supports two major mechanisms of activation of L-TGF-β and suggests that the mechanism of activation of L-TGF-β may be varied and context-dependent. For TGF-β to become biologically active the LAP has to be either released from its associations with L-TGF-β or undergo conformational change such that the LAP is not released from the L-TGF-β complex but exposes the TGF-β receptor binding site. Since TGF-β has been associated with the pathogenesis of numerous diseases, the various mechanisms of activation of L-TGF-β in context offer the possibility of controlling TGF-β activity localized to the organ of involvement and to a more specific disease process.     

TGF-β in infections and infectious diseases

Since it was first described as having the ability to inhibit macrophage activation, transforming growth factor-beta (TGF-β) has been analyzed for its role in regulating immune responses to a variety of pathogens, including viruses, bacteria, yeast, and protozoa. Most of the studies have involved organisms that infect macrophages, and this discussion will attempt to highlight these findings. Perhaps the most work has been performed with protozoan pathogens, including Trypanosoma cruzi and a variety of Leishmania species, so the discussion will begin with these organisms. Other studies have focused on mycobacteria and viruses, including human immunodeficiency virus, so these areas will also be emphasized in the discussion. For the most part, investigators have reported that TGF-β has, as expected, a negative influence on host responses and a beneficial effect on the survival and growth of intracellular pathogens. However, other studies have found that TGF-β may have a positive or beneficial effect in some models of infection. This review will attempt to highlight studies and conclusions on the roles of TGF-β in infection.     

Modulation of gene expression in the preimplantation mouse embryo by TGF-α and TGF-β

The effect of growth factors on regulating gene expression in the preimplantation mouse embryo was examined, since results of previous experiments revealed a stimulatory effect of exogenously-added growth factors on preimplantation development in vitro. Treatment of early cavitating blastocysts with either 250 pM TGF-α or TGF-β results in changes in the pattern of total protein synthesis as assessed by high-resolution two-dimensional gel electrophoresis. In some cases, the synthesis of a particular polypeptide is either up- or downregulated by each growth factor, whereas in other instances the synthesis of a polypeptide is modulated by one but not the other growth factor. Use of the mRNA differential display method permitted the identification of genes whose expression is either up- or downregulated by these growth factors. Treatment of mouse blastocysts with either TGF-α or TGF-β results in the increased expression of the b subunit of the F₀ ATPase. TGF-β also stimulates the expression of the DNA polymerase α. TGF-α treatment results in the increase in expression of a gene homologous to the human HEPG2 cDNA, as well as in a decrease in expression of fibronectin. © 1995 Wiley-Liss, Inc.     

Manipulation of TGF-β to control autoimmune and chronic inflammatory diseases

Defining the mechanisms whereby transforming growth factor-beta (TGF-β) controls physiologic inflammation and the immune response and how it contributes to pathology when it is dysregulated is critical to our ability to manipulate the levels and activity of this potent cytokine for therapeutic benefit. In keeping with its dichotomous nature, recent evidence suggests that overproduction and/or activation contribute to persistent inflammation and that antagonists of TGF-β delivered locally can break the cycle of leukocyte recruitment and fibrotic sequelae. On the other hand, systemic routing of TGF-β can also inhibit inflammatory pathogenesis by multiple mechanisms as exemplified by systemic injections of the protein and by recent gene transfer studies. In addition, enhanced levels of circulating endogenous TGF-β appear to be an instrument of suppression during the development of oral tolerance, cyclosporin treatment, and following administration of retinoic acid. Although treatment of autoimmune and chronic inflammatory diseases is an important goal, the multiplicity of actions of TGF-β and the nearly ubiquitous expression of TGF-β and its receptors dictate a cautious approach to the use of this powerful cytokine as a therapeutic agent.     

TGF-β1 regulation of dendritic cells

Dendritic cells (DCs) represent antigen-presenting cell (APC) populations in lymphoid and nonlymphoid organs which are considered to play key roles in the initiation of antigen-specific T-cell proliferation. According to current knowledge, the net outcome of T-cell immune responses seems to be significantly influenced by the activation stage of antigen-presenting DCs. Several studies have shown that transforming growth factor-beta 1 (TGF-β1) inhibits in vitro activation and maturation of DCs. TGF-β1 inhibits upregulation of critical T-cell costimulatory molecules on the surface of DCs and reduces the antigen-presenting capacity of DCs. Thus, in addition to direct inhibitory effects of TGF-β1 on effector T lymphocytes, inhibitory effects of TGF-β1 at the level of APCs may critically contribute to previously characterized immunosuppressive effects of TGF-β1. In contrast to these negative regulatory effects of TGF-β1 on function and maturation of lymphoid tissue type DCs, certain subpopulations of immature DCs in nonlymphoid tissues are positively regulated by TGF-β1 signaling. In particular, epithelial-associated DC populations seem to critically require TGF-β1 stimulation for development and function. Recent studies established that TGF-β1 stimulation is absolutely required for the development of epithelial Langerhans cells (LCs) in vitro and in vivo. Furthermore, TGF-β1 seems to enhance antigen processing and costimulatory functions of epithelial LCs.     

TGF-β1 is an organizer of responses to neurodgeneration

TGF-β1 mRNA and protein were recently found to increase in animal brains after experimental lesions that cause local deafferentation or neuron death. Elevations of TGF-β1 mRNA after lesions are prominent in microglia but are also observed in neurons and astrocytes. Moreover, TGF-β1 mRNA autoinduces its own mRNA in the brain. These responses provide models for studying the increases of TGF-β1 protein observed in βA/amyloid-containing extracellular plaques of Alzheimer's disease (AD) and Down's syndrome (DS) and in brain cells of AIDS victims. Involvement of TGF-β1 in these human brain disorders is discussed in relation to the potent effects of TGF-β1 on wound healing and inflammatory responses in peripheral tissues. We hypothesize that TGF-β1 and possibly other TGF-β peptides have organizing roles in responses to neurodegeneration and brain injury that are similar to those observed in non-neural tissues. Work from many laboratories has shown that activities of TGF-β peptides on brain cells include chemotaxis, modification of extracellular matrix, and regulation of cytoskeletal gene expression and of neurotrophins. Similar activities of the TGF-β's are well established in other tissues.     

TGF-β and fibrosis

Transforming growth factor-beta (TGF-β) isoforms are multifunctional cytokines that play a central role in wound healing and in tissue repair. TGF-β is found in all tissues, but is particularly abundant in bone, lung, kidney and placental tissue. TGF-β is produced by many but not all parenchymal cell types, and is also produced or released by infiltrating cells such as lymphocytes, monocytes/macrophages, and platelets. Following wounding or inflammation, all these cells are potential sources of TGF-β. In general, the release and activation of TGF-β stimulates the production of various extracellular matrix proteins and inhibits the degradation of these matrix proteins, although exceptions to these principles abound. These actions of TGF-β contribute to tissue repair, which under ideal circumstances leads to the restoration of normal tissue architecture and may involve a component of tissue fibrosis. In many diseases, excessive TGF-β contributes to a pathologic excess of tissue fibrosis that compromises normal organ function, a topic that has been the subject of numerous reviews [1, 2 and 3]. In the following chapter, we will discuss the role of TGF-β in tissue fibrosis, with particular emphasis on renal fibrosis.     

Bidirectional regulation of macrophage function by TGF-β

The dual role of transforming growth factor-beta (TGF-β) in modulating macrophage function is an important concept gaining increasing recognition. In addition to its role as a ‘macrophage-deactivating' agent, TGF-β functions as a monocyte activator, inducing cytoke production and mediating host defence. These functions are context-dependent, modulated by the differentiation state of the cell, the local cytokine environment, and the local levels of TGF-β in itself. In general, during the initial stages of inflammation, TGF-β locally acts as a proinflammatory agent by recruiting and activating resting monocytes. As these cells differentiate specific immunosuppressive actions of TGF-β predominate, leading to resolution of the inflammatory response. Increasing our understanding of the bidirectional regulation of macrophage function will facilitate prediction of the ultimate outcome of modulating TGF-β levels in vivo.     

The role of TGF-β in growth, differentiation, and maturation of B lymphocytes

Transforming growth factor-β (TGF-β) affects B cells at all stages in development. It appears to be involved in lymphopoiesis and is required for the development of plasma cells secreting all secondary isotypes. Its ability to inhibit proliferation and stimulate apoptosis suggest that it may be involved both in germinal center development and regulation of B-cell proliferation at sites of high antigen load such as the gastrointestinal tract. Although TGF-β appears to be required for the generation of B cells secreting secondary isotypes, it inhibits secretion of IgM and IgA from cells expressing those isotypes. In this regard, TGF-β may alter the level of RNA processing factors either directly or indirectly by inhibiting progression through the cell cycle. One of the best characterized effects of TGF-β is its ability to stimulate isotype switching to IgA in both mouse and man. There is some controversy concerning its mechanism of action in this process, but its critical role is without question. The controversy may stem in part from an inability to separate the effects of endogenous and exogenous TGF-β in the multiple models of isotype switching. The influence of endogenous TGF-β is perhaps best exemplified by analysis of production of the different classes of IgG in mouse strains producing different levels of TGF-β.     

Role of NK cells and TGF-β in the regulation of T-cell-dependent antibody production in health and autoimmune disease

Natural killer (NK) cells are a third lymphocyte population especially important in innate immunity. NK cells may also have an important role in the regulation of acquired immunity. These lymphocytes spontaneously produce large amounts of both active and latent transforming growth factor-beta (TGF-β). NK-cell-derived TGF-β1 enabled activated CD8⁺ T cells to inhibit antibody production by blocking the induction of this response. Production of lymphocyte-derived TGF-β is decreased in systemic lupus erythematosus. Insufficient levels of this cytokine in SLE and other autoimmune diseases may contribute to defective T regulatory cell function characteristic of this and other autoimmune diseases. NK cells are found in mucosal tissues and the TGF-β spontaneously released by these cells could contribute to the usual tolerogenic response of T cells to antigens presented at these sites. Thus, in addition to its well known immunosuppressive effects, TGF-β could have an equally important role in the generation of regulatory T cells.     

Regulation of GM-CSF-Induced Dendritic Cell Development by TGF-β1 and Co-Developing Macrophages

Using a culture system of bone marrow progenitor cells with GM-CSF and TGF-β1, a study was performed to analyze the effect of TGF-β1 on the development of dendritic cells (DC) and to elucidate the regulatory role of macrophages co-developing with dendritic cells. The results demonstrate that DC generated in the presence of TGF-β1 were immature with respect to the expression of CD86, nonspecific esterase activity and cell shape. Such inhibitory effects of TGF-β1 were dependent on FcR⁺ macrophages, which were depleted by panning. TGF-β1 did not appear to inhibit the commitment of progenitor cells to the DC lineage. In addition, TGF-β1 also acted directly on the intermediate stage of DC to prevent their over-maturation, which results in a preferential decrease in MHC class II, but not in CD86, in the presence of TNF-α. FcR⁺ suppressive macrophages were also shown to facilitate DC maturation when stimulated via FcR-mediated signals even in the presence of TGF-β1. These results indicate that TGF-β1 indirectly and directly regulate the development of DC and that co-developing macrophages have a regulatory role in DC maturation.     

Terminal Differentiation of Normal Human Oral Keratinocytes Is Associated with Enhanced Cellular TGF-β and Phospholipase C-γ1 Levels and Apoptotic Cell Death

Subculture of primary normal human oral keratinocytes (NHOK) results in terminal differentiation, leading to cell death. To investigate whether the subculture-induced death of NHOK is due to apoptosis, we studied transferase-mediated dUTP nick end labeling (TUNEL)-positive cells, DNA fragmentation, and expression of several apoptosis-associated genes from NHOK with different passage numbers. We also determined the effect of transforming growth factor β₁ (TGF-β₁) on the induction of apoptosis in NHOK. We were able to subculture primary NHOK up to the fifth passage, at which point cells showed morphological features of differentiation. Appearance of DNA fragmentation concurrently occurred with an increase in the number of TUNEL-positive cells with higher passage numbers. The level of cellular p53 proteins was gradually decreased by the continued passage of cells, whereas the levels of intracellular and secreted TGF-β and phospholipase C-γ1 (PLC-γ1) were significantly elevated by serial subculture. Exogenous TGF-β₁ also induced differentiation and apoptosis of proliferating NHOK. These data indicate that terminal differentiation of NHOK is associated with apoptosis, which is, in part, linked to elevated cellular levels of TGF-β and PLC-γ1.     

TGF-β3 and cancer: A review

With the development of growth factors and growth factor modulators as therapeutics for a range of disorders, it is prudent to consider whether modulating the growth factor profile in a tissue can influence tumour initiation or progression. As recombinant human TGF-β3 (avotermin) is being developed for the improvement of scarring in the skin it is important to understand the role, if any, of this cytokine in tumour progression.Elevated levels of TGF-β3 expression detected in late-stage tumours have linked this cytokine with tumourigenesis, although functional data to support a causative role are lacking. While it has proved tempting for researchers to interpret a ‘correlation’ as a ‘cause’ of disease, what has often been overlooked is the normal biological role of TGF-β3 in processes that are often subverted in tumourigenesis. Clarifying the role of this cytokine is complicated by inappropriate extrapolation of the data relating to TGF-β1 in tumourigenesis, despite marked differences in biology between the TGF-β isoforms. Indeed, published studies have indicated that TGF-β3 may actually play a protective role against tumourigenesis in a range of tissues including the skin, breast, oral and gastric mucosa. Based on currently available data it is reasonable to hypothesize that administration of acute low doses of exogenous TGF-β3 is unlikely to influence tumour initiation or progression.