This article appears in the following Rheumatology issue: Update in systemic sclerosis [View the issue table of contents]
Connective tissue growth factor: growth factor, matricellular organizer, fibrotic biomarker or molecular target for anti-fibrotic therapy in SSc?
1Division of Medicine, Research Department of Inflammation, Centre for Rheumatology and Connective Tissue Diseases, Royal Free and University College Medical School, University College London, London, UK.
Correspondence to: D. Abraham, Division of Medicine, Research Department of Inflammation, Centre for Rheumatology and Connective Tissue Diseases, Royal Free and University College Medical School, University College London, Royal Free Campus, Roland Hill Street, London NW3 PF, UK. E-mail: d.abraham{at}medsch.ucl.ac.uk
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Abstract |
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 Top Abstract Biology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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SSc is characterized by enhanced extracellular matrix (ECM) production resulting in excessive scarring and replacement fibrosis affecting the interstitial and vascular compartments of multiple organs. Although the precise molecular mechanisms driving fibrosis remain elusive, TGF-β and connective tissue growth factor (CTGF), are considered key mediators. CTGF is over-expressed in lesional tissue and enhanced levels in the circulation are an indicator of disease extent and severity. Rapidly induced by TGF-β and ET-1, CTGF activates several signal transduction pathways via surface receptors that modulate the functional activities of fibroblasts, endothelial and smooth muscle cells. In vivo, over-expression of CTGF causes ECM accumulation and promotes tissue fibrosis. In animal models of SSc, neutralization of CTGF with antibody blockade or siRNA, suppresses fibrogenesis. This article examines the role of CTGF as an integrator of extracellular signals, fibrotic biomarker and discusses the potential value of CTGF antagonism as a therapeutic strategy in SSc.
KEY WORDS: Connective tissue growth factor, The CCN family, Signal transduction, Matricellular regulatory molecule, Wound healing, Fibrosis, Tissue remodelling, Transforming growth factor-β, Fibroblast, Systemic sclerosis
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Biology of connective tissue growth factor: regulated growth factor and matricellular coordinator |
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TopAbstractBiology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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Connective tissue growth factor (CTGF), also know as CCN2, is a small cyteine-rich secreted multi-modular protein, and the founder member of the CCN (cyr61, ctgf, nov) family of immediate-early genes [1]. CTGF contains four conserved modules that resemble domains in other extraellular proteins and orchestrate many essential biological functions [2]. These modular, or more recently termed ‘matricellular’ regulatory proteins are expressed in response to growth factors, and other stimuli including hypoxia, shear stress and bio-mechanical deformation [2]. CTGF expression is primarily regulated at the level of transcription. TGF-β potently induces CTGF by the ‘classical’ Smad pathway via a Smad binding element located within the proximal promoter [2]. In development, CTGF is important for the formation of connective tissues (mesengenesis and skeletogenesis) and angiogenesis [1–3]. CTGF-deficient mice exhibit severe skeletal and ECM abnormalities due to impaired chondrogenesis, and die neonatally from respiratory failure [3]. CTGF is also involved in a diverse array of cellular processes, including proliferation, adhesion, ECM synthesis, and is important in adult wound healing [4].
The cellular effects of CTGF have been ascribed to activities encoded within the four component modular domains [1, 2, 4]. These modules encompass functional domains characteristic of other major regulatory proteins including an insulin-like growth factor binding (IGFB) domain (Module I), a chordin-like cysteine-rich domain (Module II), a thrombospondin type 1 repeat (Module III) and a C-terminal cystine-knot (Module IV) [1, 2, 4]. These domains promote interactions with membrane-associated proteins, ECM components and other growth factors. Examples of these include a region within Module III that binds low-density lipoprotein receptor-related protein (LRP) and integrins, Module IV contains both an integrin and a heparin binding domain and Module II interacts with growth factors, including TGF-β and BMP [5–7]. Furthermore, the C-terminal domain (Modules III and IV) appear to regulate fibroblast proliferation whereas the N-terminal domain (Modules I and II) mediates myofibroblast differentiation and collagen synthesis [8]. In other functions of CTGF in vitro, CTGF appears to act both directly and indirectly, with several studies showing that CTGF is required for, or acts in synergy with TGF-β in the induction of pro-fibrotic genes [2, 4, 7]. Signalling pathways and transcription factors directly stimulated by CTGF and mediating pertinent biological effects include p42/p44 MAP kinase, Akt/PKB, JNK and the Smad and NF-
B pathways. Direct activation of p42/44 MAP kinase is important in cell adhesion and ECM production [5], whereas, activation and nuclear localization of NF-
B, confers altered cell survival. Activation of the Smad pathway may involve synergy or interaction with TGF-β [7]. Here, CTGF may modulate the Smad pathway indirectly via interaction with TGF-β, thereby increasing receptor binding or by altering anti-Smad levels and prolonging TGF-β signalling [2, 7]. In SSc fibroblasts, the molecular mechanism by which the CTGF gene is regulated appears different from that of normal fibroblasts possibly accounting for the elevated and persistent expression in disease [2, 9]. Here, a polymorphic sequence within the proximal CTGF promoter appears responsible for elevated expression in certain SSc patients with defined clinical feature [9]. However, specifically how CTGF regulates the enhanced, pathological expression of collagen type I leading to fibrosis in SSc is still unclear.
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Relevance of CTGF to fibrosis |
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TopAbstractBiology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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The role of CTGF in fibrosis stems from investigations of common human diseases and animal models. CTGF has been found over-expressed in almost all human disorders, which exhibit excessive scarring and fibrosis as a facet of disease pathology. These include liver cirrhosis, keloids and hyperopic scars, diabetic nephropathy, atherosclerosis, cardiac disease, pulmonary hypertension and fibrosis, the tumour stroma of cancers and SSc [4]. In established animal models of fibrosis such as the tight skin mouse (Tsk1/+) and the bleomycin-induced tissue injury, elevated CTGF levels are also found associated with the development of fibrosis. These consistent associations of CTGF with fibrotic disease, has fuelled the notion that CTGF is an important molecular mediator of fibrosis. Several in vitro and in vivo studies have shown that addition of recombinant CTGF or gene over-expression can result in tissue scarring, or cause and augment the persistent production of collagen type I and lead to fibrosis. Adenoviral delivery of CTGF to the lungs of normal mice induced tissue scarring, suggesting the requirement of other factors to sustain fibrosis. Injection of TGF-β alone into the dermis of naïve mice resulted in the formation of transient granulation tissue, whereas serial injections of CTGF after TGF-β resulted in the development of persistent dermal fibrosis. These studies appeared to highlight the requirement of CTGF for the maintenance of fibrosis, but requiring prior induction by TGF-β. However, transgenic mice with high level of fibroblast-specific expression of CTGF develop a spontaneous fibrogenic phenotype including thickening of the dermis [10]. This data supports the notion that prolonged expression of CTGF alone might be sufficient to promote tissue fibrosis, although the precise mechanism(s) and molecular interplay between TGF-β and CTGF that induce and maintain the fibrogenic process remains a major challenge.
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CTGF as a potential biomarker for fibrosis |
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TopAbstractBiology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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Many studies have examined the potential value of measuring circulating levels of CTGF. In diabetic nephropathy, study of plasma and urinary levels has shown CTGF to be a risk marker of diabetic renal and renal vascular disease, and a predictor of end-stage disease and mortality. Plasma CTGF concentration has also been used as a novel diagnostic marker for cardiac disease, associating with patients exhibiting myocardial fibrosis and chronic heart failure [4]. Moreover in liver disease, serum CTGF levels were differentially raised in patients with fibrosis, chronic viral hepatitis and fully developed cirrhosis, suggesting that CTGF in this disease setting is a candidate marker of fibrosis and ongoing fibrogenesis in chronic liver diseases [2, 8]. In SSc, analysis of circulating levels of CTGF in sera revealed increased levels in SSc patients. These raised levels particularly of the N-terminal cleavage fragment correlated with the extent of skin disease and the severity of internal organ involvement such as pulmonary fibrosis [11]. In addition, further evaluation of CTGF cleavage products showed the presence of elevated levels of the N-terminal fragment of CTGF in the plasma and dermal interstitial fluid of SSc patients, which appeared to correlate with severity of skin disease and disease duration [2, 4].
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CTGF as an anti-fibrotic target |
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TopAbstractBiology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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In SSc, study of skin biopsies showed that CTGF mRNA and protein expression was more abundant in fibroblasts in the dermis of sclerotic lesions compared with early inflammatory lesions. In vitro studies using primary skin and lung fibroblasts isolated from SSc patients have also consistently identified CTGF as an over-expressed gene [2, 4]. Several approaches have been employed for antagonizing, blocking or inhibiting CTGF activity. In the context of SSc-associated lung disease, hepatocyte growth factor has been shown to down-regulate CTGF expression and pro-fibrogenic effects via specific signalling pathways. RNA interference studies targeting CTGF inhibits expression of the growth factor and suppressed the production of types I and III collagen in SSc fibroblasts, and indicated that CTGF is an upstream factor regulating ECM synthesis, particularly type I collagen. In the context of renal fibrosis, CTGF appears to contribute to the glomerular response to injury and mesangial cell activation, and in in vitro studies antibody blockade of CTGF significantly inhibited the TFG-β-induced expression of fibronectin by kidney mesangial cells. Moreover, in hepatic fibrosis down-regulation of CTGF expression by CTGF gene silencing using siRNA inhibited the accumulation of connective tissue proteins and attenuated the development of liver fibrosis in vivo. These data suggest a key role for CTGF in modulating ECM production, and promoting scarring and fibrosis in vivo. Interestingly, and taken together, these findings and those outlined above, appear to point to a critical role for CTGF in the development and maintenance of fibrosis rather than in the initiation of fibrogenic response in SSc. Strongly suggesting that silencing CTGF gene expression or antagonism of CTGF biological activity with for example antibody inhibition, might be a useful approach to combat the progression of fibrosis and a potential therapeutic approach for SSc.
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Acknowledgements |
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TopAbstractBiology of connective tissue...Relevance of CTGF to...CTGF as a potential...CTGF as an anti-fibrotic...AcknowledgementsReferences |
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Supplement: This paper forms part of the supplement entitled ‘Update in systemic sclerosis’. This supplement was supported by an unrestricted grant from Encysive.
Disclosure statement: D.A. has received speaker fees and travel support from Encysive Pharmaceuticals Ltd.
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