Perspectives on blockade of TGFβ overexpression
Authors: Y. Huang, W.A. Border, N.A. Noble
Journal: Kidney International (2006)

Many approaches to blocking profibrotic TGFβ overexpression are under way. Therapeutic targeting of TGFβ–Smad signaling holds promise for slowing or halting progressive renal disease. In this issue, Fukasawa et al., using the unilateral ureteral obstruction model, provide a new target for therapeutic intervention by identifying loss of the Smad corepressors Ski and SnoN as a mechanism that amplifies the profibrotic actions of TGFβ. Many approaches to blocking profibrotic TGFβ overexpression are under way. Therapeutic targeting of TGFβ–Smad signaling holds promise for slowing or halting progressive renal disease. In this issue, Fukasawa et al., using the unilateral ureteral obstruction model, provide a new target for therapeutic intervention by identifying loss of the Smad corepressors Ski and SnoN as a mechanism that amplifies the profibrotic actions of TGFβ. Progressive kidney disease is characterized by accumulation of extracellular matrix (ECM) in glomeruli and tubulointerstitium leading to loss of kidney function and kidney failure regardless of the underlying disorder. Slowing or stopping this ECM build-up is the biggest challenge facing nephrologists today. Thus a major effort has been the analysis of mechanisms involved in the pathogenesis of chronic renal fibrosis. Through understanding of these mechanisms, it might be possible to prevent the development of progressive renal insufficiency, thus minimizing the clinical, social, and economic impact of end-stage renal disease. The studies of Brenner and colleagues (reviewed by Neuringer and Brenner1.Neuringer J.R. Brenner B.M. Hemodynamic theory of progressive renal disease: a 10 year update in brief review.Am J Kidney Dis. 1993; 22: 98-104Abstract Full Text PDF PubMed Scopus (126) Google Scholar) were an early systematic analysis of factors that underlie the progression of renal disease. These studies demonstrated that hyperfiltration and increased intraglomerular pressure caused a progressive decrease of glomerular filtration, resulting in complete organ failure. Then, in 1990, three papers by Border et al. (reviewed by Border and Noble2.Border W.A. Noble N.A. Transforming growth factor beta in tissue fibrosis.N Engl J Med. 1994; 331: 1286-1292Crossref PubMed Scopus (3007) Google Scholar) suggested that TGFβ overexpression was a key mediator of fibrotic renal disease. As shown in Figure 1, this area of research has grown exponentially, so that now therapeutic targeting of TGFβ overexpression holds much hope for slowing or halting pathological ECM accumulation. The profibrotic actions of TGFβ include stimulation of ECM synthesis, inhibition of ECM degradation, and modulation of ECM receptor expression to facilitate cell–ECM interaction. Over a period of years, TGFβ overexpression has been shown to be a component of virtually every fibrotic disease in both humans and animal models. Additional basic research uncovered the existence of TGFβ isoforms and identified mechanisms of TGFβ activation and TGFβ signaling at the level of extracellular ligand activation and intracellular signal transduction. The identification of the regulation and dysregulation of post-receptor signaling shed new light on the pathways by which TGFβ contributes to fibrotic kidney disease, as Fukasawa et al.3.Fukasawa H. Yamamoto T. Togawa A. et al.Ubiquitin-dependent degradation of SnoN and Ski is increased in renal fibrosis induced by obstructive injury.Kidney Int. 2006; 69: 1733-1740Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar show in this issue. But first, some background. TGFβ is synthesized as an inactive precursor containing an amino-terminal prodomain. The prodomain, also termed latency-associated protein, is cleaved within the Golgi apparatus but remains non-covalently bound to TGFβ, thereby preventing TGFβ from binding to its receptors. This latent complex of TGFβ and latency-associated protein also associates with latent TGFβ-binding proteins before secretion from the cell. After secretion, most TGFβ is stored in the ECM as part of this inactive complex with latency-associated protein and latent TGFβ-binding proteins. The latent TGFβ complex can be activated into mature, biologically active TGFβ through the actions of proteases, such as plasmin, or by thrombospondin-1. After release from the latent complex, TGFβ dimers initiate their cellular effects by binding to cell membrane receptors. Three TGFβ receptors are involved in TGFβ signal transduction. TGFβ first binds to the type III TGFβ receptor, which then presents the ligand to the type II receptor or directly binds to the type II receptor. The type I receptor subsequently is recruited to form a stable receptor complex consisting of two receptors of each type, allowing phosphorylation of the glycine/serine-rich sequences by the type II receptor kinases. This phosphorylation activates the type I receptor kinases, resulting in autophosphorylation of the type I receptor and phosphorylation of certain members of the Smad family of signal-transducing proteins. Smad2 and Smad3 phosphorylated by the type I receptor homodimerize and then bind to Smad4. The heteromeric complex of Smad2 or Smad3 and Smad4 translocates to the nucleus, where it regulates the transcription of target genes.4.Massague J. How cells read TGF-beta signals.Nat Rev Mol Cell Biol. 2000; 1: 169-178Crossref PubMed Scopus (1650) Google Scholar Smad4 therefore seems to be critical for the Smad signaling pathway. Studies have shown that deficiency of Smad4 prevents TGFβ-induced collagen but not fibronectin expression, demonstrating that both Smad-dependent and Smad-independent signaling pathways mediate TGFβ-induced ECM overexpression.5.Tsuchida K. Zhu Y. Siva S. et al.Role of Smad4 on TGF-beta-induced extracellular matrix stimulation in mesangial cells.Kidney Int. 2003; 63: 2000-2009Abstract Full Text Full Text PDF PubMed Scopus (95) Google Scholar With the use of mouse embryo fibroblasts isolated from either Smad3 or Smad2 knockout (KO) mice, TGFβ-induced collagen type I and type III expression was ablated in Smad3 KO fibroblasts, but not in Smad2 KO fibroblasts.6.Wang W. Koka V. Lan H.Y. Transforming growth factor-β and Smad signaling in kidney diseases.Nephrology. 2005; 10: 48-56Crossref PubMed Scopus (309) Google Scholar A 900-gene chip microarray analysis further indicated that Smad3 is an essential mediator of TGFβ signaling.7.Yang Y.C. Piek E. Zavadil J. et al.Hierarchical model of gene regulation by transforming growth factor beta.Proc Natl Acad Sci USA. 2003; 100: 10269-10274Crossref PubMed Scopus (185) Google Scholar In db/db mice, a genetic model of type 2 diabetes, compared with control mice, increased nuclear localization of Smad3 in glomerular and tubular cells and increased binding of nuclear proteins to the Smad-binding element were seen.8.Hong S.W. Isono M. Chen S. et al.Increased glomerular and tubular expression of transforming growth factor-beta 1, its type II receptor, and activation of the Smad signaling pathway in the db/db mouse.Am J Pathol. 2001; 158: 1653-1663Abstract Full Text Full Text PDF PubMed Google Scholar Similar increases in nuclear localization of Smad3 and activation of the Smad-binding element were observed in kidneys of mice with streptozotocin-induced (type 1) diabetes, and these increases were prevented by administration of TGFβ-neutralizing antibodies. When Smad3 KO mice were treated with streptozotocin, diabetes-induced thickening of glomerular basement membrane, albuminuria, and upregulation of fibronectin and collagen IV expression were attenuated, which suggests that local activation of Smad3 is important in the development of diabetic nephropathy.9.Fujimoto M. Maezawa Y. Yokote K. et al.Mice lacking Smad3 are protected against streptozotocin-induced diabetic glomerulopathy.Biochem Biophys Res Commun. 2003; 305: 1002-1007Crossref PubMed Scopus (166) Google Scholar In unilateral ureteral obstruction, upregulation of TGFβ is thought to induce epithelial–mesenchymal transition of renal tubules that then go on to deposit ECM. After unilateral ureteral obstruction, Smad3 KO mice were protected from tubulointerstitial fibrosis, as evidenced by blocking of epithelial–mesenchymal transition and decreased monocyte influx and collagen accumulation. Primary cultures of renal tubular epithelial cells from Smad3 wild-type mice underwent epithelial–mesenchymal transition after TGFβ treatment, as evidenced by loss of E-cadherin expression, induction of α-smooth muscle cell actin expression, and induction of the expression of Snail (a potent repressor of transcription of the E-cadherin gene), none of which occurred in cells prepared from Smad3 KO mice.10.Sato M. Muragaki Y. Saika S. et al.Targeted disruption of TGF-beta1/Smad3 signaling protects against renal tubulointerstitial fibrosis induced by unilateral ureteral obstruction.J Clin Invest. 2003; 112: 1486-1496Crossref PubMed Scopus (697) Google Scholar These results provide clear evidence that activation of Smad3 mediates the profibrotic actions of TGFβ and is a critical feature of renal fibrosis. Three Smad transcriptional corepressors, namely Ski (Sloan-Kettering Institute proto-oncogene), SnoN (Ski-related novel gene, non-Alu-containing), and TGIF (TG-interacting factor), have been identified.11.He J. Tegen S.B. Krawitz A.R. et al.The transforming activity of Ski and SnoN is dependent on their ability to repress the activity of Smad proteins.J Biol Chem. 2003; 278: 30540-30547Crossref PubMed Scopus (90) Google Scholar Once bound to corepressors, activated Smads form transcriptionally inactive complexes. The amount of transcriptional corepressor present within a cell in a given circumstance may determine the ultimate outcome of a TGFβ response. In their new paper, Fukasawa et al.3.Fukasawa H. Yamamoto T. Togawa A. et al.Ubiquitin-dependent degradation of SnoN and Ski is increased in renal fibrosis induced by obstructive injury.Kidney Int. 2006; 69: 1733-1740Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar demonstrate a dramatic reduction of the Smad transcriptional corepressors Ski and SnoN at the protein level in the fibrotic kidney after obstructive injury. Because SnoN and Ski function as Smad transcriptional antagonists, their loss represents elimination of a crucial negative control mechanism for TGFβ action, potentially leading to unrestrained TGFβ signal transduction in the kidney under pathological conditions. The authors also show that SnoN and Ski proteins are reduced not by decreases in their mRNA expression but by accelerated degradation involving the ubiquitin–proteasome pathway. These results strongly suggest that loss of Smad corepressors is an important mechanism that amplifies profibrotic TGFβ signaling in this model. It remains to be seen whether the downregulation of Smad corepressors is involved in the progression of other forms of chronic renal disease. In summary, the profibrotic actions of TGFβ have been extensively studied for the past 15 years. Work has progressed from showing TGFβ overexpression in fibrotic diseases to uncovering a complex system where profibrotic TGFβ signaling may be dramatically amplified in the diseased kidney. This may explain why it is so difficult to completely block TGFβ signaling and halt the progression of chronic renal fibrosis. On the other hand, such a complex system also provides targets for intervention. The sheer number of available targets in this complex system offers hope that in the future it will be possible to prevent progressive renal disease by blocking the fibrotic effects of TGFβ overexpression.