Impaired Smad7-Smurf–mediated negative regulation of TGF-β signaling in scleroderma fibroblasts

253 Introduction Systemic sclerosis or scleroderma is an acquired disorder that typically results in fibrosis of the skin and internal organs (1). Although the pathogenesis of this disease is unclear, it includes inflammation, autoimmune attack, and vascular damage, leading to the activation of fibroblasts (2, 3). The reason for the presence of abnormal fibroblasts in scleroderma is not yet known, but it is possible that they develop from a subset of cells that have escaped normal control mechanisms (4, 5). TGF-β1 is a multifunctional cytokine that regulates the growth, differentiation, and function of various cell types (6). The principal effect of TGF-β1 on mesenchymal cells is its stimulation of ECM deposition. TGF-β1 has been shown to increase the expression of collagen types I, III, VI, VII, and X, fibronectin, and proteoglycans (4, 7–12). Stimulation of ECM production by TGF-β1 is further enhanced by its inhibitory effect on matrix degradation, decreasing the synthesis of proteases and increasing levels of protease inhibitors (13). Although the mechanism of dermal fibroblast activation in scleroderma is presently unknown, many of the characteristics of scleroderma fibroblasts resemble those of healthy fibroblasts stimulated by TGF-β1 (13, 14), suggesting that the dermal fibroblast activation in scleroderma may be a result of stimulation by autocrine TGF-β. This notion is supported by our recent findings that (a) scleroderma fibroblasts express elevated levels of TGF-β receptors, and this correlates with elevated levels of α2(I) collagen mRNA (15); and (b) the blockade of TGF-β signaling with anti–TGF-β Ab’s or anti–TGF-β1 antisense oligonucleotide abolished the increased expression of human α2(I) collagen mRNA in scleroderma fibroblasts (16). TGF-β1 initiates signaling through the ligand-dependent activation of a complex of heterodimeric transmembrane serine and threonine kinases consisting of TGF-β receptor type I (TβRI) and TGF-β receptor type II (TβRII). Upon binding TGF-β, the receptors rotate within the complex, resulting in the phosphorylation and activation of TβRI by the constitutively active and autophosphorylated TβRII. TGF-β1 signals from the receptor to the nucleus using a set of Smads proteins. The activated TβRI directly phosphorylates Smad2 and Smad3. Once activated, Smad2 and Smad3 associate with Smad4 and translocate to the nucleus, where such a complex regulates transcriptional responses of target genes together with additional DNA-binding cofactors (17). The inhibitory Smad7 associates with ligand-activated TβRI and interferes with the phosphorylation of Smad2 and Smad3 by preventing their interaction with activated TβRI (18). Since the expression of Smad7 is induced by TGF-β1, Smad7 inhibits TGF-β signaling by a negative feedback system (19, 20). Recently, Smad7 has Impaired Smad7-Smurf–mediated negative regulation of TGF-β signaling in scleroderma fibroblasts