Bone Morphogenetic Protein Antagonist Gremlin-1 Increases Myofibroblast Transition in Dermal Fibroblasts: Implications for Systemic Sclerosis.

OBJECTIVE: Systemic Sclerosis is an autoimmune connective tissue disease which results in fibrosis of the skin and lungs. The disease is characterized by activation of myofibroblasts but what governs this is unknown. Gremlin-1 is a BMP antagonist that is developmentally regulated and we sought to investigate its role in Systemic Sclerosis. METHODS: Dermal fibroblasts were transfected with Grem1pcDNA3.1 expression vectors or empty vectors. Various markers of myofibroblasts were measured at the mRNA and protein levels. Scratch wound assays were also performed. Media Transfer experiments were performed to evaluate cytokine like effects. Various inhibitors of TGF-ß signaling and MAPK signaling were used post-transfection. siRNA to Gremlin-1 in SSc dermal fibroblasts were performed to evaluate the role of Gremlin-1. Different cytokines were incubated with fibroblasts and Gremlin-1 measured. Bleomycin was used as model of fibrosis and immunohistochemistry performed. RESULTS: Overexpression of Gremlin-1 was achieved in primary dermal fibroblasts and lead to activation of quiescent cells to myofibroblasts indicated by collagen and α-Smooth muscle actin. Overexpression also led to functional effects. This was associated with increased TGF-ß1 levels and SBE luciferase activity but not increased Thrombospondin-1 expression. Inhibition of Gremlin-1 overexpression cells with antibodies to TGF-ß1 but not isotype controls led to reduced collagen and various TGF-ß pathway chemical inhibitors also led to reduced collagen levels. In SSc cells siRNA mediated reduction of Gremlin-1 reduced collagen expression and CTGF gene and protein levels in these cells. IL-13 did not lead to elevated Gremlin-1 expression nor did IL-11. Gremlin-1 was elevated in an animal model of fibrosis compared to NaCl-treated mice. CONCLUSION: Gremlin-1 is a key regulator of myofibroblast transition leading to enhanced ECM deposition. Strategies that block Gremlin-1 maybe a possible therapeutic target in fibrotic diseases such as SSc.

Fibroblast growth factor receptor 3 activates a network of profibrotic signaling pathways to promote fibrosis in systemic sclerosis.

Aberrant activation of fibroblasts with progressive deposition of extracellular matrix is a key feature of systemic sclerosis (SSc), a prototypical idiopathic fibrotic disease. Here, we demonstrate that the profibrotic cytokine transforming growth factor ß selectively up-regulates fibroblast growth factor receptor 3 (FGFR3) and its ligand FGF9 to promote fibroblast activation and tissue fibrosis, leading to a prominent FGFR3 signature in the SSc skin. Transcriptome profiling, in silico analysis and functional experiments revealed that FGFR3 induces multiple profibrotic pathways including endothelin, interleukin-4, and connective tissue growth factor signaling mediated by transcription factor CREB (cAMP response element-binding protein). Inhibition of FGFR3 signaling by fibroblast-specific knockout of FGFR3 or FGF9 or pharmacological inhibition of FGFR3 blocked fibroblast activation and attenuated experimental skin fibrosis in mice. These findings characterize FGFR3 as an upstream regulator of a network of profibrotic mediators in SSc and as a potential target for the treatment of fibrosis.