Background/Purpose: Systemic sclerosis (SSc) is a chronic fibrotic connective disease of unknown etiology that affects the skin and internal organs. Fibroblast growth factor receptor 3 (FGFR3) belongs to a family of 4 different FGF receptors. Alterations of FGF signaling receptors have been linked to a variety of skeletal dysformations and are also found in some forms of cancer. An FGFR3 isoform binds FGF-1 (acidic FGF) and FGF-9 and binding of one of these FGFs to FGFR3 induces homodimerization and activates the receptor tyrosine kinase function. The purpose of this study was to elucidate the role of FGF9/FGFR3 signaling in Systemic Sclerosis.

Methods: The expression of FGF9 and FGFR3 in skin tissue and in human dermal fibroblasts was determined by real-time PCR, Western blot and immunohistochemistry. Knock-down and overexpression strategies were used to evaluate the effect of both on fibroblast activation. The outcome of mice with fibroblast-specific knockout of FGF9 or non-conditional knockout of FGFR3 was evaluated in bleomycin-induced skin fibrosis; fibrosis induced by overexpression of a constitutively active TGF-β receptor I (TBRIact) and in the Tsk model. FGFR3 pharmacological inhibition with PD 173074 was evaluated in vitro as well as in murine models of fibrosis.  

Results: Our results show an upregulation of FGFR3 and in particular of its ligand FGF-9 in the skin of SSc patients. The upregulation of FGF9 was induced by TGFβ in vitro and in vivo and inhibition of TGFβ signaling blocked the upregulation of FGF9 in experimental fibrosis. SSc fibroblasts were more responsive to FGF-9 stimulation with enhanced activation of downstream mediators such as ERK and p38. Stimulation of fibroblasts with recombinant FGF9 or overexpression of a constitutively active FGFR3 induced myofibroblast-differentiation of resting fibroblasts, stimulated the collagen release and induced the expression of profibrotic mediators such as CTGF, ET-1 and IL4R. Inactivation of FGFR3, FGF9 or of the downstream mediators ERK and p38 reduced fibroblast activation and prevented the upregulation of pro-fibrotic target genes. Consistent with the induction of FGF9 by TGFβ and the profibrotic effects of FGF9, fibroblasts lacking FGF9 or FGFR3 were less sensitive to TGFβ stimulation. Inactivation of FGF9 or of its receptor FGFR3 exerted potent antifibrotic effects in several preclinical models. Knockout of FGF9 or FGFR3 ameliorated experimental fibrosis in bleomycin, TBRIact and Tsk animal model. Moreover, pharmacological inhibition of FGFR3 by the selective inhibitor prevented not only progression of fibrosis in experimental fibrosis, but also induced regression of pre-established fibrosis.

Conclusion: We demonstrate a novel role of FGF9/FGFR3 signaling in fibroblast activation and tissue fibrosis. FGF9/FGFR3 signaling is activated in SSc in a TGF-β dependent manner and induces several well-known profibrotic pathways to promote fibroblast-to-myofibroblast transition and collagen release. Pharmacologic or genetic inactivation of FGF9 or FGFR3 ameliorated fibroblast activation and fibrosis in several murine models.