Session Type: ACR Poster Session B
Session Time: 9:00AM-11:00AM
Background/Purpose: Systemic sclerosis (SSc) is a connective tissue disease characterized by systemic fibrosis. The dysregulation of transforming growth factor-β (TGF-β) signaling causes proliferation of myofibroblasts and results in the uncontrolled release of extracellular matrix. We have published that fucosyltransferase-1 (Fut1), an α-1,2 fucosyltransferase, plays an important role in rheumatoid arthritis synovial fibroblast proliferation. In this study, we examine the role of Fut1 in TGF-β receptor (TGF-βR) fucosylation, downstream signaling pathways, and target genes involved in scleroderma pathogenesis.
Methods: qPCR and ELISA were performed to assess the levels of Fut1 in SSc dermal fibroblasts and patient sera, respectively. To determine fucosylation of TGF-βR1, TGF-βR1 was immunoprecipitated from wild type (WT) dermal fibroblasts and immunoblotted with ulex europaeus agglutinin 1, which detects α-1,2 fucosylated protein. To confirm TGF-bR1 fucosylation, Fut1 was knocked down in SSc dermal fibroblasts using Fut1 shRNA. We evaluated Fut1 involvement in TGF-β signaling, myofibroblast differentiation, and TGF-b-associated target genes in WT and Fut1-/- dermal fibroblasts via immunofluorescence (IF), qPCR, and Western blotting. An in vivo wound healing model was performed with Fut1-/- and WT mice and wound healing was assessed over 8 days. To elucidate the role of Fut1 in an animal model of scleroderma, bleomycin was injected intradermally into Fut1-/- and WT mice for 9 days. Mice were euthanized and skin harvested for Masson’s trichrome staining and hydroxyproline assay, a quantitative measure of collagen.
Results: Fut1 mRNA and protein were significantly elevated in SSc compared to NL dermal fibroblasts and patient sera, respectively. We found via immunoprecipitation that TGF-βR1 is fucosylated by Fut1. We found less TGF-bR1 in SSc dermal fibroblasts when transduced with Fut1 shRNA, confirming the role of Fut1 in TGF-bR1 fucosylation. TGF-β-induced phosphorylation of Smad2, Erk1/2, and Jnk was markedly reduced in Fut1-/- dermal fibroblasts compared to WT, indicating that Fut1 is essential to TGF-βR1-mediated signaling pathways. Impaired transcription of myofibroblast differentiation inducing genes, Twist2 and Snail, was observed in Fut1-/- dermal fibroblasts. We found a significant decrease in α-smooth muscle actin in Fut1-/- compared to WT dermal fibroblasts as determined by IF and Western blotting, indicating a critical role for Fut1 in myofibroblast differentiation. In the wound repair model, Fut1-/- mouse wounds healed more slowly than WT mouse wounds. Fut1-/- mice developed significantly less bleomycin-induced skin fibrosis and collagen deposition compared to WT mice as determined by Masson’s trichrome staining and hydroxyproline assay, respectively.
Conclusion: Fut1 expression is elevated in SSc dermal fibroblasts and patient sera. Fucosylation of TGF-βR1 is critical to myofibroblast differentiation and TGF-β signaling. In addition, Fut1-/- mice exhibit delayed wound repair and diminished bleomycin-induced skin fibrosis, indicating an essential role for Fut1 in SSc pathology. Fut1 may be a novel therapeutic target for SSc.
To cite this abstract in AMA style:Stinson WA, Tsou PS, Du Y, Cui H, Cealey E, Lepore N, Ohara RA, Edhayan G, Arwani S, Morgan R, Khanna D, Fox DA, Amin MA. Fucosyltransferase-1 Mediated Fucosylation of TGF-βR1 Is Critical to TGF-β Signaling in Scleroderma and in Bleomycin-Induced Fibrosis [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/fucosyltransferase-1-mediated-fucosylation-of-tgf-%ce%b2r1-is-critical-to-tgf-%ce%b2-signaling-in-scleroderma-and-in-bleomycin-induced-fibrosis/. Accessed October 25, 2021.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/fucosyltransferase-1-mediated-fucosylation-of-tgf-%ce%b2r1-is-critical-to-tgf-%ce%b2-signaling-in-scleroderma-and-in-bleomycin-induced-fibrosis/