Background/Purpose: Glycosylation is a common post-translational modification of proteins in eukaryotes. Fucosylated glycans are synthesized by fucosyltransferases (futs). We previously reported that sialyl Lewis^x, synthesized by futs, is involved in angiogenesis. Fucosyltransferase 1 (fut1) is an α(1,2)-fucosyltransferase responsible for synthesis of the H blood group and Lewis^y antigens. We also reported that soluble H and Lewis^y antigens are mediators of inflammatory cell adhesion and angiogenesis. However, a direct role of α(1,2)-linked fucose in rheumatoid arthritis (RA) has not been demonstrated.

Methods: Assay of total α(1,2)-linked fucosylated proteins in RA, osteoarthritis (OA) and normal (NL) synovial tissue (ST) homogenates were performed by enzyme-linked immunosorbent assay (ELISA). 2-fucosyl-lactose bovine serum albumin (2’FL-BSA) was used as a standard. We previously showed that fut1 gene deficient mice in which KRN arthritis has been induced have less joint monocyte chemotactic protein (MCP)-1/CCL2 and interleukin (IL)-1β than wild type mice. Hence, we measured IL-1β and MCP-1/CCL2 in RA, OA and other inflammatory disease synovial fluids (SFs) by ELISA to determine if these cytokines have been modified. We also examined tumor necrosis factor (TNF)-α for fucosylation, as TNF-α is a central cytokine in RA pathology. To determine whether fut1 was expressed by NL and RA synovial fibroblasts, real time polymerase chain reaction (RT-PCR) was performed. To block the expression of fut1, RA synovial fibroblasts were transfected with fut1 small interfering RNA (siRNA). After treatment with fut1siRNA, RA synovial fibroblasts were stimulated with TNF-α for 1 hour. We then measured the expression of cytokines important in RA. MCP-1/CCL2, epithelial neutrophil-activating protein 78 (ENA-78)/CXCL5 and vascular endothelial growth factor (VEGF) mRNA were measured by RT-PCR.

Results: Total α(1,2)-linked fucosylated proteins in RA ST were significantly higher compared to OA STor NL ST [mean ± SEM 74 ± 21 ng/ml (n=5), 25 ± 4 ng/ml (n=7) and 29 ± 6 ng/ml (n=9), respectivel, p<0.05]. Specifically, α(1,2)-linked fucosylated IL-1β and MCP-1/CCL2 in RA synovial fluids were 6 ± 2% of total fucosylated proteins (n=9) and 20 ± 4% of total fucosylated proteins (n=12), respectively. In addition, α(1,2)-linked fucosylated TNF-α in RA synovial fluids was significantly higher than in OA and other inflammatory disease synovial fluids [950 ± 220 pg/ml (n=22), 150 ± 90 pg/ml (n=13) and 30 ± 20 pg/ml (n=11), respectively, p<0.05]. We also found that fut1 messenger RNA (mRNA) in cultured RA synovial fibroblasts was significantly elevated compared to NL synovial fibroblasts (p<0.05). Additionally, RA synovial fibroblasts transfected with fut1 siRNA had significantly lower mRNA expression for MCP-1/CCL2, ENA-78/CXCL5 and VEGF than did control siRNA transfected synovial fibroblasts.

Conclusion: These data show that α(1,2)-linked fucosylated proteins are upregulated in RA ST compared to other STs. We also show that fut1 in RA synovial cells is necessary for production of variety of cytokines. Targeting fut may be a novel way to alter joint cytokine production.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/%ce%b112-linked-fucosylated-cytokines-are-upregulated-in-rheumatoid-arthritis/