靶向siRNA DKK-1对类风湿关节炎患者滑膜细胞影响的研究

Objective Investigate the change of fibroblast-like synoviocytes (FLSs) derived from rheumatoid arthritis (RA) to small interfering RNA (siRNA) targeting dickkopf-1 (DKK-1). Methods Six siRNAs targeting DKK-1 were transfected into FLSs of patients suffering from RA. IL-1β, IL-6, IL-8, MMP2, MMP3, MMP9, TGF-β1, TGF-β2 and MCP-1 levels in the cell culture supernatant were detected by ELISA. Invasion assay and 3H incorporation assay were utilized to investigate the effects of siRNA targeting DKK-1 on FLSs invasion and cell proliferation, respectively. Western blot and radioimmunoassay were performed to analyze the change of NF-κB, IRAK1, ERK1, JNK and β-catenin in FLSs. Data were expressed as arbitrary units after normalization to levels of β-actin. Meanwhile, DKK-1 monoclonal antibody was employed as a positive control. Results 1. DKK-1 targeting siRNA inhibited the expression of DKK-1 in FLSs (p <0.01) and led to significant reduction of IL-6, IL-8, MMP2, MMP3 and MMP9 in FLSs comparing with the control group (p <0.01); 2. DKK-1 targeting siRNA also inhibited the proliferation and invasion of FLSs (p <0.05); 3. Important molecules of pro-inflammatory signaling pathway in FLSs, including NF-κB, IRAK1, ERK, JNK, were decreased by the inhibition of the expression of DKK-1 in FLSs. In contrast, β-catenin, a pivotal down-stream molecule of Wnt signaling pathway was increased. Conclusion By inhibiting DKK-1, we were able to inhibit the proliferation, invasion and pro-inflammatory cytokine secretion of FLSs derived from RA, which indicates that siRNA targeting DKK-1 is a promising novel method for anti-inflammatory and delay of joint destruction in RA.