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Figure 7 | Arthritis Research & Therapy

Figure 7

From: Cyr61 is involved in neutrophil infiltration in joints by inducing IL-8 production by fibroblast-like synoviocytes in rheumatoid arthritis

Figure 7

Identification of binding sites essential for transcription of IL-8 in Cyr61 treated FLS. (A) Wild-type (IL-8WT) and mutant vectors (mAP-1, mC/EBPβ and mNF-κB) were cotransfected with the control vector pRL-TK into HSF cells for 4 hours, cultured for 20 hours and incubated for 2 hours in the presence or absence of Cyr61 (5 μg/ml). The luciferase activity relative to control was indicated after normalization with Renilla luciferase activity. Data represent the mean ± SEM of at least three independent experiments. * P <0.05, ** P <0.01. (B) FLS were stimulated with Cyr61 (5 μg/ml), and the c-Jun, C/EBPβ and p65 bound to the response element in the IL-8 promoter were detected by ChIP assay. It is noted that pre-incubation of cells with 093G9 (20 μg/ml) or control IgG (20 μg/ml) 1 hour before the Cyr61 stimulation suppressed the c-Jun, C/EBPβ and p65 DNA binding. The relative amount of IL-8 promoter DNA bound to c-Jun, C/EBPβ and p65 was detected by PCR and quantified by densitometric scanner. ChIP, chromatin immunoprecipitation; FLS, fibroblast-like synoviocytes; HSF, human skin fibroblasts; IgG, immunoglobulin G; SEM, standard error of the mean.

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