Figure 1. R9-SOCS3-KIR suppressed the C5a-mediated induction of the NF-kB promoter. A: The ARPE-19 cells in a serum-free medium were treated with R9-SOCS3-KIR or the control peptide (20 μM) for 1 h, followed by treatment with C5a (50 ng/ml) for 4 h. The cells were then co-transfected with a mixture of plasmids with NF-kB promoter-driven firefly luciferase and another plasmid with thymidine kinase-driven Renilla luciferase as an internal control and grown for 24 h in a 1% FBS-containing medium. The cell lysates were harvested to measure the relative luciferase units using a dual luciferase kit from Promega. ****, p<0.0001. Error bars indicate the standard deviation. Abbreviations: K, R9-SOCS3-KIR; Ctrl, R9-SOCS3-KIR-scrambled peptide; UT, untreated. B. R9-SOCS3-KIR suppressed the C5a-induced nuclear translocation of the NF-κB subunit p65. The ARPE-19 cells were grown overnight in eight-well plates, placed in a serum-free medium, and treated with R9-SOCS3-KIR or its control peptide at 20 μM for 1 h, followed by the addition of C5a (50 ng/ml) for 30 min. The cells were stained with an antibody to p65, Cy3-conjugated secondary antibody, and DAPI and then imaged in a fluorescence microscope using the same exposure time and light intensity. Abbreviations: K, R9-SOCS3-KIR; Ctrl, R9-SOCS3-KIR-scrambled peptide; UT, untreated. Scale bar: 50 μm.