Figure 2.

OA enhances NF-κB transcriptional activity. (A) Schematic representation of the two IL-6 promoter constructs and the pGAL4p65, pGAL4dbd, and pGAL4tkluc plasmids applied in the transfections of TF-1 cell line. Fragments of 122 (pIL6(-122)luc) and 60 (pIL6(-60)luc) basepairs, respectively, of the human IL-6 promoter are fused to the luciferase gene. Binding site for the transcription factors NF-κB is denoted above the largest construct. The Gal4-transactivator fusion proteins, pGal4p65 and pGal4dbd, are exclusively nuclear and are regulated independently of IκB. The reporter gene is under the control of five Gal4-binding sites. (B) Schematic representation of NF-κB- and p65-mediated promoter activity in the transiently transfected TF-1 cells. TF-1 cells were cultured as described in ‘Methods' and subsequently transiently transfected by means of electroporation with the respective plasmids. Six hours after transfection cells were stimulated for 24 h with medium or OA (30 ng ml⁻¹). Twenty-four hours after stimulation cells were lysed and analysed for the amount of produced luciferase protein. Basal promoter activity for each construct when treated with medium alone is set at 1. Mean fold induction and standard error of the mean represent six or more identical experiments. ^*P<0.05.