Antioxidant effect maps to the NF-κB element. (A) Serial 5′ deletions of the IL-8 promoter analyzed for their DMSO-sensitivity. Triplicate cell cultures were transfected with the indicated IL-8/LUC reporter plasmids and an SV40/alkaline phosphatase plasmid as an internal control. Cells were unstimulated/stimulated with 20 ng/mL TNF in the absence or presence of 2% (vol/vol) DMSO. Fold-induction of the Luciferase activity of stimulated cells is shown (P < .0001 for –99, -162, -1498: nonpretreated vpretreated with 2% (vol/vol) DMSO). (B) DMSO-sensitivity of IL-8 promoter point mutations. Triplicate cell cultures were transfected with the indicated IL-8/LUC reporter plasmids and an SV40/alkaline phosphatase plasmid as an internal control. Cells were unstimulated/stimulated with 20 ng/mL TNF in the absence/presence of 2% (vol/vol) DMSO. Fold-induction of the Luciferase activity of stimulated cells is shown (P < .0001 for nonpretreatedv DMSO-pretreated, except ▵NF-κB). (C) DMSO effects on IL-8 multimers. Multimers of NF-κB, NF-IL6, and AP-1 sites ligated upstream of an inert hIL-8 TATA box were analyzed for their DMSO sensitivity. Triplicate cell cultures were transfected with the indicated IL-8/LUC reporter plasmids and a SV40/alkaline phosphatase plasmid as an internal control. Cells were untreated or stimulated with 20 ng/mL TNF in the absence or presence of 2% (vol/vol) DMSO. Fold-induction of stimulated Luciferase activity is shown (for NF-κB multimer nonpretreated v DMSO-pretreated, P < .0001).