Anacardic acid represses NF-κB–dependent reporter gene expression induced by TNF and various plasmids. (A) Anacardic acid inhibits the NF-κB–dependent reporter gene expression induced by TNF. A293 cells were transiently transfected with a NF-κB–containing plasmid for 24 hours. After transfection, the cells were incubated with the indicated concentrations of anacardic acid for 4 hours and then treated with 1 nmol/L TNF for an additional 24 hours. The supernatants of the culture media were assayed for SEAP activity. Data are presented as mean (± SD). (B) Anacardic acid inhibits the NF-κB–dependent reporter gene expression induced by TNF, TNFR1, TRADD, TRAF2, NIK, IKK, p65, and TAK1/TAB1. Cells were transiently transfected with a NF-κB–containing plasmid alone or with the indicated plasmids. After transfection, cells were incubated with 25 μmol/L anacardic acid for 4 hours and then incubated with the relevant plasmid for an additional 24 hours. TNF-treated cells were incubated with 25 μmol/L anacardic acid for 4 hours and then treated with 1 nmol/L TNF for an additional 24 hours. The supernatants of the culture media were assayed for SEAP activity. Data are presented as mean (± SD). (C) Anacardic acid inhibits the COX-2 promoter activity induced by TNF. Cells were transiently transfected with a COX-2 promoter linked to the luciferase reporter gene plasmid for 24 hours and treated with the indicated concentrations of anacardic acid for 4 hours. Cells were then treated with 1 nmol/L TNF for an additional 24 hours, lysed, and subjected to a luciferase assay. Data are presented as mean (± SD).