GRP increases IL-8 luciferase activity, and Ets1 regulates GRP-induced IL-8 secretion in SK-N-SH cells. (A) For IL-8 promoter activity assays, cells were seeded in 24-well plates, transfected with IL-8 luciferase plasmid DNA, and then treated with GRP (10-7 M). (B) For IL-8 secretion assay, cells in 12-well plates (1 x 105 cells/well) were treated with GRP (10-7 M) for 24 hours. The culture supernatants were collected, and IL-8 in the culture supernatant was measured by ELISA (data represent triplicate determinations; mean ± SEM; *P < .05 vs control). (C) Ets1 siRNA transfection yielded a significantly decreased Ets1 protein expression by Western blot analysis. (D) Ets1 knockdown using siRNA significantly decreased IL-8 secretion. In addition, GRP-stimulated increase in IL-8 secretion was markedly attenuated in SK-N-SH cells transfected with Ets1 siRNA vector when compared to control cells transfected with control siRNA vector (mean ± SEM; *P < .05 vs control siRNA vector; †P < .05 vs no GRP). All data represent triplicate determinations.