Thrombin stimulation of Egr-1 promoter activity in primary endothelial cells is mediated by an ERK1/2 MAPK–dependent pathway.

HUVECs were transiently transfected with 0.5 μg of full-length wild-type 1200-bp Egr-1-Luc plasmid, serum-starved in 0.5% FBS for 18 hours, treated in the absence (Control) or presence of 1.5 U/mL thrombin for 6 hours, and harvested for luciferase activity. Where indicated, the transfected cells were preincubated with 20 μM PD98059 (PD), 3 μM SB203580 (SB), 5 μM BIM, or 10 μM LY294002 (LY) for 10 minutes. The results show the means and standard deviations of luciferase light units (relative to untreated cells) obtained in triplicate from 4 independent experiments. Luciferase light units were corrected for transfection efficiency as described in “Materials and methods.”