15(S)-HETE induces HMG-CoA reductase expression in HDMVECs. (A-B) Quiescent HDMVECs were treated with and without 0.1μM 15(S)-HETE for the indicated time periods and either total cellular RNA was isolated or cell extracts were prepared. The RNA was analyzed for HMG-CoA reductase mRNA levels by QRT-PCR (A) and the cell extracts were analyzed for HMG-CoA reductase protein levels by Western blotting (B). (C-D) Quiescent HDMVECs were treated with 0.1 μM 15(S)-HETE in the presence or absence of 20 μg/mL actinomycin D or 10 μg/mL cycloheximide for 2 hours and HMG-CoA reductase mRNA and protein levels were measured as described in panels A and B, respectively. (E) Quiescent HDMVECs that were pretreated with actinomycin D (20 μg/mL) for 2 hours were treated with and without 15(S)-HETE (0.1μM) for 10 minutes and cell extracts were prepared. An equal amount of protein from control and each treatment was immunoprecipitated with anti-Farnesyl antibodies and immunocomplexes were analyzed by Western blotting for Rac1 using anti-Rac1 antibodies. Total cellular Rac1 levels were measured by Western blotting. The bar graphs in panels A, B and C represent the mean ± SD values of 3 independent experiments. *P < .01 versus control; **P < .01 versus 15(S)-HETE. HMG-CoAR indicates HMG-CoA reductase.