Figure 1. Induction of HO-1 protein and AMPK activation by OA-NO₂.

A) BAECs were incubated with OA-NO₂ at the indicated concentrations or with BSA (vehicle) for 16 h, and western blot analysis was performed as described in Materials and Methods to detect HO-1 protein expression and AMPK phosphorylation at Thr172. The blot is representative of those obtained from three separate experiments. Corresponding densitometric analyses of phosphorylated AMPK and ACC are shown. *p<0.05 vs. control. B) BAECs were incubated with 2.5 µM OA-NO₂ for the indicated times, and western blotting was performed as above. The blot is representative of three blots obtained from three separate experiments. *p<0.05 vs. corresponding control. C) Confluent BAECs were exposed to vehicle or OA-NO2 (2.5 µM) for 16 h. AMPKα was immunoprecipitated from cell lysates (1 mg) with a specific antibody. AMPK activity was assayed by ³²P-ATP incorporation into the SAMS peptide. *p<0.05 vs. control. D) BAECs were incubated with the indicated concentrations of OA for 16 h. Western blotting was performed as described in Materials and Methods. E) BAECs were infected with Ad-DN-AMPK (MOI = 50) or Ad-GFP (control). Infected and non-infected cells were treated with 2.5 µM OA-NO₂ for 16 h. AICAR and metformin were used as positive controls. The blot is representative of three blots obtained from three separate experiments.