Figure 3. Effect of avicin D on GRE-dependent gene expression.

(A) A549 cells transfected with p (GRE)₂-50huIL6P-luc+ were treated with avicin D (1 µM) or Dex (1 µM) for 2–16 hrs. Luciferase activity was measured in cell lysates, and normalized for TK renilla luminescence as described in the methods. (B) Effect of avicin D and RU486 on Dex-induced luciferase activity. A549 cells transfected with p (GRE)₂-50huIL6P-luc+ were pre-treated with avicin D (1 µM) or RU486 (1 µM) for 2 hrs, prior to being exposed to Dex (1 µM) or avicin D (1 µM) for 16 hrs. Luciferase activity was measured in cell lysates, and normalized for TK renilla luminescence as described in the methods. Luciferase activity has been expressed as relative luciferase units (RLU). (C) A549 cells were treated with Avicin D (1 µM) for 0–120 min or with Dex (1 µM) for 60 min. Western blot analysis of the nuclear extracts was performed using anti-phospho GR (Ser211) antibodies. Actin levels have been shown as a loading control.