Effects of S 36578-2 on HUVECs spread on vitronectin. The compound S 36578-2 was added to cells 3 hours after seeding in experimental medium. (A) Cells were processed for F-actin detection 30 minutes after the addition of S 36578-2. Scale bar represents 20 μm. Phase contrast micrographs were taken 24 hours after treatment. Scale bar represents 100 μm. Results from a representative experiment, repeated at least 3 times, are shown. (B) Floating and attached cells were collected 24 hours after treatment. Apoptosis was assessed by flow cytometry after annexin V and propidium iodide staining. Results of 1 of 2 experiments with similar results are shown. (C) PARP cleavage was assessed by Western blotting 24 hours after treatment of cells with S 36578-2. Where indicated, cells were treated for 1 hour with the caspase inhibitor Z-VAD-FMK before S 36578-2 was added. Erk1/2 was used as loading control. Results of 1 of 3 experiments with similar results are shown. (D) Procaspase-8, procaspase-9, cleaved caspase-3, and PARP were detected by Western blotting. Erk1/2 was used as loading control. Results of 1 of 12 experiments with similar results are shown.