Celastrol kills p53-null H1299 cells via ROS-mediated JNK activation.

<p>7a. Celastrol enhances HIF-1α expression and induces Akt activation in H1299 cells. H1299 cells were treated with the indicated doses of Celastrol for 6 h, and the proteins were detected by western blotting. 7b. Celastrol time-dependently induces ROS accumulation in H1299 cells. H1299 cells were treated with 4 µM Celastrol for the indicated times. The levels of ROS were measured by DCFH-DA staining and subsequently assayed by flow cytometry. 7c. Suppression of ROS-induced JNK activity could prevent Celastrol-mediated H1299 cell death. H1299 cells were pretreated with either the ROS scavenger 5 mM NAC or the JNK kinase inhibitor 40 µM SP600125 for 1 h and then treated with or without 4 µM Celastrol under normoxia for 24 h. Cell death was examined by microscopy (200×) and measured by Annexin-V/PI staining and flow cytometry. The proteins were detected by western blotting.