Treatment with PDA-66 induces apoptosis via cleavage of caspases. (A) Cells were treated with PDA-66 for up to 72 h and stained with Annexin V FITC and Propidium iodide (PI). Rates of early apoptotic (FITC+, PI-) and late apoptotic and necrotic (FITC+, PI+) cells were measured by flow cytometry. The upper diagrams display the rate of apoptotic cells after 48 h (left) and 72 h (right). The lower diagrams show the results for necrosis measurement, respectively. Significant induction of apoptosis could be observed in all cell lines after 48 h of incubation as well as tendential induction of necrosis at both points of time. Results are displayed as the mean + SD of three independent experiments. *Significant treatment effect vs. DMSO control, α = 0.05. (B) Cells were treated with different concentrations of PDA-66 and total cell lysates (25 μg) were analyzed by Western blot to detect cleavage of Caspase 3, 7 and PARP. GAPDH was used as loading control. Exemplary results of PDA-66 treated SEM cells after 24 and 48 h are displayed. Induction of apoptosis was confirmed by an increase of the cleaved forms of Caspase 3, 7 and PARP.