ROS production in response to combined CI-1040 and retinoid treatment. OCI-AML3 cells were pretreated with the MEK inhibitor CI-1040 (0.5 μM) for 30 minutes and subsequently exposed to either ATRA (1 μM) or 9-cis RA (0.1 μM). At the indicated time points, control and treated cells were then incubated with DHE for 15 minutes, and its oxidation to the fluorescent product ethidium was then monitored by flow cytometric analysis. (A) Primary data from 1 experiment representative of at least 3 performed with superimposable results. The percentage of ROS-positive cells after 96 hours of exposure to the indicated treatments is shown in the figure. (B) Results are expressed as the net ROS induction (percentage of ROS-positive cells in treated samples minus percentage of ROS-positive cells in control samples) and represent the average ± SD of at least 3 independent experiments. (C) Apoptosis was evaluated by flow cytometric analysis of FITC-conjugated annexin V binding, while simultaneously assessing membrane integrity by PI exclusion. Results are expressed as the net apoptosis induction (percentage of apoptosis in treated cells minus percentage of apoptosis in control cells) and represent the average ± SD of at least 3 independent experiments.