Ivermectin induces generation of ROS. OCI-AML 2 leukemia cells were treated with increasing concentrations of ivermectin overnight (A) or 6μM ivermectin for increasing incubation times (B). After incubation, ROS was detected by staining cells with Carboxy-H2DCFDA (final concentration 10μM) and flow cytometric analysis. Data represent the mean ± SD fold change in ROS production, compared with control. Representative experiments performed in triplicate are shown. Differences in change of ROS compared with control were analyzed by an unpaired t test. ***P < .001; **P < .005. (C) U937 and TEX leukemia cells and DU145 and PPC-1 prostate cells were treated with ivermectin at 6μM for 2 hours. After treatment, ROS generation was measured as described above. Data represent the mean ± SD fold change in ROS production compared with each of their buffer treated controls. Representative experiments performed in triplicate are shown. Differences in change of ROS compared with control were analyzed by an unpaired t test. ***P < .001. Primary AML cells (n = 3) and normal hematopoietic stem cells (PBSCs, n = 3) were treated with ivermectin (6μM) for 6 hours. After treatment, ROS generation was measured as described above. Data represent the mean ± SD fold change in ROS production compared with each of their buffer treated controls for experiments performed in triplicate. Differences in ROS production compared with control were analyzed by an unpaired t test. ***P < .001. (D) OCI-AML2 cells were treated simultaneously with ivermectin (3μM), the ROS scavenger, NAC (5μM), or the combination of NAC and ivermectin. After 48 hours of treatment, cell growth and viability were measured by the MTS assay. Data represent the mean ± SD percent viable cells from a representative experiment performed in triplicate. Differences in change of cell viability, compared with control, were analyzed by an unpaired t test. ***P < .001.
