PA4 enhances ROS generation to induce lymphoma cell apoptosis. (A) Summary of gene-set enrichment analysis reveals the overrepresentation of indicated pathways in PA4 treated group. Genes are ranked into an ordered list based on relative expression in control and PA4-treated H9 cells. (B) Cells were treated with indicated dosages of PA4 for 24 hours and stained with annexin V and 4′,6-diamidino-2-phenylindole for apoptosis analysis. Data are shown as mean ± SD, n = 3. ∗∗P < .01 vs DMSO treatment (C) and H9 wild-type and H9^{CAMK2G−/−} cells (D). (E) Cells were treated with DMSO or 1 μM PA4 for 24 hours and stained with 2′,7′-dichlorodihydrofluorescein diacetate (H₂DCFH-DA) for intracellular ROS, and MitoSOX for mitochondrial superoxide analysis. Data are shown as mean ± SD, n = 3. ∗∗P < .01 and ^##P < .01 vs H9 + DMSO treatment. (F) Representative western blots for ER stress markers after PA4 treatment in wild-type H9 and H9^{CAMK2G−/−} cells for 24 hours. CHOP, C/EBP homologous protein; FDR, false discovery rate; G2M, cell cycle 2 phase to M phase transition; PARP, poly-(adenosine 5′-diphosphate–ribose) polymerase.