Figure 2. Synergistic effects of PARP inhibitors and cisplatin on DNA damage responses and clonogenic survival. (A and B) Effects of cisplatin (CDDP, 20 µM), PJ34 hydrochloride hydrate (PJ, 30 µM) or CEP 8983 (CEP, 10 µM), alone or in combination on DNA damage foci. Six hours after exposure to the indicated drug combinations, A549 cells were fixed, permeabilized and stained for the visualization of phosphorylated histone H2AX (γH2AX) in the presence of 2 µM Hoechst 33342 (H33342), which was used for nuclear counterstaining. Representative fluorescence microphotographs are shown in (A), and their quantitation (as the percentage of cells bearing > 5 γH2AX foci per nucleus) are shown in (B). (C and D). Effect of CDDP, PJ and/or CEP on clonogenic survival. After incubation with the indicated agents (1 µM CDDP, 5 µM PJ, 1 µM CEP) for 48 h, cells were cultured in drug-free medium, and colonies were rendered visible 10 d later by crystal violet staining. Representative photographs are shown in (C), and the frequency of colonies (referring to the untreated control as 100% value) is shown in (D). Results are means of triplicates ± SD. ^§p < 0.05 (Student’s t-test), compared with the sum of the cytotoxic effects caused by each agent alone.