Figure 4

Forskolin augmented radiation-induced apoptosis by inhibiting ATM activation in the mouse lung. (A) Effects of forskolin on the phosphorylation of PP2A B56δ and ATM in B16F10 mouse melanoma cells. Mouse melanoma cells were pretreated with 40 μM forskolin for 30 min and irradiated with γ-rays (5 Gy). After incubation for 30 min, the phosphorylation of PP2A B56δ and ATM were analyzed by western blotting. (B) Effects of forskolin on the phosphorylation of PP2A B56δ and ATM in the mouse lung. (C) Effect of forskolin on apoptosis in the mouse lung analyzed by western blotting. (D) Effect of forskolin on apoptosis in the mouse lung analyzed by the TUNEL assay. Four-week-old male BALB/c mice (20 g, n = 6) were injected intraperitoneally with forskolin (20 μg/g) or DMSO, and after 6 h, the mice were exposed to whole body γ-ray irradiation (10 Gy). After incubation for 30 min, the animals were sacrificed, and the lung tissues were excised and homogenized for western blot analysis of ATM and PP2A. For apoptosis analysis, the animals were sacrificed at 24 h after irradiation, and the lung tissues were excised. A portion of the lung tissues were homogenized and analyzed for cleavage of caspase-3 and PARP by western blotting. The other portion of the tissue was immediately fixed with formaldehyde and examined by the TUNEL assay. The arrows indicate the stained cells undergoing apoptosis.