Fig. 4

Induction of caspase-related apoptosis in oral cancer cells by limocitrin. SCC-9 and SCC-47 cells were treated with limocitrin for 24 h. (A) Annexin-V/propidium iodide double staining for apoptosis detection. (B) Proportion of apoptotic cells. (C, D) Chromatin condensation was assessed using DAPI staining following the treatment of cells with limocitrin (0, 10, 20, and 40 μM) for 24 h. Scale bar = 50 µm. White arrows denote chromatin condensation. (E) Western blot analysis of cleaved caspases and PARP, with β-actin as the internal control. (F) Protein expression levels are displayed graphically. (G) Cells were pretreated with the caspase inhibitor, Z-VAD-FMK, and then with limocitrin for 24 h, with β-actin as the internal control. (H) Quantitation of cleaved caspase-related proteins and PARP is illustrated. All data are presented as mean ± standard deviation (n = 3). *p < 0.05, indicating statistical significance compared with the control group.