Mitochondrial pathway of the lysine demethylase 5C inhibitor CPI-455 in the Eca-109 esophageal squamous cell carcinoma cell line

Figure 1 Eca-109 cell viability following treatment with CPI-455 at different times and concentrations. Data are expressed as means ± SE (n = 3). ^aP < 0.05 vs control.

Figure 2 Changes in Eca109 cells after 48 h culture with and without CPI-455 treatment. Left panels show the morphology of the mitochondria of Eca109 cells. A: Arrows indicate mitochondria with complete and clear structure. B: Arrows indicate mitochondria with a fuzzy structure and weak staining of cells. Right panels show TUNEL assay results (bar = 10 μm) and laser confocal scanning microscopy × 600 of nuclear staining and DNA fragmentation of Eca109 cells C: without, and D: with CPI-455 treatment. Data are reported as means ± SE (n = 3).

Figure 3 Eca-109 induced by CPI-455 is dependent on the generation of reactive oxygen species. A: Eca109 untreated (0 h); B: Eca-109 induced by CPI-455 for 24 h; C: Eca-109 induced by CPI-455 for 48 h; D: Eca-109 induced by CPI-455 for 72 h. Data are reported as means ± SE (n = 3). ROS: Reactive oxygen species.

Figure 4 Mitochondrial membrane potential assayed by flow cytometry. A: Eca109 untreated (0 h); B: Eca109 treated with CPI-455 for 48 h; C: Gray scale results of membrane potential energy of mitochondria (Blue: Results of the first assay; Red: Results of the second assay; Green: Results of the third assay). Data are reported as means ± SE (n = 3). ^aP < 0.05 vs control.

Figure 5 Western blot assays of p53, Bax, lysine-specific demethylase 5C, cleaved Caspase 3, and cleaved Caspase 9 protein expression in Eca-109 cells. Lane 1: Eca109 untreated (0 h); Lane 2: Eca109 induced by CPI-455 for 24 h; Lane 3: Eca109 induced by CPI-455 for 48 h; Lane 4: Eca109 induced by CPI-455 for 72h. KDM5C: Lysine-specific demethylase 5C; GAPDH: Glyceraldehyde-3-phosphate dehydrogenase.

Figure 6 Densitometry analysis of the immunoblotting data of P53, Bax, lysine-specific demethylase 5C, cleaved Caspase 3, and cleaved Caspase 9 proteins in Eca-109 cells. Data are reported as means ± SE (n = 3). ^aP < 0.05 vs control; ^bP < 0.01 vs control.