Figure 3.

miR-3174 Inhibits Mitochondria-Dependent Apoptosis in GC Cells

(A) After infection of MKN45 cells with LV-miR-3174 or LV-miR-NC and BGC823 cells with ANTI-3174 or ANTI-NC, apoptotic rates were detected with flow cytometry. (B) Quantification of the apoptotic rate of each group is shown. (C) Caspase-3 activity was measured using a Caspase-3 activity detection kit in MKN45 cells transfected with LV-miR-NC or LV-miR-3174 or treated with Z-VAD-FMK after LV-miR-NC transfection and in BGC823 cells with or without miR-3174 suppression or exposure to Z-VAD-FMK after inhibition of miR-3174. (D) CCK-8 assays were performed to test cell viability after 48 hr culture. (E) Detection of the mitochondrial membrane potential by using JC-1 as a fluorescence probe. (F) Quantification of the fluorescence ratios converted into histograms. (G) Upper panel, western blot measurement of pan-capsase-3 (inactivated form of caspase-3), cleaved caspase-3 (c-caspase-3), Bax, and Bcl-2 protein levels in MKN45 or BGC823 cells. β-actin was used as an internal control. Lower panel, western blot analysis of cytochrome c (cyto.c) protein levels in cytosol or mitochondrial (mito) fraction of cells. β-actin, internal control in cytosol; cox IV, internal control in mitochondrial fragments. Graph represents mean ± SEM; *p < 0.05, **p < 0.01, and ***p < 0.001.