Fig. 6. SUPT5H depletion promotes apoptosis, generates ROS, and change mitochondrial membrane potential (MMP) in MDA-MB-231. (A) Flow cytometric analysis demonstrated the levels of cell apoptosis in MDA-MB-231 cells. (B) Western blot analysis of apoptosis regulatory proteins in MDA-MB-231 treated cells with siRNA-SUPT5H or siRNA-scr (representative blots are shown from two independent experiments). (C) qRT-PCR analysis of the expression of genes related to apoptosis. (D) SUPT5H depletion promotes ROS production and the levels of ROS were diminished in presence of N-acetyl-L-cysteine (NAC). Scale bar, 100µm and magnification, 20X. (E) Flowcytometric analysis of the change of MMP in SUPT5H silenced MDA-MB-231 cells using JC-1 dye. (F) Treatment of NAC reagent to SUPT5H depleted MDA-MB-231 cells rescues the cells from apoptosis as indicated by flow cytometric analysis. The mean ± was shown in bar plots (n=3). *p<0.05, **p<0.01, ***p<0.001, not significant, ns.