Fig. 4: TTK knockdown promotes mitochondrial apoptosis of BC cells.
A Representative immunofluorescence images of MitoSOX staining and quantification of MitoSOX mean fluorescence intensity (MFI) in RT-112 and UM-UC-3 cells stably transfected with shNC, shTTK#1, or shTTK#2 at 48 h. (Scale bar: 10 μm). B Mitochondrial H2O2 production in RT-112 and UM-UC-3 cells stably transfected with shNC, shTTK#1, or shTTK#2, assessed by MitoB oxidation. C Intracellular reactive oxygen species (ROS) levels in RT-112 and UM-UC-3 cells transfected with shNC, shTTK#1, or shTTK#2, were assessed by DCFH-DA staining and flow cytometry. D Mitochondrial membrane potential in RT-112 and UM-UC-3 cells transfected with shNC, shTTK#1, or shTTK#2 was assessed by JC-1 staining and flow cytometry. E Schematic diagram illustrating the proposed mechanism by which mitophagy inhibition leads to accumulation of damaged mitochondria, excessive mitochondrial ROS (mtROS) production, and induction of mitochondrial apoptosis. F Western blot analysis of apoptosis-related proteins in TTK knockdown RT-112 and UM-UC-3 cells, with additional evaluation after treatment with the mtROS scavenger Tempo. G Flow cytometry analysis of apoptosis in TTK knockdown RT-112 and UM-UC-3 cells, with additional evaluation after treatment with the mtROS scavenger Tempo. H Colony formation assays of TTK knockdown RT-112 and UM-UC-3 cells, with additional evaluation after treatment with the mtROS scavenger Tempo. Data are expressed as mean ± SD from three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001.
