Figure 5.

Loss of mitofilin results in mitochondrial functional abnormality. Mitofilin (red) versus control (black) siRNA-treated HeLa cells were stained with 40 nM DiOC₆ (A) or 2 μM 2-hydroethidium (B) and analyzed by flow cytometry to assess mitochondrial membrane potential and ROS production, respectively. Metabolic flux was measured by the oxidation of ³H-labeled palmitate to H₂O (C). For oxygen consumption, HeLa cells were suspended in medium A (250 mM sucrose, HEPES-KOH, pH 7.5, 1 mM ADP, and 2 mM K₂HPO₄) and introduced into a chamber equipped with a Clark electrode. Respiration was induced by the addition of 5 mM malate/glutamate. Rates of oxygen consumption were calculated from the slopes of state 3 respiration (D). All the assays were performed with the HeLa cells after 48 h of the second-round siRNA treatment. Filled and open bars represent mitofilin versus control siRNA-treated cells, respectively.