Fig. 1: Distinct effects of Tfam, Cox10 and Trx2 silencing on mitochondrial function.

HUVECs were transfected with control, Tfam, Cox10 or Trx2 siRNAs. 48 h after transfection, cells were subjected to various assays. a Cell lysates were analyzed by Western blotting for TFAM, COX10 and TRX2 depletion with respective antibodies. Protein levels were quantified and presented as fold changes by taking control siRNA as 1.0. n = 3 (three independent experiments). b Mitochondrial DNA contents were quantitated by qPCR and presented as fold changes compared to control siRNA, n = 6 (duplicates from three independent experiments). c Mitochondrial ROS (mtROS) were assessed by a mitochondrial-specific ROS probe mitoSOX and fluorescence intensity was presented as arbitrary fluorescence unit (AFU). n = 6. d ATP production was measured and presented as mmol/cell, n = 6 (duplicates from three independent experiments). e–i The oxygen consumption rate (OCR) measured by Seahorse. e A diagram for typical mitochondrial stress test profiles. The addition of the coupled respiration inhibitor oligomycin (1 μM) was used to assess ATP production and proton leak. Maximal respiration was measured by adding 1 μM carbonyl cyanide-4-(trifluoromethoxy) phenylhydrazone (FCCP), and spare respiratory capacity and non-mitochondrial respiration was measured by adding 0.5 μM rotenone and 0.5 μM antimycin A. f–h OCR in siRNA-transfected HUVECs. i The basal, the ATP-coupled and the maximum oxygen consumption rate were calculated. n = 6 (Duplicates from three independent experiments). Data are means ± SEM. P values are indicated, using one-way ANOVA followed by Tukey’s multiple comparisons test. ns: non-significance (P > 0.05). Source data are provided as a Source Data file.