Fig. 4.

FDX1 regulates metabolism and its downregulation facilitates mitophagy of HCC cells. (A) Non-targeted metabolomics analysis of metabolites in indicated HCC cells. The volcano diagram shows the metabolites that were significantly altered after overexpression or knockdown of FDX1 (top panel). Venn diagram shows the common altered metabolites in HCC cells after overexpression or knockdown of FDX1 (lower left panel). Heatmap shows these common altered metabolites in positive ion mode and negative ion mode (lower right panel). (B) KEGG enrichment analysis for metabolism by differential metabolites from non-targeted metabolomics analysis. (C) Representative images of MHCC-97H^FDX1, PLC/PRF/5^shFDX1 and their control cells after transfecting with mRFP-GFP-LC3 lentivirus. Hoechst 33342 was used for nuclear staining. Scale bars, 5 μm. (D) Mitochondria and lysosomes of MHCC-97H^FDX1, PLC/PRF/5^shFDX1 and their control cells were stained by MitoTracker-Red and LysoTracker-Green, respectively. Hoechst 33342 was used for nuclear staining. Scale bars, 5 μm. (E) Representative transmission electron microscopy images of MHCC-97H^FDX1, PLC/PRF/5^shFDX1 and their control cells. Scale bars, 1 μm. (F) Western blot analysis of PINK1, Parkin, Tom20, p62, and LC3 I/II in HCC cells with FDX1 overexpression or knockdown. (G) Protein expression of FDX1, Parkin and LC3 was examined by IHC in subcutaneous tumors derived from the indicated cells. Scale bars, 100 μm. Data presented as mean ± SD of triplicate independent experiments. **P < 0.01; ***P < 0.001. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)