PFKM inhibits doxorubicin-induced cardiotoxicity by enhancing oxidative phosphorylation and glycolysis

Figures

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  Figure 1. DOX treatment inhibited PFKM expression in H9c2 cells. (A–C) PFKM expression in DOX-treated H9c2 cells. (D–F) PFKM expression in DOX-treated H9c2 cells for different time points. *P < 0.05, ***P < 0.001 vs 0 μM or 0 h. (see Supplementary Fig. S1–S4)

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  Figure 2. PFKM upregulation inhibited DOX-induced cell apoptosis and DOX-decreased OXPHOS and glycolysis. PFKM-overexpressing H9c2 cells were treated with DOX for 24 h, and (A) cell viability, (B, C) TUNEL staining, (D, E) expression of PFKM, Bcl-2 and Bax, (F) OCR, (G) ECAR, (H) ATP and (I) lactate level were measured. Scale bar: 100 μm. ***P < 0.001 vs Control; ##P < 0.01, ###P < 0.001 vs DOX + vector. (see Supplementary Fig. S5–S8)

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  Figure 3. PFKM downregulation promoted cell apoptosis and inhibited OXPHOS and glycolysis. H9c2 cells were transfected with PFKM siRNA or siNC, and (A) cell viability, (B, C) TUNEL staining, (D, E) expression of PFKM, Bcl-2 and Bax, (F) OCR, (G) ECAR, (H) ATP and (I) lactate level were measured. Scale bar: 100 μm. ***P < 0.001 vs siNC. (see Supplementary Fig. S9–S12, more information for Supplementary Fig. S12 can be found in Supplementary Information 1.)

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  Figure 4. PFKM regulates DOX-mediated cell growth and apoptosis via the glycolysis pathway. PFKM-overexpressing H9c2 cells were treated with DOX with/without 2-DG for 24 h, and (A) cell viability and (B, C) TUNEL staining was measured. Scale bar: 100 μm. ***P < 0.001 vs Control; ##P < 0.01, ###P < 0.001 vs DOX + vector + vehicle; ΔΔP < 0.01, ΔΔΔP < 0.001 vs DOX + PFKM + vehicle.

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  Figure 5. HDAC1 inhibited H3K27ac-induced transcription of PFKM in DOX-induced H9c2 cells. (A, B) H3K27ac and HDAC1 levels in DOX-treated H9c2 cells. (C) ChIP assay of H3K27ac on the PFKM promoter in DOX-treated H9c2 cells. (D) ChIP assay of H3K27ac on the PFKM promoter and (E–G) expression of PFKM and H3K27ac in H9c2 cells treated with 2 μM DOX and 0.1 μM MGCD for different time points. *P < 0.05, **P < 0.01, ***P < 0.001 vs 0 h or control. (see Supplementary Fig. S13–S18)

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  Figure 6. HDAC1-induced PFKM transcriptional repression regulated DOX-mediated OXPHOS and glycolysis in H9c2 cells. H9c2 cells were transfected with PFKM siRNA or nonspecific siRNA (siNC) and treated with 2 μM DOX in the absence or presence of 0.1 μM MGCD for 24 h, and (A) cell viability, (B) OCR and (C) ECAR, (D) ATP and (E) lactate level were measured. (F) Schematic representation of the regulation of DOX-induced cardiotoxicity via OXPHOS and glycolysis through HDAC1-induced PFKM transcriptional repression. ***P < 0.001 vs Control; ###P < 0.001 vs DOX + siNC + vehicle; ΔΔP < 0.01 vs DOX + siPFKM-1 + vehicle.