Fig. 2: McPGK1 is required for liver TIC self-renewal.

A Quantitative real-time PCR to confirm the knockdown efficiency in mcPGK1 silenced cells. B FACS detection of mcPGK1 silenced cells for CD133 expression. C Quantitative real-time PCR analysis of TIC markers and TIC-associated TFs in mcPGK1 silenced and control cells. D Sphere formation of mcPGK1 silenced and control cells. Typical sphere images were in the left panel and sphere diameters of n = 30 spheres from three independent experiments were measured in the right panel. D3, day 3. Scale bars, 100 μm. E Sphere formation of mcPGK1 silenced and control HCC #2 sample cells. Scale bars, 500 μm. F McPGK1 fluorescence in situ hybridization and Ki67 staining in mcPGK1 silenced and control Huh7 cells. Typical images and Ki67⁺ ratios were shown. For each group, n = 10 fields were observed. Scale bars, 10 μm. G, H Tumor initiation assay of gradient numbers of mcPGK1 silenced and control cells derived from spheres derived from primary #2 cells. mcPGK1 detection for knockdown efficiency was shown in G. n = 7 6-week-old male BALB/c nude mice were used for H. TIC ratios and P-value were calculated by extreme limiting dilution analysis (ELDA) (http://bioinf.wehi.edu.au/software/elda/). Scale bars, 30 μm. I 1 × 10⁶ mcPGK1 knockdown and control luciferase labeled primary #2 cells were used for in vivo propagation. Typical liver images and calculated results were shown. n = 6 mice per group. J Sphere formation with 1000 CD133⁺ control and shmcPGK1 Huh7 cells, which were sorted via FACS assay. K Sphere formation with mcPGK1 knockdown and control cells, for which shmcPGK1 and control plasmids were delivered into primary HCC cells derived from sample #2 with mitochondria-targeting nanoparticles (Mito-NP). Scale bars, 500 μm. For A, B, C, E, J, K, n = 4 independent experiments. In all panels, data are shown as mean + s.d. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001, by two-tailed Student’s T-test. Source data are provided as a Source Data file.