Fig. 2
Establishment of the iOVCAR-3-OSKM model, which exhibited stemness and cancer-initiating cell properties. A Schematic of the transduction procedure and establishment of iOCICs. The cells were infected with SeV containing four factors (OSKM) at day 0 and replated onto MEF dishes. Tra-1–60 positive colonies were picked up on day 21 and day 28. Scale bars: 200 µm. B Morphology of iOVCAR-3-OSKM colonies. Six-day 7-iOVCAR-3-OSKM clones on feeder cells. Scale bars: 200 µm. C Pluripotent gene expression and SeV silencing in iOVCAR-3-OSKM clones. Expression of endogenous pluripotent genes (OCT4, SOX2, KLF4, and NANOG), SeV, and the housekeeping gene GAPDH. cDNA of human ESCs (H9 cell line) and a human induced pluripotent cell line (iPBMCF) were used as positive controls. Neg Ctl: negative control (PCR mixture without cDNA). The samples derive from the same experiment and gels were processed in parallel. D qRT–PCR of the gene expression of four factors and markers previously reported to be related to OCICs in iOVCAR-3-OSKM cells. mRNA expression levels were normalized to those of GAPDH. Relative expression levels compared to those in parental OVCAR-3 cells are shown in log2 scale (n = 3). Error bars indicate SEM. *P < 0.05, **P < 0.005, Student's t test. E Sphere-formation ability in vitro. A total of 105 cells were plated on low attachment dishes and cultured for 14 days. The numbers of spheroids (≥ 25 µm) were counted under a microscope. Spheres numbers of iOVCAR-3-OSKM #48, #38 and SP were significantly increased compared #2 and parental OVCAR-3 cells. The error bars indicate the SD. ***P < 0.0005, Student’s t test. NS means nonsignificant. Scale bars: 200 µm. NS means nonsignificant. F Tumorigenicity of iOVCAR-3-OSKM cells after implantation in the subcutaneous regions of immunodeficient NOD-SCID mice. A total of 103 cells were subcutaneously injected into both flanks of immunodeficient female NOD-SCID mice (n = 8). Tumor volume and tumor formation rate (n = 16) are shown