Incomplete radiofrequency ablation promotes the development of CD133+ cancer stem cells in hepatocellular carcinoma cell line HepG2 via inducing SOX9 expression

Cancer stem cells (CSCs) accelerate the growth of hepatocellular carcinoma (HCC) residual after incomplete radiofrequency ablation (In-RFA). The present study aimed to detect the effects of In-RFA on stemness transcription factors (STFs) expression which are important for the production and function of CSCs, and to find which STFs promote HCC stemness after In-RFA. HepG2 cells were used for in vitro and in vivo studies. Flow cytometry and sphere-formation assays were used to detect the level and function of CD133+CSCs in the models. PCR array and ELISA were applied to analyze the altered expression of 84 STFs in CD133+CSCs in two models. Specific lentiviral shRNA was used to knockdown STFs expression, followed by detecting In-RFA's effects on the levels and function of CD133+CSCs. In-RFA was identified to induce CD133+CSCs and increase their tumorigenesis ability in vitro and in vivo. The mRNA levels of 84 STFs in CD133+CSCs were detected by PCR array, showing that 15 and 22 STFs were up-regulated in two models, respectively. Meanwhile, the mRNA levels of seven common STFs were up-regulated in both models. ELISA assay demonstrated that only the protein of sex determining region Y-box 9 (SOX9) was up-regulated in both models, the protein levels of the other 6 common STFs did not increase in both models. Finally, SOX9 was identified to play an important role in inducing, maintaining stemness and promoting tumorigenesis ability of CD133+CSCs in both models. In-RFA-induced SOX9 stimulates CD133+CSCs proliferation and increases their tumorigenesis ability, suggesting that SOX9 may be a good target for HCC treatment.