In this study, for the first time, we found that Tregs enhanced the ‘stemness’ of HCC cells, demonstrated by increased TICs ratio, upregulated expression of TICs-related genes CD133, Oct3/4, Sox2, c-Myc, Klf4, Nanog, CD13, EpCAM, elevated tumor sphere formation, and tumorigenic ability. Here, NANOG is linked to hepatocellular carcinoma.