Selective enrichment of hepatocellular cancer stem cells by chemotherapy

This study investigated the enrichment of cancer stem cells (CSCs) in hepatocellular carcinoma (HCC) by chemotherapy. SMMC-7721 cells were inoculated into mice treated with 0, 2, 5 or 10 mg/kg cyclophosphamide (CTX). Tissue from the resulting tumours was re-inoculated into CTX-treated mice two more times, thus producing three generations of tumour cells for each dose of CTX. Chromosome and fluorescence-activated cell sorting analyses were performed to determine the purity of the enriched cells. Sphere culture, colony formation and proliferation assays demonstrated that the self-renewal potential, proliferative activity and clonogenicity of the enriched cells in vitro increased with increasing chemotherapy dose and generation. The ability of the enriched cells to produce xenograft tumours in mice was also dependent on chemotherapy dose and generation. In conclusion, subjecting HCC cells to chemotherapy in vivo enriched the samples for HCC CSCs in a dose- and generation-dependent manner.