Analysis of radiosensitivity of cancer stem-like cells derived from canine cancer cell lines.

Cancer stem-like cells (CSCs) are self-renewing cells comprising a small subpopulation in tumours, and generate differentiated progeny through asymmetric division. It has been shown that CSCs are resistant to ionizing radiation, and this feature could be one of the mechanisms of tumour recurrence after radiation therapy. Much attention has been focused on to target CSCs; however, difficult of isolating CSCs and lack of knowledge on their radiosensitivity have limited this kind of research in veterinary medicine. In the present study, sphere-forming cells (SC), cultured using sphere formation method, were isolated from four type of canine tumour cell lines and evaluated if they have CSCs-like properties by expression of CSCs markers (real-time polymerase chain reaction) and capacity of tumorigenesis (xenograft transplantation in nude mice), and were assessed radiosensitivity (clonogenic survival assay) and DNA repair kinetics (immunofluorescence staining for p53-binding protein 1) after X-ray irradiation in comparison with the corresponding normal adherent culture cells (AC). All SCs were isolated using sphere formation and showed high gene expression of CD133 and tumorigenic ability as compared with AC. All SCs were significantly resistant against X-ray irradiation as compared with AC. In addition, the amount of DNA double-strand breaks after X-ray irradiation were significantly lower in SC compared with the corresponding AC. These results indicate that SC isolated through sphere formation possess CSCs-like characteristics and CSCs are important factor that affect radiosensitivity in canine tumours. In addition, radioresistance of CSCs may depend on reaction of DNA double-strand break after X-ray exposure.