Changes in the Relative Number of SP Cells of Melanoma Line B16 after Radiation Exposure in vivo

The quantitative regularities of changes of cancer stem cell (CSC) population were explored after local γ-irradiation of experimental tumors (murine melanoma line B16). CSCs were detected by the ability of these cells to exclude Hoechst 33342 fluorescent dye and to form a so-called side population (SP) under flow cytometry study. In the control group of unexposed mice a positive correlation was found between the proportion of CSCs (SP) and tumorweight at the initial stage of growth (R = 0.77, p = 0.009). In the advanced stages of tumor growth similar relationship was not revealed. Statistically significant increase in the proportion of CSCs (SP) occurred 2-5 days after exposure of tumor to a dose of 10 Gy as compared to control; this index returned to the control level 8 days after irradiation. On the second day after exposure to radiation a linear correlation between the percentage of CSCs and a radiation dose in the range of 2-10 Gy was established (R = 0.98, p = 0.003), confirming a higher radioresistance of this population as compared to other cells not only in vitro (as it was previously shown by us and other authors), but also in vivo. These results suggest the possibility of application of this model system to assess the CSC sensitivity to various antitumor agents in vivo, including preclinical trials, and clarify the details of the practical application of this method.