Figure 3. CSC have an increased rate of DSB clearance after ionising radiation.

Cells were treated with 2Gy IR and immuno-stained to investigate the level of DNA damage and repair following treatment. (A) CA1 cells were fixed at set time-points (1 h, 18 h) following treatment. Double strand DNA breaks (DSB foci) were measured by immunofluorescence against anti- γH2A.x and anti-pCHK2. Cells were counterstained with DAPI for nuclear visualisation. DSB foci rapidly accumulated in CA1 cells within an hour after irradiation, regardless of the type of population that was exposed to IR. DNA repair, or DSB foci clearance, was assessed by measuring the remaining γ-H2A.X foci 18 hours post-IR. CD44^hi/ESA^hi and CD44^hi/ESA^low cells displayed a reduced amount of DNA double strand breaks, suggesting quicker and/or more efficient DNA repair. (B) DSB foci were counted in each population for both CA1 and Luc4 cells in ImageJ using the cell counter tool. Each data point represents the average amount of nuclear DSB foci (double-positive for γH2A.x and p-CHK2) per field. An average of approx. 10 cells was quantified per high power field. Data are representative of two separate experiments performed in two replicate wells each. *P < 0.05, **P < 0.01, ***P < 0.001.