Skip to main content
. 2018 Jul 31;19(8):2233. doi: 10.3390/ijms19082233

Figure 4.

Figure 4

The genetic or pharmacologic inhibition of Akt1-phosphorylation affects DNA repair upon IR. TrC1 stably overexpressing Akt1-WT, pre-treated for 2 h with 0 or 4 µM MK-2206, or the phosphorylation-deficient Akt1-TA, -SA or -TASA mutants were exposed to irradiation with 0 Gy, 3 Gy (A,B) or 40 Gy (C,D) as indicated. (A,B) Cells were fixed in 3% para-formaldehyde (PFA), permeabilized with 0.2% Triton X-100 in phosphate-buffered saline (PBS) at distinct time points between 0 h and 24 h upon irradiation with 3 Gy, and stained with Hoechst33342, to visualize the nuclei (blue), and γH2A.X (magenta), to visualize sites of DNA DSB. (A) The number of γH2A.X foci at 2–24 h after irradiation with 3 Gy using the Focinator v 2.2. software [20] was normalized to the number of foci detected at 0.5 h time point. (C,D) Cells were processed by applying neutral comet assay to quantify the amount of damaged DNA in the form of DSB at a fixed time-point. The quantification was performed by the OpenComet software and depicts the comet tail length of each indicated Akt1 mutant 4 h upon 40 Gy. (A,C) Data show means ± SD from 3 independent experiments with 50 analyzed nuclei per trial and condition. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ANOVA test with Tukey correction. (B,D) Data show representative pictures out of 3 independent experiments.