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. 2016 Oct 20;23(12):846–856. doi: 10.1038/gt.2016.61

Figure 6.

Figure 6

(a, b) Reversal of genome instability by FXN gene transfer. (a) Analysis of genome instability via Imagestream flow cytometry capture of phosphorylated γ-H2AX recruited to DNA DSB seen as foci in cell nuclei. Mean number of γ-H2AX foci per cell measured by ImageStream. In both human and mouse primary fibroblasts means were calculated over three experiments for normal and five experiments for FRDA and LV FXN-treated. P-values were calculated using the single values of each cell of each experiment (n=28 000 for Human normal GMO7492; n=40 464 for Human FRDA GMO3665, and n=39664 for LV FXN-treated FRDA), (n=15000 for mouse normal Y47; n=19 000 for mouse FRDA YG8sR and n=23 000 for LV FXN-treated FRDA). In human cell lines, FRDA fibroblasts with the mean of 4 showed higher numbers of γ-H2AX foci per cell compared with normal cells with 2.1 and LV FXN-treated cells with 1.99 foci per cell. In mouse cell lines, FRDA fibroblast with the mean of 4.5 showed higher numbers of γ-H2AX foci per cell compared with Normal cells with 2.8 and LV FXN-treated cells with 2.7 foci per cell. Mean calculated from two independent experiments performed in duplicate. Error bars show the s.e.m. Significance levels are represented by asterisks: *P<0.05, **P<0.01 and ***P<0.001. (b) Mean number of γ-H2AX foci per cell measured by immunocytochemistry in human primary fibroblasts, normal fibroblasts (GMO7492, n=400), FRDA fibroblasts (GMO3665, n=400) and FXN-treated FRDA fibroblasts (GMO3665 n=400) and mouse primary fibroblasts, normal fibroblasts (Y47, n=400), FRDA fibroblasts (YG8sR, n=400) and FXN-treated FRDA fibroblasts (YG-FXN n=400). In human cell lines, FRDA fibroblast with the mean of 0.9 showed higher numbers of γ-H2AX foci per cell compared with Normal cells with 0.3 and FXN-treated cells with 0.46 foci per cell. In mouse cells, FRDA fibroblast with the mean of 1.9 showed higher numbers of γ-H2AX foci per cell compared with Normal cells with 0.56 and FXN-treated cells with 0.8 foci per cell. Mean calculated from two independent experiments performed in duplicate. Error bars show the s.e.m. Significance levels are represented by asterisks: *P<0.05, **P<0.01 and ***P<0.001.