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. 2015 Feb 28;6(8):5978–5989. doi: 10.18632/oncotarget.3468

Figure 6. Perturbation of thiamine metabolism results in persistent DNA-damage and reduced nucleotide pools.

Figure 6

A) HeLa cells were irradiated following transfection with 40 nM Ambion siRNA (siNT, siTPK1 and siTHTR1), or treatment with either DMSO (control) or 10 μM PyrH. Cells were fixed 24 hrs post-irradiation and probed for γ-H2AX (red), 53BP1 (green) and nuclei (blue). Representative images of irradiated cells treated and stained as indicated are shown. B) Quantification of DNA damage foci in HeLa cells irradiated after siRNA transfection as indicated, counting a minimum of 500 cells per well. Representative experiment of n=3 is shown, data represented as mean +/− SD from triplicate wells. Unpaired two-sided students t-tests comparing siNT to candidate knockdown **p<0.01, ***p<0.001. C) Quantification of DNA damage foci in HeLa cells irradiated after 4 days incubation with DMSO (control) or 10 μM PyrH. Cells stained and quantified as described in B. D) High performance liquid chromatography of HeLa cells incubated with or without PyrH for 4 days. Representative of n=3.