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. 2016 Oct 24;6:36022. doi: 10.1038/srep36022

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Copyright © 2016, The Author(s)

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(A,B) Clonogenic survival of EM9 (XRCC1^−/−) and AA8 (XRCC1^+/+) CHO cells (A) or PARP1^+/+ (wt) and PARP1^−/− MEFs (B) after a treatment with CDT^wt or CDT^H153A. Results represent the mean ± SD of at least 3 independent experiments. Statistics were calculated by unpaired Student’s t-test (**P < 0.01; ***P < 0.001). Graphs in linear scales are presented in Fig. S9. (C) Alkaline COMET assays on HCT116 control and shXRCC1 cells non-treated (NT) or treated for the indicated times with 25 ng/ml of CDT^wt or CDT^H153A. Left panel: Tail DNA percentage. Data are the mean ± SD of at least 3 independent experiments. Statistics were calculated by unpaired Student’s t-test (*P < 0.05; **P < 0.01; ***P < 0.001). Right panel: representative images of non-treated cells (NT) or cells treated with 25 ng/ml of CDT^wt for 24 h and evaluated with alkaline COMET assays. (D) Representative images of 53BP1 immunostaining in AA8 (XRCC1^+/+) and EM9 (XRCC1^−/−) non-treated (NT) CHO cells, treated with 75 pg/ml of CDT^wt for 24 h or for 2 h followed by a 22 h recovery time (repair 22 h). Scale bar = 20 μm. (E) Quantification of AA8 (XRCC1^+/+) and EM9 (XRCC1^−/−) CHO cells positive for 53BP1 foci formation after CDT^wt or CDT^H153A exposure at the indicated doses for 24 h or for 2 h followed by a 22 h recovery time (2 h). NT: non-treated cells. Data are expressed as the mean ± SD of at least 3 independent experiments. Statistics were calculated by unpaired Student’s t-test (*P < 0.05; ***P < 0.001). (F) Cell cycle analysis by flow-cytometry of HCT116 control and shXRCC1 cells non-treated (NT) or exposed for 24 h to CDT^wt (0.75 ng/ml). Upper panel: Graphs show the cell cycle profiles obtained for one representative experiment. Lower panel: Cell cycle index. Data represent the mean ± SD of at least three independent experiments. Statistics were calculated by unpaired Student’s t-test (*P < 0.05; ***P < 0.001).