Effect of PARPs inhibitors or of NAD⁺ supplementation on hepatic cells after H₂O₂ treatment.

<p>(A) The viability of WRL68 cells (assessed with the MTT assay) was analysed after 16 h of pre-incubation, exposure to H₂O₂ for 30 min and 24 h of post-incubation, always in the presence of PARPs inhibitors. Cell viability was expressed as the percentage of viable cells with respect to those determined in the control (non-treated cells), which was considered to be 100%. Results from the PJ34 and AG14361 inhibitors are shown on bar graphs (left for PJ34, showing the mean ± SEM of five independent experiments; right for AG14361, showing the mean ± SEM of three independent experiments). Significant differences: **<i>P</i> < 0.01 and ***<i>P</i> < 0.001 with respect to the control (non-treated cells) and ^##<i>P</i> < 0.01 with respect to 3.5 mM H₂O₂. (B) Bar graphs showing the percentage of cells containing ROS. The presence of ROS was measured by staining cells with H₂DCFDA. WRL68 cells pre-incubated for 16 h with PARPs inhibitors were treated with H₂O₂ for 15 min and the proportion of cells positively marked with DCF fluorescence was recorded by flow cytometry. Bar graphs show the mean ± SEM of three independent experiments. Significant differences: ***<i>P</i> < 0.001 with respect to the control (non-treated cells). (C) Percentage of viable WRL68 cells treated with H₂O₂ for 30 min and 24 h of post-incubation, always in the presence of 250 μM of NAD⁺. Cell viability was determined by the MTT method and expressed as the percentage of viable cells with respect to those determined in the controls (non-treated cells), which was considered to be 100%. Bars represent the mean ± SEM of four independent experiments. Significant differences: ***<i>P</i> < 0.001 with respect to the control (non-treated cells) and ^##<i>P</i> < 0.01 with respect to 3.5 mM H₂O₂.