Figure 6.

Effect of SIRT1 and SIRT2 modulation in differentiated SH-SY5Y cells. (a) SIRT1 activity was measured by a fluorometric assay on 40 μg of total protein extract from either uninfected (Ctrl) and cells infected with adenoviral vectors coding for Wt-SOD1 (Wt) and G93A-SOD1 (G93A) treated with one of the following: 15 μM AGK2, 3 μM Ex527 and 20 μM SIRTinol that inhibit SIRT2, SIRT1 and both Sirtuins, respectively; and 10 nM SRT1720 that activates SIRT1. Activity is reported as percent of the relative uninfected DMSO-treated control and reported as the mean±S.D. of three independent experiments with each sample in triplicate. Values significantly different from relative controls are indicated with *P<0.05 and **P<0.01 with respect to Ctrl; and ^#P<0.05 and ^##P<0.01 with respect to DMSO-treated cells. (b) SH-SY5Y cells either uninfected (Ctrl) or infected with adenoviral vectors coding for Wt-SOD1 (Wt) and G93A-SOD1 (G93A) were treated with 15 μM AGK2, 3 μM Ex527, 20 μM SIRTinol and 10 nM SRT1720. Cell viability was assessed by the MTS assay 48 h after infection and drug treatments. Absorbance at 490 nm are expressed as percent of the relative uninfected control cells and reported as the mean±S.D. of three independent experiments made in triplicate. Values significantly different from relative controls are indicated with *P<0.05 with respect to Ctrl and ^## P<0.01 with respect to DMSO-treated cells. (c) Cells either uninfected (Ctrl) or infected with adenoviral vectors coding for Wt-SOD1 (Wt) and G93A-SOD1 (G93A) were treated with three increasing doses of AGK2, Ex527, SIRTinol or SRT1720. Cell protein extracts were assayed, 48 h after infection and drug treatment, for Caspase-3 activity by a fluorescence enzymatic assay and reported as percent of the relative uninfected control cells. Mean±S.D. of four independent experiments is given. Values significantly different from the relative controls are indicated with *P<0.05 with respect to Ctrl and ^##P<0.01 with respect to DMSO-treated cells