Figure 3

Resveratrol increases PP2A activity and dephosphorylates Tau at PP2A-sensitive sites in primary cortical neurons. Representative western blots and quantifications of several independent experiments are shown in a–g. Band intensities of phospho-Tau were normalized to total Tau (Tau-5). For each experiment the respective control sample was set to 100%. (a) Primary cortical neurons of wild-type mice were treated with increasing concentrations of resveratrol for 20 hours. Cell lysates were analysed on western blots using antibodies detecting Tau phosphorylation at S202, total Tau (Tau-5), and actin. n = 3, *p < 0.0001 (b) Primary cortical neurons from wild-type mice were treated with 100 µM resveratrol over increasing time intervals. Cell lysates were analysed on western blots using antibodies detecting Tau phosphorylation at S202, total Tau (Tau-5), and GAPDH. n = 6, *p < 0.05. (c) Primary cortical neurons of wild-type mice were treated with increasing concentrations of resveratrol for 20 hours. Cell lysates were analysed on western blots using antibodies detecting Tau phosphorylation at S396, total Tau (Tau-5), and actin or GAPDH as loading controls. n = 3. (d) Primary cortical neurons of wild-type mice were treated with 100 µM resveratrol over increasing time intervals (lower panel). Cell lysates were analysed on western blots using antibodies detecting Tau phosphorylation at S396, total Tau (Tau-5), and actin or GAPDH as loading controls. n = 3. (e) Primary cortical neurons from wild-type mice were treated with increasing concentrations of resveratrol for 20 hours. Cell lysates were analysed on western blots using antibodies detecting phospho-S6K (n = 3, *p < 0.05) and phospho-S6 (n = 5, *p < 0.05), total S6K as well as total S6, and actin. Band intensities of phospho S6/S6K were normalized to total S6/S6K. For each experiment the respective control sample was set to 100%. (f) Primary cortical neurons from wild-type mice were treated with 100 µM resveratrol over increasing time intervals. Cell lysates were analysed on western blots using antibodies detecting the PP2A targets p-S6K (n = 4, *p < 0.05) and p-S6 (n = 3, *p < 0.05), total S6K as well as total S6, and actin. (g) Primary cortical neurons from wild-type mice were treated with 100 µM resveratrol and/or the PP2A inhibitor okadaic acid (10 nM). Cell lysates were analysed on western blots using antibodies detecting Tau phosphorylation at S202, total Tau (Tau-5), phospho-S6, total S6 and actin (n = 4, *p < 0.05). (h) Cell viability is not affected by resveratrol. Primary neurons were treated with increasing concentrations of resveratrol for 20 hours and cell viability was measured in a WST-1 assay. Columns represent mean values +/− SEM (n = 6).