Fig. 6

Effect of TDZ on NF-kB /CREB activation and on the autophagic pathway. a, b Human astrocytes were treated with medium alone or TDZ (1 μM) for 24 or 72 h, and then with LPS-TNF-α for an additional 24 h. NF-kB p65 protein levels were evaluated in cytoplasm and nuclei by western blot analysis. GAPDH and H3 were the loading controls. a Representative western blots. b Densitometric analysis of the immunoreactive bands was performed using ImageJ. The data are expressed as the percentage of optical density of the immunoreactive band relative to that of the control, which was set at 100 % and are the mean values ± SEM of three different experiments. c Human astrocytes were treated with medium alone, or TDZ (1 μM) or FLUOX (10 μM) for 24 h or 72 h, and then with LPS-TNF-α for an additional 24 h. At the end of treatment, CREB activation was determined by ELISA, as described in the Methods section. The data are expressed as percentages relative to untreated cells (control), which were set at 100 %, and represent the mean ± SEM of two independent experiments, each performed in duplicate. d, e Human astrocytes were treated with medium alone, TDZ or FLUOX or Rapamycin (RAPA) for 24 or 72 h. Following incubation, the protein levels of LC3B were evaluated by western blot analysis. GAPDH was the loading control. d Representative western blots. e Densitometric analysis of the immunoreactive bands. The data are expressed as the percentage of optical density of the immunoreactive band relative to that of the control, which was set at 100 %, and are the mean values ± SEM of three different experiments. Statistical significance was determined using a one-way ANOVA-Tukey HSD post hoc test: **P < 0.01, ***P < 0.001 vs. control; ^## P < 0.01, ^### P < 0.001 vs. cells treated with LPS-TNF-α