Figure 2
From: Cannabidiol induces autophagy via ERK1/2 activation in neural cells
CBD induces autophagy in a concentration-dependent manner. (A) The autophagic flux was evaluated in SH-SY5Y cells treated with 5, 10 and 50 µM of CBD for 2 h in the presence or absence of the lysosome inhibitor NH4Cl (10 mM) added at the last hour of the treatment. The concentrations of 10 and 50 µM CBD increased LC3-II levels in the NH4Cl treated group; Two-way ANOVA (F3,52 = 3.499) followed by Sidak’s post-hoc test; *p < 0.05, **p < 0.01 and ***p < 0.001. (B) Time-course for 1, 5, and 10 µM of CBD on the autophagic flux evaluated at 1, 2 and 4 h in the presence or absence NH4Cl added at the last hour of the treatment. The concentration of 10 µM CBD increased LC3-II levels after 1, 2 and 4 h of treatment; Three-way ANOVA (F6,78 = 3.96) followed by Sidak’s post-hoc test, *p < 0.05 and ***p < 0.001, respectively vs. group treated with NH4Cl. Samples were subjected to western blotting using anti-LC3 and anti-α-tubulin primary antibodies. Representative images of LC3-II and a graphical summary reported as the means ± S.E.M of LC3-II levels after α-tubulin normalization are also displayed. (C) LC3 puncta count were analyzed in SH-SY5Y cells overexpressing Cherry-LC3 treated with CBD (10 µM) or under starvation conditions (STV) for 2 h. Representative fluorescent images are shown (scale bar, 10 µm). Data are expressed as mean ± S.E.M. One-way ANOVA (F2,22 = 16.148), followed by Dunnett´s post-hoc test. **p < 0.01 and ***p < 0.001 relative to CTR group. The entire blots are presented in Supplementary Figure S1.
