Fig. 8

Spautin-1 abrogates induction of BECLIN-1-dependent autophagy in Human Gingival Mesenchymal Stem Cells cultivated in osteoblastic differentiation condition. HGMSCs were cultured for 21 days in control medium or in differentiation medium supplemented with 1 μM resveratrol (Diff + RV) in the absence or in the presence of spautin-1 (Sp1). At the end, cell homogenates were processed for western blotting analysis of the expression of a BECLIN-1 (target of spautin-1) and of b LC3 (marker of autophagosome). Densitometry (arbitrary units) of the specific bands is included. Data were reproduced in three independent experiments. The ratio LC3-II/LC3-I is assumed as an index of autophagosome and autolysosome accumulation in the cell. c HGMSCs were plated on sterile coverslips, let adhere and cultured for 21 days in control medium or in differentiation medium supplemented with 1 μM resveratrol (Diff + RV) in the absence or the presence of spautin-1 (Sp1). At the end, the coverslips were fixed and processed for immunofluorescence staining of the autophagy interactome markers Vps34 (PI3KC3) and BECLIN-1. Fluorescence staining was quantified with the ImageJ software. Integrated fluorescence intensity of co-labeled area (yellow) was calculated and reported in histograms. Data from three coverslips per condition reproduced in three separate experiments