Fig. 3

Oli-neu differentiation and lipid homeostasis is impaired upon reduction of β-glucocerebrosidase activity. a Representative western blot of CNP and MAG protein levels in differentiated Oli-neu cells upon vehicle (veh) and CBE treatment. β-actin (ACTB) was used as loading control. b Densitometric quantification of western blots for CNP and MAG upon dbcAMP treatment (t3) in the presence of vehicle (veh) or CBE (*, p < 0.05; unpaired two-tailed Mann–Whitney U test; CNP, n = 4; MAG, n = 3 independent experiments). Error bars indicate s.e.m. c qRT-PCR for Plp1 (left panel) and Mag (right panel) gene expression in Oli-neu cells at t0 and t3, upon vehicle (veh) or CBE treatment (*, p < 0.05; **, p < 0.01; unpaired two-tailed Student’s t test; n = 9 samples from 3 independent experiments). Error bars indicate s.e.m. d, e Representative images of TUNEL test and quantification of TUNEL-positive nuclei normalized on the total number of cells at t0 and t3 differentiated Oli-neu treated with vehicle (veh) or CBE (Kruskal–Wallis test and Dunn's test for multiple comparison; n = 4 independent experiments). Scale bar = 100 μm. f, g Representative western blot (f) and densitometric quantification of cleaved caspase-3 (CL-CASP3) in t0 and t3 differentiated Oli-neu cells upon treatment with vehicle (veh) and CBE. β-actin (ACTB) was used as loading control (Kruskal–Wallis test and Dunn's test for multiple comparison; n = 3 independent experiments). Error bars indicate s.e.m. h Filipin III staining of differentiated Oli-neu cells treated with vehicle (veh) or CBE at t3. Scale bar = 50 μm. i High Performance Thin Layer Chromatography (HPTLC) analysis of glucosylceramide (GlcCer), lactosylceramide (LacCer), sphingomyelin (SM) and globotriaosylceramide (Gb3), reported as relative levels to total resolved lipids (unpaired two-tailed Student’s t-test; n = 4 independent experiments). Error bars indicate maximum and minimum values