Fig. 4

Silvestrol treatment induces autophagy in MDA-MB-435 cells. a Cells were treated with either DMSO, 5 μM SAHA, 30 nM homoharringtonine (HHT), or 25 nM silvestrol for 24 h and subsequently stained with monodansylcadaverine (MDC) or acridine orange (AO). Both silvestrol and SAHA promoted vacuole formation (indicated by blue staining in MDC and red staining in AO). Images are representative of a pattern of staining observed in at least three independent experiments. b Silvestrol induces autophagy by mediating EGFP-LC3 translocation. Representative pictures of cells treated with DMSO, 5 μM SAHA or 25 nM silvestrol for 24 h. c Cells were treated with 25 nM silvestrol in the presence or absence of 3MA (10 mM), bafilomycin A1 (50 nM) and then harvested for protein analysis. Cell lysates were resolved in SDS-PAGE and probed with specific antibodies against LC3 and Actin. d Wild-type (WT) or Atg7^-/- MEFs were treated with 25 nM silvestrol for 72 h. Cytotoxicity was determined by MTS assay with the viability of control cells defined as 100 %. The data are represented as means ± SEM, * p ≤0.05