<p>(A–D) MCF7 cells were left untreated, treated with Oligofectamine (Oligo) or transfected with control siRNA (CT) or indicated siRNA pools. (A) After 60 h, cells with enlarged acidic compartment (late endosomes and lysosomes) were identified by flow cytometry (FL-2A) of LysoTracker Red-stained cells. The threshold for high intensity staining was defined so that 90% of control siRNA-transfected cells were below. (B) After 72 h, total cysteine cathepsin activity (zFR-AFC cleavage) was determined. HSPA1 and CTSB siRNAs served as internal controls. (C, D) After 60 h, cells were stained for Lamp-2 (C) or F-actin (D) and analyzed by confocal microscopy. Representative images are shown. <i>Arrows</i>, aggregation of lysosomal structures in cell protrusions/periphery. <i>Bars</i>, 20 µm. Values represent means + SD of a minimum of three independent experiments. *<i>P</i><0.05, **<i>P</i><0.01, ***<i>P</i><0.001, vs. control siRNA-transfected cells.</p
