Fig. 7From: Autophagy modulators influence the content of important signalling molecules in PS-positive extracellular vesiclesPS-EVs influence the autophagy flux, expression of senescence-associated proteins, and metabolic profiles of fibroblasts. (a) Autophagic flux in HGF cells after 12 h-lasting effect of CM2 treatment traced with an mRFP-GFP-LC3 tandem construct. Autophagosomes and autolysosomes are labelled in yellow and red, respectively. LysoTracker-based fluorescent signal is specific for lysosomal structures and acidic organelles. blank = exofree medium incubated at 37 °C 24 h. control = CM2 medium conditioned by non-treated FaDu cells. Scale bar equals 50 µm. Representative fields of view of minimally 11 per treatment. (b) CPD18-CM2 medium from FaDu cells pre-treated by CPD18 enhanced autophagy flux in HGF. The intensities of the yellow signal (calculated as meanRFP-times-GFP) and the meanRFP signal from the mRFP-GFP-LC3 tandem construct were significantly enhanced. noCM = HGF without conditioned medium; control = CM2 medium conditioned by non-treated FaDu cells. N indicates number of cells. (c) The expression level of proteins involved in the autophagy machinery or senescence after treatment (12 h; 48 h) with CM2 containing PS-EVs. Uncropped western blots for this figure are shown in Additional file 3. (d) and (e) ATP production from glycolysis and OXPHOS based on Seahorse parameters OCR (oxygen consumption rate) and ECAR (extracellular acidification rate) obtained from XF Real-Time ATP Rate Assay before and after autophagy inhibitors treatment; PER = Proton efflux rate, rate of protons extruded into the extracellular medium during glycolysis. blank = exofree medium incubated at 37 °C 24 h. FaDu CM = CM2 medium conditioned by non-treated FaDu cells. OM = oligomycin; ROT = rotenone; AA = antimycin A. P-values from group comparisons based on the t-test are shown. Asterisks represent statistical significance (* p < 0.05; ** p < 0.01; *** p < 0.001). Each dot represents a measurement in well. N indicates number of cellsBack to article page