Fig. 4: miR-218-5p impairs DXR-induced mitophagy.

A Total lysate from MCF7 cells transfected with GFP or GFP/miR-218-5p vectors, and then treated with DXR 30 µM for 24 h, were immunoblotted with the indicated antibodies. The graphs show the mitochondrial markers COXII and COXIV protein levels normalised on Vinculin, which has been used as a loading control. Solid dots represent the value respect to the control condition, from 3 independent experiments. B Representative immunofluorescence images of the mitochondrial network, visualised through TOMM20 staining, in the indicated conditions. Histograms represent the mean of the cellular area occupied by mitochondria, expressed as %, for each cell analysed, respect to the control condition. At least, 25 cells from 3 independent experiments were analysed. Scale bar, 10 μm is shown. C mtDNA/nDNA ratio from MCF7 grown as mammospheres, expressing GFP or GFP/miR-218-5p, and treated with DXR 2 µM for 72 h. Dots represent the mean of Ct of at least 3 technical replicates, from 5 independent experiments. D Total lysate from MDA-MB-231 cells transfected with GFP or GFP/miR-218-5p and treated with DXR 30 µM for 24 h, were immunoblotted with the indicated antibodies. The graphs show the mitochondrial markers COXII and COXIV protein levels normalised on β-actin that has been used as a loading control. Solid dots represent the value respect the control condition, from 3 independent experiments. SD are shown on the graph in red. One-way ANOVA (Tukey’s multiple comparison) was performed for A, B and D. Unpaired t-test (Welch’s correction) was performed for C. *p < 0.05, **p < 0.01, ***p < 0.001.