Fig. 5

Exosomes internalization blockage inhibited target cells transformation. a Exosomes were isolated and labeled with PKH-26. Cells were treated or not with Cytostatin (1.4 μg/ml), Heparin (10 μg/ml) and the anti-β4 integrin antibody (ASC-8; 10 μg/ml). In parallel, exosomes were treated or not with RGD (300 nM) and Collagenase I (500 μg/ml). Cells were exposed to exosomes and analyzed by flow cytometry after gating on cells (G1 population). Data are expressed as the percentage of PKH-26 positive cells. Values in brackets are the mean fluorescence intensity (MFI). Note that antagonists treatments reduced exosomes internalization. b Viability of cells treated as in (a). Note that treatments slightly affected cell viability. Values are mean +/− SD, (n = 3 independent cell cultures). c NanoSight analyses of exosomes treated or not with collagenase I. Note that exosome sizes are identical. (D-F) BRCA1-KO fibroblasts and exosomes were treated as in (a). Cells were washed and mixed with treated exosomes. This treatment was repeated every second day for 2 weeks. Antagonists untreated cells exposed to untreated exosomes served as control. Both cell population were transplanted into NOD/SCID mice. d 4 weeks after injection, mice injected with control cells (i) and blocked cells (ii) were photographed and euthanized. Representative pictures of tumors are shown. e Tumor volumes at euthanasia. Values are mean +/− SD, (n = 2 mice per group), P < 0.05. f Formalin-fixed paraffin-embedded xenotransplant samples were processed for H&E staining. Note that tumors obtained with treated cells displayed areas of necrosis. Scale bars, 50 μm