Fig. 5: hMSC-derived let-7f is incorporated by 4T1 tumor cells impairing proliferation and invasion.

A qRT-PCR analysis of absolute let-7f expression in 4T1 mammary carcinoma cells after 24 h of cultivation under serum-free conditions without (control) and upon addition of 24 h-cell culture supernatants obtained from hMSCs that had been transfected with let-7f mimics or control oligonucleotides grown in the absence (−) and presence (+) of an inhibitor of exosome formation, GW4869. B hMSCs were transfected with Cy5-labeled let-7f and analyzed by fluorescence microscopy after nuclear staining with DAPI. Scale bars 5 µm. After 48 h of cultivation under serum-free conditions, hMSC culture supernatants were collected and added to subconfluent 4T1 cells. After 24 h of incubation, 4T1 cells were analyzed for Cy5-let-7f uptake (white arrows) adopting fluorescence microscopy. C Assessment of cell proliferation rates in 4T1 cells upon 72 h of incubation without (untreated) and with 24 h-cell culture supernatants obtained from hMSCs transfected with let-7f mimic or control oligonucleotides by use of the WST-8 assay. D, E 4T1 cells were transfected with let-7f mimics, let-7f inhibitor, or control oligonucleotides. D After 72 h of incubation, cell proliferation was determined by use of the WST-8 assay. E Cells were analyzed for their invasive potential using the Transwell assay with human ECM as a migration barrier and 10% human serum as chemoattractant. Results are given relative to control cells set as 100%. A, C, D Data represent the mean ± SD of triplicate measurements (n = 3); ***P < 0.001, **P < 0.01, *P < 0.05. B, E Results are mean values ± SD of a triplicate measurement and representative for three independent experiments (n = 3); ***P < 0.001, *P < 0.05.