Fig. 3

Exosomal transfer of miR-196a from CAFs to HNC cells. a miR-196a expression in NFs, CAFs, normal oral epithelial cells (titled normal), primary cancer cells, and HNC cell lines was analyzed using real-time PCR. b CAL 27 and HN4 cells were incubated with control CM, CAF-CM, or exosome-depleted CAF-CM for 24 h. The miR-196a expression level in these cells was determined using real-time PCR. c CAL 27 and HN4 cells were co-cultured with DMSO-treated HNC cells, DMSO-treated CAFs, or GW4869-treated CAFs for 24 h. The miR-196a expression level was then detected in HNC cells using real-time PCR. d CAFs transiently transfected with Cy3-tagged miR-196a (Cy3-miR-196a) were co-cultured with CAL 27 or HN4 cells for 48 h. Fluorescence microscopy was used to detect the red fluorescent signals in HNC cells (scale bar, 20 μm). e Real-time PCR analysis of miR-196a expression in CAF-CM treated with RNase A (2 mg/mL) alone or combined with Triton X-100 (0.1%) for 20 min. f Real-time PCR analysis of miR-196a expression in exosomes, exosome-depleted CM, and whole CM derived from CAFs. g miR-196a expression in CAL 27 and HN4 cells was detected by real-time PCR at 24 h after incubation with exosomes (25 μg/mL) from HNC cells (Ctrl Exos), CAFs, or CAFs transfected with or without miR-196a mimics. h miR-196a expression in CAL 27 and HN4 cells was detected by real-time PCR at 24 h after incubation with exosomes (25 μg/mL) from HNC cells (Ctrl Exos), CAFs, and CAFs transfected with or without anti-miR-196a. i, j CAL 27 and HN4 cells were treated with the indicated exosomes (25 μg/mL) for 6 days, and the cisplatin response in these cells was determined with MTT assays. (ns, no significant difference; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001)