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. 2017 May 29;3:17015. doi: 10.1038/cddiscovery.2017.15

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Copyright © 2017 The Author(s)

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Bystander uptake of fluorescently labeled cfCh and activation of DNA damage and inflammation following co-cultivation of NIH3T3 cells with irradiated and un-irradiated GalNAc-T2-GFP HeLa cells. (a) Upper panel, fluorescent microscopy at 36 h showing uptake by bystander NIH3T3 cells of fluorescent cfCh particles released from BrdU pre-labeled dead, but not live, GalNAc-T2-GFP HeLa cells; lower panel, quantitative analysis of mean fluorescence intensity (MFI) of images given in upper panel. (b–e) Upper panels, fluorescent microscopy at 36 h showing activation of H2AX (b), p-ATM (c), NFκB (d) and active Caspase-3 (e) in bystander NIH3T3 cells. Lower panels, quantitative analysis of MFI of images given in upper panel. One thousand nuclei were gated and analyzed for quantifying MFI in each case. *P = 0.05; ****P = 0.0001; NS=Not Significant. Results (mean±S.E.) were analyzed by Student’s t-test. (f) Fluorescent microscopy of NIH3T3 cells co-cultivated with irradiated B16-F10 melanoma cells at 6 h showing BrdU and γ-H2AX co-expressing cells (upper panel); BrdU and IL-6 co-expressing cells (middle panel), and γ-H2AX and IL-6 co-expressing cells (lower panel).

Bystander uptake of fluorescently labeled cfCh and activation of DNA damage and inflammation following co-cultivation of NIH3T3 cells with irradiated and un-irradiated GalNAc-T2-GFP HeLa cells. (a) Upper panel, fluorescent microscopy at 36 h showing uptake by bystander NIH3T3 cells of fluorescent cfCh particles released from BrdU pre-labeled dead, but not live, GalNAc-T2-GFP HeLa cells; lower panel, quantitative analysis of mean fluorescence intensity (MFI) of images given in upper panel. (b–e) Upper panels, fluorescent microscopy at 36 h showing activation of H2AX (b), p-ATM (c), NFκB (d) and active Caspase-3 (e) in bystander NIH3T3 cells. Lower panels, quantitative analysis of MFI of images given in upper panel. One thousand nuclei were gated and analyzed for quantifying MFI in each case. *P = 0.05; ****P = 0.0001; NS=Not Significant. Results (mean±S.E.) were analyzed by Student’s t-test. (f) Fluorescent microscopy of NIH3T3 cells co-cultivated with irradiated B16-F10 melanoma cells at 6 h showing BrdU and γ-H2AX co-expressing cells (upper panel); BrdU and IL-6 co-expressing cells (middle panel), and γ-H2AX and IL-6 co-expressing cells (lower panel).