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. 2018 Dec 12;8:17780. doi: 10.1038/s41598-018-36036-5

Figure 3.

Figure 3

Characterisation of multinucleated senescent arrest and escape. (a) Representative fluorescence images of HCT-116 cells untreated (CTR) or treated with 5 nM CNF1 for 72 h (CNF1), or polyploid HCT-116 cell-derived daughter cells (Dau), stained with phalloidin and DAPI. White arrowheads indicate filopodium-like structure at the cell periphery; Red arrowheads indicate micronuclei in the multinucleated cells. Bars, 50 μm. (b) Representative confocal images of HCT-116 cells untreated (CTR) or treated with 5 nM CNF1 for 72 h (CNF1). White arrowheads indicate filopodium-like structure at the cell periphery. Bars, 10 μm. (c) Percentage of cells containing multinuclei or micronuclei in HCT-116 cells untreated or treated with 5 nM CNF1 for 72 h, or polyploid HCT-116 cell-derived daughter cells. Data are mean ± SD of three independent experiments. **p < 0.01. (d) Representative images of SA-β-gal staining of HCT-116 cells (upper panel) and percentage of SA-β-gal positive cells (lower panel). Bars, 50 μm. Data are mean ± SD of three independent experiments. **p < 0.01. (e) Representative immunoblot analysis of p53, p21, p16, pRb, HMGA2, p-H3, H3 and β-actin in HCT-116 cells untreated or treated with 5 nM CNF1 at 24 h, 48 h and 72 h, or polyploid HCT-116 cell-derived daughter cells.