Fig. 7
From: FoxM1 repression during human aging leads to mitotic decline and aneuploidy-driven full senescence
FoxM1 governs aneuploidization-driven cellular senescence in elderly cells. a–d FACS sorting of senescent cells from neonatal a, elderly b, FoxM1 siRNA-depleted neonatal c, and 84 y/87 y with FoxM1-dNdK d cell populations with high β-galactosidase activity. The gates were defined accordingly to the respective auto-fluorescent control. e Relative intensity levels of the fluorogenic substrate DDAOG in the sorted cell populations. f Aneuploidy index in FACS-sorted β-gal-positive fibroblast subpopulations (β-gal +) vs. unsorted populations (∅) as determined by FISH analysis for three chromosome pairs. g Experimental layout of live-cell/fixed-cell correlative microscopy analysis shown in h, i. Mitotic elderly fibroblasts expressing H2B–GFP and respective daughter cells were imaged for 72 h. SA-β-Gal assay and immunostaining for 53BP1/p21 were performed at the end of imaging. h Movie frames of representative phenotypes observed for elderly cells expressing H2B–GFP. Top panel, correct chromosome segregation, with cycling daughter cells (Supplementary Movie 9). Middle panel, correct chromosome segregation with non-cycling daughter cells staining negative for SA-β-Gal and 53BP1/p21 (Supplementary Movie 10). Bottom panel, chromosome mis-segregation leading to micronuclei formation (arrowheads), with non-cycling daughter cells staining positive for senescence markers (SA-β-Gal and 53BP1/p21) (Supplementary Movie 11). Dashed line indicates the tracked daughter cell. Time, hour:minute. Scale bar, 30 µm (movie frames) and 15 µm (immunostaining). i Single-cell analysis of daughter cell fate (cell death, cell cycle arrest, and cell senescence) from mitoses with apparent correct chromosome segregation or with mis-segregation (leading to micronuclei formation). Non-cycling daughter cells were stained for senescence markers (β-gal and 53BP1/p21). Values are mean ± SD of two independent experiments. Sample size (n) is indicated for f and i. NS, p > 0.05, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, and ****p ≤ 0.0001 by two-tailed χ2 statistical tests
