Fig. 2.

Characteristics of DIPS-associated growth arrest according to cell distribution in the cell cycle and the analysis of cell cycle inhibitory proteins and cell cycle-promoting cyclins. Analysis of cell distribution in particular phases of mitotic cycle using flow cytometry with cells stained with propidium iodide. The cells gathered in the G1 phase are marked in red, whereas cells belonging to the G2/M stage are marked in blue. The quantification (%) of cells arrested in G0/G1, S, and G2/M phases is shown on the representative histograms (a). Determination of senescence-specific cell cycle inhibitors (p16, p21, and p53) in young and DIPS cells. White arrows indicate exemplary positive cells. Magnification × 200, objective aperture 0.35 (b). Changes in cyclin B1 and D1 level during DIPS (c) quantified with densitometry (d). Because the cytostatic effect of drugs is diverse for individual samples, the blots represent results obtained for all tested cultures (Y—young cells; S—senescent/DIPS cells). Results derive from 6–8 independent experiments with HGSOCs obtained from different donors. Flow cytometry was performed using 10 000 cells, whereas the quantification of cell cycle inhibitors was based on the analysis of 500 randomly selected cells per group. The results are expressed as the means ± SEM. *p < 0.05; **p < 0.01; ***p < 0.001 vs. young cells