Fig. 2.

Nestin expression is required for the stability of lamin A/C and evasion of senescence in cancer cells. a qPCR analysis of the expression of lamins in control and Nestin-knockdown cells. b Immunoblotting analysis of lamins expression in control and Nestin-knockdown cells. c Immunofluorescence staining of lamin A/C in control and Nestin-knockdown cells. Scale bars, 5 μm. d Immunofluorescence staining of lamin A/C in control, Nestin-knockout, and Nestin-knockout/lamin A/C-rescue H1299 cells. Scale bars, 20 μm. e Immunoblotting analysis of lamin A/C in control, Nestin-knockout, and Nestin-knockout/lamin A/C-rescue cells. f SA-β-gal staining was used to identify the frequency of senescent cells in vitro (left panel, ×400). Scale bars, 100 μm. g Quantification of the SA-β-gal-positive cells in f. h Immunofluorescence staining of H3K9me3 in control, Nestin-knockout, and Nestin-knockout/lamin A/C-rescue H1299 cells. Scale bars, 20 μm. i Nuclear circularity was analyzed with the Cellprofiler software. Shown were the quantifications of mean circularity and the percentage of cells with deformed nuclei in cells subjected to Nestin knockdown and lamin A/C overexpression. For all quantifications, the data represented the mean ± SEM of three independent experiments, as assessed using unpaired t-test. *P < 0.05, **P < 0.01, ***P < 0.001