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. Author manuscript; available in PMC: 2015 Jan 31.
Published in final edited form as: Mol Cancer Res. 2014 Aug 20;13(1):41–49. doi: 10.1158/1541-7786.MCR-14-0239

Figure 4.

Figure 4

SIRT1 silencing induces cell senescence through the p27kip1 pathway. A. SIRT1 knockdown induces cell senescence in NSCLC cells. 5×105 SIRT-silenced or shRNA-control H1299, H460 and A549 cells were seeded onto 6 well plates and cultured for 72 hrs, then SA-β-Gal analysis was performed. A representative experiment from three independent experiments is shown (left panels). The SA-β-Gal positive cells were quantitated under the microscope and the percentage of SA-β-Gal positive cells in each condition was calculated. The histograms represent the mean ± SD of triplicates (middle panels). Asterisks indicate significant differences between two groups (* p<0.01). Immunoblot analysis shows the SIRT1 levels in the H1299, H260 and A549 knockdown cells that were used for the SA-β-Gal analysis (right panels). B. p27kip1 knockdown rescues SIRT1-silencing-induced cell senescence. SIRT1-silenced H1299 cells were transfected with two independent p27 siRNA (#L-003472-00-005 & #12324) or non-targeting siRNA control (# D-001810-01-20) at 25nM for 72 hrs, then the SA-β-Gal analysis was performed. A representative experiment from three independent experiments is shown (the left panels). SA-β-Gal positive cells were quantitated under the microscope and the percentage of SA-β-Gal positive cells in each condition was calculated (the middle panel). Asterisks indicate significant differences between two groups (* p<0.01). The p27 levels were determined by immunoblot analysis with p27 antibody (the right panel).