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. 2018 May 16;6(2):181–198. doi: 10.1016/j.jcmgh.2018.05.002

Figure 4.

Figure 4

E47 altered expression of cellular senescence biomarkers in a prosenescence direction. (A) Cell size was measured after staining cell membrane glycoproteins with wheat germ agglutinin conjugated to Alexa 488. The median cell size was quantified based on pixel number, P values were determined by Mann–Whitney test, boxes represent the 2 middle quartiles, and whisker ends represent minimum and maximum data points. More than 50 cells were measured in each condition. (B) SA-βgal activity was assayed at pH 6 after 72 hours of culture. For each condition in each cell line, data represent the mean of 3 separate fields, each with a minimum of 50 cells counted and compared by unpaired t test. Error bars are ± SEM. Similar data were observed in 3 independent experiments. (C) Western blot analysis of CEBP-α, Lamin B1, and CENP-A after induction of E47. (B and C) Scale bars: 62.5 μm; magnification, 200×. (D) Western blot analysis of the effect of E47 induction on ERK expression and activation via phosphorylated ERK (pERK) T202/Y204. pERK and ERK signals were normalized to vinculin protein loading control. The ratio of pERK to ERK after normalization is shown. (E) Western blot analysis of whole-cell extracts from MIA PaCa-2/E47 and PANC-1/E47 cells after 72 hours of treatment with 5 μm SCH772984 ERK inhibitor. (F) SA-βgal activity assayed in PANC-1/E47 cells after 72 hours of E47 induction and simultaneous treatment with 5 μm SCH772984. Data represent the mean of 6 separate fields, each with a minimum of 100 cells analyzed by unpaired t test. Error bars are ± SEM.