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Fig. 2 | Journal of Experimental & Clinical Cancer Research

Fig. 2

From: TRIM56 promotes malignant progression of glioblastoma by stabilizing cIAP1 protein

Fig. 2

TRIM56 loss inhibits GBM progression in vitro. a Western blot showing TRIM56 protein levels in GBM#P3, LN229 and U118MG GBM cell lines stably transduced with lentiviruses containing constructs for shRNAs against TRIM56. b Growth curves generated from cell number counts taken over 72 h (at 24-h intervals) to generate growth curves of GBM#P3-, LN229- and U118MG-sh-TRIM56-NC, -S1 and -S2 glioma cells. c Fluorescence images and quantification of EdU assays performed to assess the proliferative capacity of GBM#P3-, LN229- and U118MG-sh-TRIM56-NC, -S1 and -S2 glioma cells (scale bar, 500 Î¼m). d Fluorescence images and quantification of immunofluorescence staining for cleaved caspase-3 in the indicated cells. Cells were fixed and nuclei were stained with DAPI (scale bar, 500 Î¼m). e Cell cycle analysis for the indicated cells. The percentage of cells arrested in the G0/G1 phase is analyzed in bar graphs (right panels). f Representative images and quantification of colony formation assays for the indicated cells. Cells were seeded at 1000 cells/well, cultured for 2 weeks, fixed and stained with crystal violet, and quantified. g Representative images of 3D tumor sphere invasion assays and quantification for GBM#P3-sh-TRIM56-NC, -S1 and -S2 cells. Representative images were acquired at 0 h, 24 h and 48 h, and plots show percentage of invaded area (scale bar, 200 Î¼m). h Model and representative fluorescence images of co-culture invasion assays of GBM#P3-sh-NC, sh-TRIM56-S1 or sh-TRIM56-S2. Invasion capacity was assessed at 24 h, 48 h and 72 h. Scale bars = 200 Î¼m (magnified insets) and the results were quantified. i Representative images and quantification of Transwell assays performed on LN229- and U118MG-sh-TRIM56-NC, -S1 and -S2 cells (scale bar, 50 μm). Cells were fixed, stained with crystal violet and counted. Comparisons between two independent samples and among multiple samples were performed using two-tailed t tests and one/two-way ANOVA, respectively. Error bars indicate at least three independent experiments, and data are shown as mean ± SEM. ∗ p < 0.05, ∗  ∗ p < 0.01, ∗  ∗  ∗ p < 0.001, and ∗  ∗  ∗  ∗ p < 0.0001

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