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. 2016 Jul 19;7(34):54883–54896. doi: 10.18632/oncotarget.10702

Figure 3. FoxO3 knockdown slightly reduces the expression of stemness markers and strongly increases cell death after γIR or combination treatment with γIR and PI-103.

Figure 3

(A and D, left) Western blot analyses of the expression of FoxO proteins, stemness markers, and the differentiation marker GFAP in control shRNA- or FoxO3 shRNA-transduced GBM-SCs (n = 3 experiments; for statistical analysis, see Figure S4). (A and D, right) Absolute cell numbers after 3 or 4 days of incubation in CSC medium. (B and E, left) Confirmation of the transduction efficiency by GFP expression analysis. The bar represents 100 μm. (B and E, right) Sphere-forming capacity after 3 days of incubation in CSC medium. The bar represents 500 μm. (C and F) Assessment of cleaved caspase 3 (1 of 2 experiments is shown, each with similar results) and apoptosis by annexin V staining. Cells were treated with 0.5 μM PI-103 for 1 h and then irradiated with 10 Gy. The analyses were performed after 2 (GBM22) or 4 days (GBM4). Data for cell and sphere numbers and apoptosis in (A)–(F) represent means ± SD from 3 independent experiments. KD, knockdown.