Figure 4

SOX1 reduces EMT, stimulates cell differentiation and triggers cellular senescence, whereas transient knockdown of SOX1 partially reverses the malignant phenotype. (A) Presence of the EMT-related proteins E-cadherin and Vimentin was detected via IF and WB. IF (left panel) Blue, DAPI; Red, E-cadherin; Green, Vimentin. EMT-related markers were detected by WB (right panel) in HONE1 cells with forced SOX1 expression. GAPDH served as an internal control. (B) Morphology of differentiated HONE1 cells induced by SOX1 was observed under microscopy. WB was used to detect the cell surface markers related to cell differentiation in HONE1 cells with or without SOX1 overexpression. (C) HONE1 cells with or without SOX1 overexpression were fixed and stained for SA-β-gal activity. The number of senescent colonies within each 20× field of HONE1 cells increased from 8.33 ± 1.53 to 50.33 ± 9.87. Bar represents mean ± SD of three independent experiments. (*p <0.05, Student’s t test) (D) SOX1 expression level in SOX1-overexpressing-NPC cells following transfection of SOX1-specific siRNA was determined using WB. (E, F) Ki67 staining rate and colony formation ability following knockdown of SOX1 using siRNA in SOX1-overexpressing NPC cells. Ki67 staining rate increased from 0.17 ± 0.02 to 0.49 ± 0.10 in CNE2 cells and from 0.14 ± 0.02 to 0.28 ± 0.06 in HONE1 cells. Colony formation ability increased from 6.82 ± 1.77% to 15.47 ± 5.28% in HONE1 cells. Bar represents mean ± SD of three independent experiments (*p <0.05, **p <0.01, Student’s t test). (Scale bars, 50 μm in A, 25 μm in B, C, 50 μm in E).