Fig. 3

IFITM3 overexpression is central to cancer stemness, cancer progression of GC and pulmonary metastasis in vivo. A, E The efficiencies of IFITM3 overexpression in TMK-1 and AGS cells were determined by analyzing IFITM3 protein expression using western blots. B, F Measurements of cellular growth curve by viable cell counts from control or IFITM3-overexpressing TMK-1 and AGS cells, respectively. C, G Transwell migration assays were conducted using control or IFITM3-overexpressing and TMK-1 and AGS cells, respectively. Left panels: representative images of migrated cells from each treatement group. Right panel: quantitative and statistical analyses (**p < 0.01). D, H Sphere formation assays were utilized to assess impacts of IFITM3 overexpression on sphere-forming capabilities of TMK-1 and AGS cells (Scale bar, 500 μm). Left panels: representative spheres formed on TMK-1 or AGS1 cells. Right panels: quantitative and statistical analyses (**p < 0.01). I, J Chemoresistance of control- or IFITM3-overexpressing TMK-1 and AGS cells was determined based on percentages of surviving cells after treatment with 5’FU (20 μg/ml) or cisplatin (3 μM) for 72 h. Viable cells after treatments were measured using MTS assay. K Control or IFITM3-overexpessing AGS cells (2 × 10⁶) were subcutaneously injected into nude mice for monitoring tumor growth. Tumors from the tumor-bearing mice (n = 6 for each group) were collected as shown in the left panel image. Tumor growth curves were plotted and statistical differences between control and IFITM3-oeverexpression groups were derived using Student’s t-test (**p < 0.01). (L) SCID mice that received intravenous injection of 1 × 10⁶ control- or IFITM3-overexpressing AGS cells (n = 6 per group) were used to investigate if IFITM3 contributed to metastasis. All animals were sacrificed 4 weeks after cancer cell injections, and lungs from control or IFITM3-overexpresing groups were removed for tumor biopsy. Left panel: Representative H&E staining images from lungs of the control and IFITM3 groups of mice. Scale bar: 200 μm. Right panel represents the lung metastatic index (average fold increases in the density of tumor foci as per cm² of lung area)