Fig. 4. PVT1 KD suppresses the metastasis of HNSCC cells.

A, B Extreme limiting dilution analysis (ELDA) of EpCAM⁺ ALDH^high tumor cells transfected with shCtrl and shPVT1 in vivo (n = 5). The frequency of allograft formation was displayed for each cell dose injected. ELDA software was used to analyze the data. Scale bar: 1 cm. C Schematic diagram of the time points of ASO PVT1 treatment and sacrifice (Sac) of mice injected with ALDH^high CSC-like cells from HN6 cells. D Representative image of nude mice orthotopic tumors in the tongue from the mice in the different treatment groups. The lesion areas are circled with a white dotted line. Scale bar, 2 mm. E Quantification of the tumor volume of nude mice in the different treatment groups. Data are shown as the mean ± SD from at least two independent experiments. **p < 0.01 using an unpaired Student’s t-test. F H&E-stained HNSCC from nude mice in the different treatment groups. Scale bar, 500 μm. G Quantification of the number of tumors in the nude mice in the different treatment groups. Data are shown as the mean ± SD from at least two independent experiments. **p < 0.01 using a χ² test. H Quantification of the area of the tumors in the nude mice in the different treatment groups. Data are shown as the mean ± SD from at least two independent experiments. **p < 0.01 using an unpaired Student’s t-test. I A representative image of anti-PCK immunostaining of cervical lymph nodes. Scale bar, 200 μm. J Quantification of lymph node metastatic areas of nude mice in the indicated treatment groups. Data are shown as the mean ± SEM from at least two independent experiments. **p < 0.01 using an unpaired Student’s t-test.