Fig. 2

Gemcitabine resistance induced by PVT1 was associated with increased Wnt/β-catenin signaling pathway and autophagic activity. a Whole cell lysates from PANC-1 and ASPC-1 cells transfected with PVT1 overexpression plasmid or PVT1 siRNA were analyzed by western blotting using the indicated antibodies. b The expression levels of CyclinD1 and C-myc in PANC-1 and ASPC-1 cells transfected with PVT1 overexpression plasmid or PVT1 siRNA were analyzed by real-time qRT-PCR. c and d Representative confocal images of GFP-LC3 puncta in PANC-1 cells transfected with PVT1 siRNA. The number of GFP-LC3 puncta was quantified using ImageJ software. (n = 10). Scale bars: 10 μm. e Representative H&E staining images and immunohistochemical images of CyclinD1, C-myc and p62 in excised xenograft tumor tissues. Scale bars: 100 μm. f PANC-1 cells stably expressing PVT1 were treated with gemcitabine (1 μM), XAV-939 (10 μM) or CQ (10 μM) for different durations as indicated, and cell viability was measured by MTT assay. g PANC-1 cells stably expressing PVT1 were transfected with β-catenin siRNA or ATG7 siRNA, and the cells were then treated with gemcitabine (1 μM) for different durations as indicated. Cell viability was measured by MTT assay. h and i Apoptotic cells among PVT1 overexpressed PANC-1 cells treated with gemcitabine (1 μM), XAV-939 (10 μM) or CQ (10 μM) were analyzed by TUNEL assay, and the number of TUNEL-positive cells was quantified. Scale bars: 100 μm. j and k Representative images and quantification of EdU incorporation in PVT1 overexpressing PANC-1 cells treated with gemcitabine (1 μM), XAV-939 (10 μM) or CQ (10 μM). Scale bars: 100 μm. Data were represented as mean ± SD, *P < 0.05; **P < 0.01; ***P < 0.001