Figure 4.

STAT3 signaling is critical for SC-induced pancreatic cancer cell migration and invasion. A. Immunoblotting assay demonstrating increased STAT3 phosphorylation in pancreatic cancer cells following exposure to co-cultured conditioned medium (co-CM) for the indicated times. Cells without co-CM treatment were used as a control (Ctrl). B. Western blotting of pancreatic cancer cells shows reduced level of phosphorylated STAT3 (p-STAT3) following the addition of IL6 neutralizing antibodies (50 ng/mL) to co-CM compared with those only treated with co-CM. The duration of co-CM treatment in both groups was 30 min. Cells with no treatment were used as a control. C. Representative images of the immunofluorescence assay results for anti-p-STAT3 (red) and DAPI (blue) staining. Pancreatic cancer cells were treated with co-CM or co-CM plus IL6 neutralizing antibodies (50 ng/mL) for 30 min and then collected for immunofluorescence analysis. Cells without any treatment were used as a control. Scale bar: 10 μm. D. Immunoblotting analysis showing disrupted STAT3 activation following co-CM incubation at the indicated times in STAT3 siRNA#1 targeted pancreatic cancer cells. Scrambled siRNA targeted cancer cells exposed to co‑CM were used as a control. E and F. Representative images of wound healing assay showing that STAT3 interference undermined co-CM induced pancreatic cancer cell migration. Control siRNA targeted tumor cells incubated with co-CM were used as a control. G and H. Scrambled siRNA and STAT3 siRNA- targeted MIA PaCa-2 and AsPC-1 cells were used for Transwell migration and invasion assays. Co-CM was used as a chemoattractant in all groups. Scale bar: 100 μm. *p < 0.05, **p < 0.01, ***p < 0.001.