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. Author manuscript; available in PMC: 2015 Oct 28.
Published in final edited form as: Nat Commun. 2015 Apr 28;6:6885. doi: 10.1038/ncomms7885

Figure 2. GALR2 promotes tumour invasion and metastasis.

Figure 2

(a) UM-SCC-1-GALR2 cells are more invasive than control UM-SCC-1-pcDNA cells in a Boyden chamber chemoinvasion assay with GAL as the chemoattractant (scale bar = 100 μm). Invasive cells (arrows) from both groups were quantified. Data are representative of three independent experiments with three replicates in each experiment. (*P < 0.05, two sample t-test; data represent mean + SD). (b) UM-SCC-1 cells were transfected with non target (NT) siRNA or siGAL (#5, #6, #7, #8). Downregulation of GAL was verified in RNA (upper panel) and whole cell lysates (lower panel) from these cells. (c) After verification of downregulation of GAL by Q-RT-PCR (left panel), conditioned medium from UM-SCC-1 (parent) cells transfected with NT siRNA or siGAL7 (siGAL) was used as the chemoattractant in a chemoinvasion assay performed with UM-SCC-1-GALR2 (right panel). Cells are significantly more invasive when stimulated with CM from UM-SCC-1-NT than from UM-SCC-1-siGAL7 cells. Data are representative of two experiments each with three replicates. (d-f) UM-SCC-1-GALR2 CAM tumours were (d) larger (scale bar = 5 mm), (e) more invasive (cancer cells are labelled green and highlighted by arrows, scale bar = 200μm), and (f) more disruptive of the basement membrane than control (UM-SCC-1-pcDNA) tumors. Collagen IV and dashed lines label basement membrane (scale bar =100μm). (g) Metastases (yellow, arrows) from the upper CAM to the lower CAM were observed (scale bar = 5 mm). Metastases to the lower CAM and liver were also quantified with quantitative ALU-PCR. (pcDNA group: lower CAM n=4; liver n=5; GALR2 group: lower CAM n=6, liver n=6).