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. 2016 Apr 25;7(23):34630–34642. doi: 10.18632/oncotarget.8969

Figure 1. S100A4 drives the invasive potential of lung cancer cells.

Figure 1

A and B. Lung cancer cells as indicated were lysed in RIPA buffer, total cell lysates (80μg) were subjected to 15% SDS-PAGE, transferred and immunoblotted with rabbit anti-S100A4 antibody (A). RNA was isolated from cells and quantitative real time PCR (Q-PCR) was used to assess S100A4 expression levels (B). C-G. A549 cells with stable knockdown of S100A4 by shRNA targeting S100A4 (shS100A4) or expressing a non-targeting control (shCont) were generated. S100A4 expression was assessed by immunoblot analysis (C) and Q-PCR (D). Cell proliferation in standard (2D) culture was assessed by MTT (E). Cells were grown in 3D Matrigel for 5 days and representative phase contrast images for control and knockdown cells are shown (F). The diameter of 70-120 colonies from randomly chosen fields was measured, quantified for average colony volume and presented in (G). H-J. H1299 cells, stably transfected with pIRES-GFP-S100A4 (GFP-S100A4) or pIRES-GFP alone (GFP), were assessed for S100A4 expression by immunoblot analysis (H) or for invasion toward 1% FBS overnight (I) or grown in 3D Matrigel for 5 days (J). Representative data from at least three independent experiments are shown. Error bars represent the SEM of the mean in (G) and the SD of the mean from at least three replicates in (B, D, E and I). Arrows indicate invasive growth. Scale bar in (F and J) = 50μm. * indicates p<0.05.