Effects of Smad3 overexpression on the inhibitory effect of Nar on TGF-β1-mediated migration.

<p>(A) Transwell migration assay, in non-Matrigel-coated cell culture insert, panc-1 cells were transfected with GFP-Smad3 or GFP-C1 plasmid for 6 h, then washed, changed to complete DMEM with 0.1% DMSO, 100 µM Nar for 24 h, then exposure to 5 ng/ml TGF-β1 or the combination of 5 ng/ml TGF-β1 and 100 µM Nar for next 24 h. As described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0050956#s4" target="_blank">Materials and methods</a>. The cells that had invaded to the lower surface of the filter were stained with crystal violet and photographed. (B) Transwell migration assays. In non-Matrigel-coated cell culture insert, to quantify migration, the stained cells were lysed in DMSO and their absorbance was measured. These experiments were done triplicate. *p<0.05; **p<0.01. ***p<0.005. (C) and (D) Panc-1 cells were transfected with GFP-Smad3 or GFP-C1 plasmid for 6 h, then washed, changed to complete medium, 100 µM Nar for 24 h, then exposure to 5 ng/ml TGF-β1 or the combination of 5 ng/ml TGF-β1 and 100 µM Nar for next 24 h for qRT-PCR (C) or western blot (D).