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. 2012 Apr 25;3:59. doi: 10.3389/fendo.2012.00059

Figure 2.

Figure 2

Effect of Nodal overexpression and knockdown on glioblastoma cell proliferation. (A) Four cell lines, U87 transfected with a control empty vector (U87–EV), U87 transfected with Nodal shRNA (U87-shNodal), U87 transfected with Nodal plasmid (U87-Nodal), and 87 transfected with both Nodal expression plasmid and Nodal shRNA (U87-Nodal/shNodal) were generated. Proteins extracted from these cell lines were analyzed by Western blotting using antibodies against Nodal, phosphor-Smad2 (pSmad2), total Smad2 and Smad3 (Smad2/3), and β-actin as the loading control. Overexpression and activation of Smad2 were confirmed in the U87-Nodal cells. Numbers on the Nodal blot are densitometry readings of the bands. (B) Cell growth assays. The stable cell lines were cultured for 48 h and cell numbers were determined by the trypan blue exclusion method. Data represents mean ± SEM (n = 3). The experiment was done three times and similar results were obtained. *P < 0.01 vs. U87–EV and U87-shNodal, #P < 0.05 vs. U87–EV and U87-Nodal. (C) Comparison of the effect of Nodal overexpression in U87 and OV2008 cells. Two pairs of cell lines, control and Nodal-overexpressing OV2008 and control and Nodal-overexpressing U87 cells, were cultured for 2 days before pictures were taken.