Fig. 2

Nodal protein levels are dowregulated during GBM differentiation. a, b OB1 stem cells forming oncospheres were immunostained for Nodal (red) and Nestin (green). DAPI (blue). OB1 stem cells present Nodal immunostaining localized to vesicles-like particles symmetrically distributed in the cytoplasm of GBM stem cells that are Nestin-negative (c, d). Upon differentiation, less differentiated cells were found positive for both Nodal and Nestin. More differentiated GBM cells were found negative for Nodal and positive for Nestin (69 % of cells; d, asterisk). Cells presenting a less elongated morphology were found Nodal-positive and Nestin-negative (26 % of cells). e Quantification of intracellular Nodal protein in OB1 stem cells and upon differentiation by Western blot. Nodal protein levels decrease in 50 % upon differentiation (quantification of average across three separate experiments). Nodal protein normalization through α-Tubulin immunoblotting. f Quantification of extracellular Nodal protein in protein precipitation of proteins present in conditioned medium by Western blot, presenting visible decrease in Nodal levels in OB1 cells upon differentiation. Nodal protein normalization through Coomassie Blue staining of gel. Data are mean ± SD. ***P < 0.001 by unpaired t test, n = 3)