A. Log2-gene expression ratios normalized to the mean expression in NB samples are shown for diffuse astrocytoma WHO grade II (AII), anaplastic astrocytoma WHO grade III (AAIII), secondary glioblastoma WHO grade IV (sGBIV) and primary glioblastoma WHO grade IV (pGBIV). Median RNA expression is indicated by horizontal bars; boxes show the 25th and 75th percentile range, whiskers mark the 5th and 95th percentiles; maximum and minimum values are depicted as horizontal bars.
B, C. The specificity of the antibody against Evi was confirmed by siRNA silencing of the target protein. U87MG cells were transfected with three-independent siRNAs to silence Evi. Silencing of gene expression was validated by Western blot and quantified real-time RT-PCR confirming robust downregulation of Evi expression. β-Actin was detected as loading control. PCR-data are expressed as mean ± SD of three-independent experiments (**p < 0.01).
D, E. Representative immunohistochemical stainings for Evi on tissue sections of NB and astrocytic gliomas of different WHO grades. (D) NB, Evi-positive vascular smooth muscle cells; (E) NB, Evi-positive ependymal cells.
F, G. Evi-positive tumour cells in a diffuse astrocytoma WHO grade II.
H, I. Evi-positive tumour cells in a primary glioblastoma WHO grade IV. Scale bar: 100 µm.
