Figure 10

Inhibition of TWIST1 expression decreases glioma cell invasion and stem cell activity. (A) Top panel: Endogenous levels of TWIST1 protein expression in nuclear extracts from SNB19 cells transduced with shGFP (shCtrl) or shTWIST1 (shTW). Levels of Ini1 protein were used as a loading control for Western blot analysis. Bottom panel: Quantification of SNB19 shTW invasion relative to control cells (shCtrl) accepted as 100%. Representative images of membranes demonstrating reduced invasiveness of SNB19 shTW cells through matrigel relative to control cells are shown. (B) Endogenous levels of TWIST1 protein expression in nuclear extracts from T98G cells transduced with shGFP (shCtrl) or shTWIST1 (shTW). Levels of Ini1 protein were used as a loading control for Western blot analysis. Bottom panel: Quantification of T98G shTW invasion relative to control cells (shCtrl) accepted as 100%. Representative images of membranes demonstrating reduced invasiveness of T98G shTW cells through matrigel relative to control cells are shown. (C) Inhibition of TWIST1 mRNA expression by 80% was achieved in GBM6 glioma stem cells transduced with shTWIST1 resulting in reduced sphere size. (D) Phase contrast image of representative GBM6 spheres transduced with control or TWIST1 specific shRNA lentivirus shows dramatic qualitative reduction in sphere size (E) Quantification of mean sphere diameter in GBM6 control or shTW cells confirms significant decrease in sphere size due to inhibition of TWIST1 expression (p < 0.0001).