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. 2018 Sep 13;20(10):1045–1058. doi: 10.1016/j.neo.2018.08.008

Figure 2.

Figure 2

Depletion of PDZ-RhoGEF expression inhibits glioma cell migration and invasion and TROY-stimulated glioma cell migration. (A) U87MG cells were transfected with two independent siRNAs targeting PDZ-RhoGEF (PRG-1, PRG-2) or with a control siRNA targeting firefly luciferase (Luc). Cell lysates were analyzed for PDZ-RhoGEF by immunoblotting. Immunoblotting of β-tubulin protein was used as a loading control. (B) Invasion of U87MG cells transfected with siRNAs targeting PDZ-RhoGEF (PRG-1, PRG-2) or luciferase (Luc). Forty-eight hours after cell seeding onto the brain slice, glioma cell invasion into the brain slices was quantified using confocal microscopy. The data are depicted as the mean values (+/− SEM) from three separate experiments. ***P < .001. (C) Migration of T98G cells and T98G cells overexpressing TROY (T98G/TROY-HA) transfected with siRNAs targeting PDZ-RhoGEF (siPRG-1, siPRG-2) or a nonsilencing control (siCTRL). Data represent the mean values (+/− SEM) (n = 3, **, P < .01; ***, P < .001). Cell lysates were analyzed by immunoblotting with indicated antibodies. Immunoblotting of tubulin was used as a loading control. (D) Knockdown of PDZ-RhoGEF expression in patient xenograft GBM43 cells transduced with the shRNA-targeting PDZ-RhoGEF (PRG) or the control (CTL) nontargeting shRNA. Cell lysates were analyzed by immunoblotting with indicated antibodies. Immunoblotting of β-actin protein was used as a loading control. (E) Primary xenograft GBM43 cells transduced with a control nonsilencing shRNA (NS Ctrl) or shRNA targeting PDZ-RhoGEF (PRG) were treated with the indicated concentration of TMZ. Cell viability was assessed after 72 hours by CellTiter-Glo assay. Data are depicted as the mean values (+/− SD) of six replicates. ***P < .001.