Chemotherapeutic Stress Induces Transdifferentiation of Glioblastoma Cells to Endothelial Cells and Promotes Vascular Mimicry

Figure 3

TMZ increases population of immature and mature endothelial cells with functional endothelial cell properties. A Dil-Ac-LDL uptake assay was performed, where acetylated (Ac), fluorescently labeled low-density lipoprotein (LDL) specifically binds to a receptor on the surface of the endothelial cells delivering cholesterol via endocytosis. (a) FACS analysis was performed at day 2 and day 8 post-TMZ exposure to evaluate the uptake of Dil-Ac-LDL by the CD105- (a) and CD31- (b) positive cells. (c) The PDX line GBM43 was treated with 50 μM TMZ or equimolar DMSO and then plated in EGM (optimized endothelial cell media with 2% FBS and VEGF for rapid proliferation), EBM (EGM media without any supplement and growth factors), or DMEM media. Results show that EGM media independently promoted tube formation which was significantly augmented in the presence of TMZ. In addition, while DMEM media alone did not promote tube formation, the addition of therapeutic stress significantly increased tube formation. Images were taken and analyzed by a blinded experimenter. Bars represent the means of three independent experiments, and error bars show the standard error measure. Tubes were quantified based on the number of tubes per field with images taken across the plate of cells. The effect of various treatments was compared across cells by the Student -test with Tukey’s post hoc test.

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