Fig. 3. U87 glioma xenograft slice culture characterization and drug responses.

a Retention of tissue integrity in U87 flank xenograft slice cultures by H&E staining, cell proliferation detected by Ki-67 immunostaining (red, DAPI counterstain) and cell death by SYTOX Green (SG)/Hoechst (blue) staining or by cleaved caspase 3 (CC3)/hematoxylin staining. b Xenograft slices contain endothelial (CD31), immune (CD45) and macrophage (IBA-1) cells. Confocal images identify U87 glioma cells (arrow, GFP⁺/HuNu⁺), vimentin-positive mesenchymal stromal cells (vim, red, solid arrowhead) and additional non-tumor cells (open arrowhead). c–f Slices treated for 2 or 3d and assessed for cell death by SG/Hoechst staining. Tanespimycin (AAG), geldanamycin, bortezomide (BORT, 3 d), mocetinostat (MOC, 3 d), parthenolide, and YM155 (YM). Cell death was quantified by mean fluorescence (dotted regions) c, d or by % SG⁺/total nuclei e, f. Red outline in c is a positive control crush lesion. g, h Apoptotic cell death quantified as CC3-stained area. No primary antibody control is shown. i Cell death in U87 cells by loss of CellTox Green fluorescence²⁸ when grown in conventional (blue) or slice (purple) medium, versus U87 xenograft slices (black line) in slice medium (SG index). For d, f, h, average ± SEM; n = 17, 3, 6, 5, 5, 3, 3, 4, 6, 6, 3, 3, 7, 6, 9, 4, 6, 8, 6, 9, 9 slices for each drug condition, in order. One-way ANOVA versus DMSO with Dunnett’s multiple comparison test. *p < 0.05, **p < 0.001. Cell line assays were done in duplicate. Scale bars: 100 μm a, g, 20 μm b, e, 1 mm c.