Figure 4
From: Investigation of brain tissue infiltration by medulloblastoma cells in an ex vivo model
Quantification of relative growth and proliferation in response to HGF or EGF stimulation. (A) Confocal microscopy analysis of DAOY LA-EGFP cells spheroid co-cultured for five days in the OCSCs and exposed to the growth factors as indicated. EGFP signal was volumised and volume quantified. EdU-positive nuclei were marked and counted as well. (B) Quantification of volume, occupancy (tumour area/slice area) and number of EdU-positive nuclei. Mean and SD of two independent experiments are shown. ANOVA statistical analysis was performed (* = p < 0.05). (C) Higher magnification of EGF-stimulated DAOY cells invading the molecular layer. D) Purple asterisks in LA-EGFP grey scale panel highlight the proliferating cells. E) Magnification of the invasion front (boxed inset in (D). Inverted grey scale image shows F-actin cytoskeleton of leading edge cells with distinct filopodia and lamellipodia (arrows). (F) EGF-induced invasion of DAOY cells into molecular layer (green: LA-EGFP, red: anti-Calbindin). Orthogonal XYZ section shows infiltration of tumour cells into Purkinje cell layer. Red and green channels are also shown as inverted grey scale images for better visualization of the different layers. (G) Comparison of growth and invasion of untreated and EGF-stimulated UW228 tumour spheroids. Red arrows in LA-EGF grey-scale image indicate approximate length of invading cell streams. (H) Higher magnification of tissue infiltration of UW228 cells in response to EGF. Leader cells (green) at invasion front 600 µm deep in the host tissue and Purkinje cells (blue) are shown. Inverted grey scale image shows elongated F-actin cytoskeleton of the leader cell with pronounced lamellipodium. (I) Morphology of EGF-stimulated UW228 cells during spheroid-proximal infiltration. Image to the right shows volumised F-actin cytoskeleton with marked invasive protrusions.
