BACKGROUND: Glioblastomas are highly malignant tumours characterized by an invasive phenotype, which limits response to standard treatment including surgery, radiation and chemotherapy. Targeting the migratory properties of glioma cells may offer an alternative means to improve disease management. We recently developed a 96 well-based in vitro collagen invasion assay to study the effect of anti-migratory drugs on glioma spheroids generated from patient derived and established cell lines. One limitation of this 3D technique is the quantitative assessment of drug activity as data is subjected to 2D analysis. Here we explored the feasibility of analysing spheroids and migratory cells using 3D image reconstruction. METHODS: Glioma spheroids (U87) were allowed to migrate into a collagen matrix over 72h and prepared for immunohistochemistry. Paraffin embedded spheroids were serially sectioned at 5 µM. A total of 50 slides per spheroid were stained using H&E for reconstruction of the whole spheroid and surrounding migratory rim. Slides were scanned with Aperio AT scanner at 20x magnification, and custom image registration software was used to generate and view 3D volumes. This software uses a novel method for slice-to-slice image registration using automatic registration algorithms custom designed for virtual slides and histopathology images RESULTS: By visualizing the fixed spheroids in 3D, we determined the number, morphology and distance traversed by migratory cells. We also ascertained differences between core and migrating cell morphology, number of cells and voxel sizes. CONCLUSIONS: We have demonstrated for the first time the feasibility of generating 3D histological reconstruction of glioma spheroids and migratory cells encased in collagen including whole slide scanning, image serving, registration and visualization in one procedure. Analysis of the glioma spheroids and the migratory cells allowed the quantitative analysis of research-relevant parameters. This technology can be used to analyse migratory behavior of glioma cells and drug activity in a 3D setting.
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|Published - Nov 2014
|19th Annual Scientific Meeting of the Society for Neuro-Oncology - Loews Hotel South Beach, Miami, United States
Duration: 13 Nov 2014 → 16 Nov 2014
Conference number: 19