Glioblastoma multiforme (GBM), is the most common and the most aggressive type of primary brain malignancy. Glioblastoma stem-like cells (GSCs) are able to migrate in vascular niches within or away from the tumour mass, increasing tumour resistance to patient treatments and contributing to relapses. To study individual GSCs migration and their interactions with the microenvironment in the vasculature, there is a need to develop a model of human blood vessels in vitro. Herein, we report a systematic study on the interaction between patient-derived glioma stem-like cell lines with different organotypic perivascular niche models. A microfluidic chip integrated with an extracellular matrix was fabricated to support the culture of rounded microvessels, formed with endothelial cells from three different organs, (1) human brain microvascular endothelial cells (hCMEC/D3), (2) human umbilical vein endothelial cells (HUVECs) and, (3) human lung microvascular endothelial cells (HMVEC-L). Three-dimensional (3D) cell culture retains selected adherent and tight junction markers of the endothelial cells, and the stemness-related genes of GSCs. We optimized the experimental protocol to perform qPCR, and western blot on the co-cultured GSCs with endothelial cells forming microvessels. Endpoint biological assays showed upregulation of neovascularization-related genes in endothelial cells (e.g., angiopoietins, vascular endothelial growth factor receptors) resulted after their co-culture with GBM cells. Moreover, we measured cancer cell speed and polarization during migration towards the endothelial cell formed vessel by live-cell imaging showing that organotypic (brain cancer cells - brain endothelial microvessel) interactions differ from those within non-tissue specific vascular niches. The development and optimization of this 3D microfluidic device could provide the next level of complexity of an in vitro system to study the influence of glioma cells on normal brain endothelium. More importantly, it enables the possibility to conduct comparative studies to dissect the influence of 3D culture, microvessel architecture and organotypic vessel types on glioma cells stemness and migration.
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