Micro-Vessels-Like 3D Scaffolds for Studying the Proton Radiobiology of Glioblastoma-Endothelial Cells Co-Culture Models

Glioblastoma (GBM) is a devastating cancer of the brain with an extremely poor prognosis. While X-ray radiotherapy and chemotherapy remain the current standard, proton beam therapy is an appealing alternative as protons can damage cancer cells while sparing the surrounding healthy tissue. However, the effects of protons on in vitro GBM models at the cellular level, especially when co-cultured with endothelial cells, the building blocks of brain micro-vessels, are still unexplored. In this work, novel 3D-engineered scaffolds inspired by the geometry of brain microvasculature are designed, where GBM cells cluster and proliferate. The architectures are fabricated by two-photon polymerization (2PP), pre-cultured with endothelial cells (HUVECs), and then cultured with a human GBM cell line (U251). The micro-vessel structures enable GBM in vivo-like morphologies, and the results show a higher DNA double-strand breakage in GBM monoculture samples when compared to the U251/HUVECs co-culture, with cells in 2D featuring a larger number of DNA damage foci when compared to cells in 3D. The discrepancy in terms of proton radiation response indicates a difference in the radioresistance of the GBM cells mediated by the presence of HUVECs and the possible induction of stemness features that contribute to radioresistance and improved DNA repair. Brain micro-vessels-like scaffolds are realized using two-photon polymerization (2PP). After co-culturing them with endothelial (HUVECs) and glioblastoma (GBM) cells, which show in-vivo-like morphologies, they are exposed to proton radiation. Higher DNA damage is observed in GBM monoculture when compared to GBM/HUVECs co-culture, with cells grown on 2D surfaces featuring a larger amount of DNA damage foci than cells in 3D.image