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Updated: Jun 7, 2025
Co-culture of Glioblastoma Stem-like Cells on Patterned Neurons to Study Migration and Cellular Interactions
Published on: February 24, 2021
Glioblastoma cells alter brain endothelial cell homeostasis and tight junction protein expression in vitro
Xolisile Mokoena1, Peace Mabeta2, Werner Cordier1
1Department of Pharmacology, School of Medicine, Faculty of Health Sciences, University of Pretoria, Dr Savage Road, Prinshof 349-Jr, Private Bag X323, Arcadia, Pretoria, 0007, South Africa.
Background:
Glioblastoma (GBM) is an aggressive therapy-resistant brain tumour that may impacts the integrity of the blood-brain barrier (BBB). The BBB is a protective barrier of the central nervous system formed mainly by endothelial cells. This study aimed to investigate the in vitro effect of GBM cells on the BBB.
Methods:
Brain endothelial (bEnd.3) cells were used as a model of the BBB. Glioblastoma-conditioned media (CM) was extracted at the 48-h (h) time-point from the U87 GBM cells and diluted to 40% with fresh media. The effect of the U87-CM collected at 48 h on bEnd.3 cell growth was evaluated following 48 and 72 h of treatment using the xCELLigence system. Additionally, bEnd.3 cell growth was also investigated in a U87 and bEnd.3 co-culture model continuously for 48 h using the xCELLigence system. The migration of bEnd.3 cells was assessed following 48 and 72 h using the migration scratch assay. The barrier integrity was evaluated continuously for 1 h using the transwell permeability, and the tight junction (TJ) protein expression was evaluated using Western blot assay following 48 and 72 h.
Results:
There was a significant decrease in bEnd.3 cell growth following 32 h (p < 0.05), 40 h (p < 0.01), and 48 h (p < 0.001) of treatment with U87-CM, while co-culturing of bEnd.3 and U87 cells increased cell growth following 16 h (p < 0.05), 24 h (p < 0.001), 32 h (p < 0.01), 40 h (p < 0.001), and 48 h (p < 0.001). The migration of bEnd.3 cells significantly increased following both 24 (p < 0.05) and 48 h (p < 0.01) of treatment with U87-CM. The permeability of bEnd.3 cells co-cultured with U87 for 48 h was significantly increased (p < 0.05) at the 15- and 30-min time points. Furthermore, the expression of ZO-1 and occludin was significantly increased (p < 0.05) in both bEnd.3 cells treated with U87-CM as well as bEnd.3 cells co-cultured with U87 cells.
Conclusion:
The current findings suggest that U87 cells alter the integrity of bEnd.3 cells possibly through the secretomes in the CM and through cell-cell interactions in co-culture models. This may assist in the understanding of the mechanisms by which GBM affects the BBB, which may aid in the management thereof.
