Human mesenchymal stromal cells increase glioblastoma radioresistance by cellular networks
OC-0595
Abstract
Human mesenchymal stromal cells increase glioblastoma radioresistance by cellular networks
Authors: Maren Strack1, Alexander Rühle1, Dieter Henrik Heiland2, Oliver Schnell2, Anca-Ligia Grosu1, Nils Henrik Nicolay1
1University Medical Center Freiburg, Department of Radiation Oncology, Freiburg, Germany; 2University Medical Center Freiburg, Department of Neurosurgery, Freiburg, Germany
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Purpose or Objective
Previous studies have identified the presence of multipotent mesenchymal stromal cells (MSCs) in the glioblastoma (GBM) microenvironment as an adverse feature regarding patient survival. We aimed to analyze whether MSCs contribute to the poor radiation response of glioblastoma cells (GBCs).
Material and Methods
An immortalized GBM cell line (U251) and two primary GBCs, isolated from surgical samples of a primary and a recurrent GBM, were directly co-cultured with human bone-marrow derived MSCs from three different donors to model intercellular interactions in vitro.
To examine the influence of MSCs on the radiosensitivity of GBCs, colony forming, proliferation and metabolic viability assays were performed. A modified Boyden chamber migration assay was carried out to investigate whether radiotherapy can mitigate the GBCs’ ability to chemotactically attract MSCs.
Cell cycle distribution and levels of cleaved caspase-3 were quantified by flow cytometry after irradiation with 2 und 4Gy, and the expression and localization of the radioresistance-related Growth Associated Protein 43 (GAP43) was analyzed based on immunofluorescence stainings. An siRNA-based knockdown was established in GBCs to evaluate whether GAP43-silencing may attenuate MSC-associated radioprotection.
Results
Direct co-culturing with MSCs enhanced the GBCs’ clonogenic survival and proliferation rates after irradiation, whereas the GBCs’ metabolic activity was unaffected by MSC-conditioned medium.
The number of MSCs migrating towards the tumor cells was found to be reduced after irradiation of the GBCs, indicating impaired secretion of chemoattractant cytokines by irradiated GBCs.
Direct co-cultivation with MSCs attenuated radiation-induced apoptosis and caused a pronounced G2/M arrest in primary GBCs after irradiation. Furthermore, the presence of MSCs increased GAP43 expression and led to a distinct pattern of GAP43 localization in primary GBCs. SiRNA-based knock-down of GAP43 resulted in a reduction of clonogenic survival of irradiated GBCs both in mono- and co-culture.
Conclusion
Bone-marrow derived MCSs increase the radioresistance of GBCs, potentially contributing to the tumor-protective effects of tumor-infiltrating MSCs in GBM patients. MSC-mediated up-regulation of GAP43 in GBCs is suggested as a potential molecular mechanism underlying the observed radioprotection of GBCs in the presence of MSCs.