Glioblastoma (GBM) is considered the most common and lethal type of brain tumor with a poor prognosis. GBM treatment has challenges due to its aggressive nature, which often causes treatment failure and recurrence. Hypoxia is one of the characteristics of glioblastoma tumors that contribute to radioresistance and malignant phenotypes of GBM. In this study, we aimed to determine the effects of hypoxia on the radiosensitivity of U87 GBM cells by the hypoxia-mimicking model.
Following the treatment of cells with different concentrations of CoCl, an MTT assay was used to evaluate the cytotoxicity of CoCl. To understand the effects of Ionizing radiation on CoCl-treated groups, cells were exposed to irradiation after pretreating with 100 μM CoCl and a clonogenic survival assay was performed to determine the radiosensitivity of U87 cells. Also, the intracellular Reactive oxygen level was measured by 2′,7′-dichlorofluorescein diacetate (DCFDA) probe staining. Additionally, the expression of hypoxia-associated genes, including HIF-1α, HIF-2α, and their target genes (GLUT-1), was monitored by reverse transcription polymerase chain reaction (RT-PCR).
Our study revealed that the cell viability of CoCl-treated cells was decreased in a concentration-dependent manner. Also, CoCl did not cause any cytotoxicity on U87 cells at a concentration of 100 μM after treatment for 24 h. Colony formation assay showed that CoCl pretreatment induced radioresistance of tumor cells compared to non-treated cells. Also, CoCl can protect cells against irradiation by the clearance of ROS. Moreover, Real-time results showed that the mRNA expression of HIF-1α and GLUT-1 were significantly upregulated following hypoxia induction and/or irradiation condition. However, the level of HIF-2α mRNA did not change significantly in hypoxia or irradiation alone conditions, but it increased significantly only in hypoxia + irradiation conditions.
Taken together, our results indicated that simulating hypoxia by CoCl can effectively increase hypoxia-associated genes, specially HIF-1α and GLUT-1, but did not affect HIF-2α gene expression. Also, it can increase the clearance of ROS, respectively, and it leads to inducing radioresistance of U87 cells.

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