Expression of oncogenic microRNA-21 in neurospheres and attached cells of a glioblastoma cell line increased after treatment with temozolomide and ionizing radiation
Glioblastomas are the most common primary malignant brain tumors, and despite current advances in therapy, they are still extremely resistant to treatment. The tumorigenic potential of glioblastomas is due to a subpopulation of neoplastic cells, called cancer stem cells (CSCs), which in culture form a suspension cluster called neurospheres. Their properties include tumorigenesis, chemoresistance, and radioresistance. Several miRNAs exhibit altered expression levels during gliomagenesis; thus they constitute one of the key mechanisms for CSC regulation. We evaluated whether temozolomide and ionizing radiation modulate the expression of the tumor suppressor microRNAs-15, -16 and of the oncogenic miR-21 in neurospheres and attached cells in a glioblastoma cell line culture (U343-MG). The methods included staining with Trypan blue to verify cell viability and real time PCR to quantify the expression of microRNAs after exposure to treatments. We observed that miR-16 was more highly expressed in neurospheres than in attached cells 48h after treatment with temozolomide, and miR-21 was highly expressed in attached cells compared to neurospheres treated with temozolomide at time 0 h (30 min after treatment); at 48 h this microRNA was highly expressed in neurospheres treated with temozolomide in association with ionizing radiation. We concluded that the microRNAs were differentially expressed when comparing the different cell types and treatment modalities. The higher expression of miR-21 in neurospheres at time 48 h may suggest a pathway of CSC radioresistance and chemoresistance, with further rapid tumor growth and recurrence or resistance to other treatment modalities.