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Lentivirus mediated silencing of the growth differentiation factor 15 gene increases chemotherapy resistance of glioma U373 cells
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Abstract:
Objective: To determine the effect of the growth differentiation factor 15 (GDF15) gene in resistance of glioma cells to chemotherapeutic agents cisplatin (DDP) and teniposide (Vumon, VM-26) in vitro. Methods: Glioma U373 cells were infected with a lentiviral vector expressing a small hairpin RNA targeting the GDF15 gene (LV-GDF15-RNAi) and a control lentiviral vector (LV-RNAi), respectively. Stably infected cells were then treated with VM-26 (01, 0.5, 2.5 and 12.5 μg/ml) and DDP (0.08, 0.4, 2 and 10 μg/ml). After treatment for 48 h, cell viability was assessed by MTT assay and Hoechst/PI staining to evaluate differences in resistance to VM-26 and DDP, and changes in Bcl-2, Bcl-xL, P53 and caspase-3 proteins were analyzed by Western blotting. Results: GDF15 protein content was significantly lower in LV-GDF15-RNAi-infected U373 cells (0.013±0.001) than in LV-RNAi-infected (0.622±0.068) and wild-type U373 cells (0.601±0.004) (P<0.01); survival rate was significantly different between LV-GDF15-RNAi-infected (91.84±2.64)% and LV-RNAi-infected (80.71±2.66)% in the presence of VM-26 and DDP at the lowest concentrations used (P<0.01) and the difference became more significant as the concentrations were increasing. Under the treatment with VM-26 or DDP at the same concentration, the number of apoptotic cells was significantly lower in LV-GDF15-RNAi-infected cells than that in LV-RNAi-infected cells (P<0.05). In association with changes in apoptosis, protein levels of Bcl-2 and Bcl-xL were significantly higher (P<0.05) but protein levels of P53 and caspase-3 were significantly lower (P<0.01) in LV-GFD 15-RNAi-infected cells than in LV-RNAi-infected cells. Conclusion: Down-regulation of the GDF15 gene may increase the resistance of glioma cells to VM-26 and DDP, possibly through regulating the expression of Bcl-2, Bcl-xL, P53and caspase-3.
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Project Supported:
Project supported by the Youth Foundation of the National Natural Science Foundation of China (No. 30701030)，the Project of Research Foundation in Application and Frontier Technology of Tianjin City (No. 11JCYBJC14900)，and the National High Technology Research and Development Program (863 Program) of China (No. 2012AA021003)
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